scientific report 2003-2004 - ipb halle · jörg ziegler publications and other scientific...

SCIENTIFIC REPORT 2003-2004

Leibniz Institute of Plant BiochemistryWeinberg 306120 HalleGermany

Phone: +49 (0) 3 45 - 55 82 11 10Fax: +49 (0) 3 45 - 55 82 11 09

Email: [email protected]

Message from the Managing Director 4

Departmental Organization 5

Research Mission Statement 6

Department of Natural Product Biotechnology 8Head: Toni M. Kutchan, Professor

Research Groups: Alkaloid Biosynthesis 10Toni M. Kutchan

Opium Poppy Biotechnology 12Susanne Frick

Mode of Action of Jasmonates 14Claus Wasternack & Otto Miersch

Papaver-Gene Expression Analysis 16Jörg Ziegler

Publications and Other Scientific Activities 2003 18


Department of Bioorganic Chemistry 20Head: Ludger Wessjohann, Professor

Research Groups:Plant and Fungal Metabolites/Microanalytics 22Norbert Arnold & Jürgen Schmidt

Structural Analysis & Computational Chemistry 24Wolfgang Brandt & Andrea Porzel

Research Group: Isoprenoids 26Ludger Wessjohann

Research Group: Synthesis and Method Development 28Ludger Wessjohann & Bernhard Westermann



Department of Stress and Developmental Biology 36Head: Dierk Scheel, Professor

Research Groups:Molecular Communication in Plant-Pathogen Interactions 38Wolfgang Knogge

Cellular Signaling 40Dierk Scheel & Justin Lee

Induced Pathogen Defense 42Dierk Scheel & Sabine Rosahl

Metal Homeostasis 44Stephan Clemens

Metabolite Profiling in Arabidopsis and Crop Plants 46Stephan Clemens, Jürgen Schmidt, Ludger Wessjohann, Dierk Scheel



Table of Contents

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Department of Secondary Metabolism 50Head: Dieter Strack, Professor

Research Groups: Molecular Physiology of Mycorrhiza 52Michael H. Walter

Cell Biology of Mycorrhiza 54Bettina Hause

Biochemistry of Mycorrhiza 56Willibald Schliemann

Glycosyl- and Methyltransferases 58Thomas Vogt

Hydroxycinnamic Acids 60Dieter Strack



Administration and Technical Services 64Head: Lothar Franzen

Administration Employees 66

Employee Statistics 67

Involvement of the IPB in National and International Scientific Networks 68

Resources and Investments 69

Funds from External Sources 70

Seminars, Workshops and Colloquia 2003 74

Seminars, Workshops and Colloquia 2004 76

Guest Researchers and Fellows 78

Press and Public Relations 80Head: Sylvia Pieplow

Publications and Press Releases 2003 84

Publications and Press Releases 2004 85

Map and Impressum 87

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The Leibniz Institute of Plant Biochemistry (IPB) is a research center that isrich in contemporary German history. Founded as the Institute for Bio che -

mistry of Plants in 1958 by Kurt Mothes, this former research institute of the EastGerman Academy of Sciences was unique in emphasizing a multidisciplinary ap -proach to studying plant natural products and hormone-regulated processes.Scientists from around the globe were regularly invited to visit and the instituteenjoyed international recognition. The IPB, as we know the institute today, wasfounded in 1992 and has developed into a thriving center of modern plant re -search. The multidisciplinary style of research introduced by Kurt Mothes conti-nues and has been expanded upon.Within four research departments, more than170 scientific staff, doctoral and diploma students and guests from the world-overcontribute to the exciting bustle of state-of-the-art research.

The fundamental research programs at the IPB center on, in a broad sense, natu-ral product research and molecular interactions. From natural product isolationand structure determination through to investigations of biosynthesis and regu-lation, new biologically active molecules are sought and production sources opti-mized through chemical synthesis and genetic engineering. Cellular func tion asregulated by molecular interactions are investigated at the level of receptor-ligand, enzyme-substrate and protein-protein interactions using computer mode-ling and a variety of biochemical and genetic techniques. The years 2003-2004were very productive for us and intriguing new research developments within thedepartments at the IPB in these areas are highlighted with in this scientific report.

Scientists at the institute are participating in an increasing number of local, natio-nal and international research networks. A new Priority Program of the GermanResearch Foundation (DFG, SPP 1152, Evolution of Metabolic Diversity) was ini -tia ted and is coordinated by scientists at the IPB. In 2004, members of the insti -tute contributed to the successful application to the German Research Foun -dation for a new Collaborative Research Center (SFB 648, Molecular mechanismsof in for mation processing in plants) at the Martin Luther University of Halle-Wit -ten berg.

We have further improved upon our research facilities during the past two years;the institute dedicated a new 500 square meter shared-use building in Spring2004. The construction provides space for a mass spectrometry center, nucleo -tide sequence analysis, fermentation and synthetic organic chemistry laborato-ries. Our greenhouses are also being expanded; we began in November 2004with the construction of an additional 346 square meters of floor space that willmeet the growing demand of the IPB's research programs with transgenic plants.These new plant growth facilities will be ready for occupancy in Spring 2005.

We hope that in this report, we are able to share our excitement for plant bio-chemistry and its manifold applications with you.We are proud of how our insti-tute has developed over the years and if you are planning on being in the Halle(Saale) area, please do stop by to learn more about us.

Toni M. Kutchan

Message from the Managing Director

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Board of Directors

Dierk Scheel, ProfessorManaging Director until November 2004Head of the Department of Stress andDevelopmental Biology

Toni M. Kutchan, ProfessorManaging Director since November 2004Head of the Department of Natural ProductBiotechnology

Lothar FranzenHead of Administration and TechnicalServices

Dieter Strack, ProfessorHead of the Department of SecondaryMetabolism

Ludger Wessjohann, ProfessorHead of the Department of BioorganicChemistry

Board of Trustees

Ministerialrat Thomas ReitmannChairman of the Board of Trustees,Ministry of Education and Cultural Affairs ofthe State of Saxony Anhalt

MinisterialratJürgen Roemer - Mähler, Dr.Vice Chairman of the Board of Trustees,Federal Ministry of Education and Research

Wilhelm Boland, ProfessorMax Planck Institute for Chemical Ecology,Jena, Chairman of Scientific Advisory Board

Alfons Gierl, ProfessorTechnical University of Munich,Vice Chairman of Scientific Advisory Board

Reinhard Neubert, ProfessorVice Rector for Research and PostgraduateStudents of the University of Halle

Jörg Stetter, ProfessorBayer AG, Leverkusen

Scientific Advisory Board

Wilhelm Boland, ProfessorChairman of Scientific Advisory Board,Max Planck Institute for Chemical Ecology,Jena

Alfons Gierl, ProfessorVice Chairman of Scientific Advisory Board, Technical University of Munich, Department of Genetics

Raoul J. Bino, ProfessorUniversity of Wageningen, The NetherlandsLaboratory of Plant Physiology

Thomas Boller, ProfessorUniversity of Basel, Institute of Botany

Horst Kunz, ProfessorUniversity of Mainz, Institute of Organic Chemistry

Birger Lindberg MØller, ProfessoerUniversity of Copenhagen, Department of Plant Biology

Günter Strittmatter, Dr.KWS SAAT AG, Einbeck

Lutz F. Tietze, ProfessorUniversity of Göttingen, Institute of Organic Chemistry

Lothar Willmitzer, ProfessorMax Planck Institute of Molecular Plant Physiology, Potsdam-Golm

Ulrich Wobus, ProfessorInstitute of Plant Geneticsand Crop Plant Research, Gatersleben

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Departmental Organization

Department ofNatural ProductBiotechnologyHead: T. M. Kutchan,

Department ofBioorganic ChemistryHead: L. Wessjohann,Professor

Department ofStress and DevelopmentalBiologyHead: D. Scheel, Professor

Department ofSecondary MetabolismHead: D. Strack, Professor

Administration and TechnicalServicesHead: L. Franzen

Alkaloid BiosynthesisToni M. Kutchan

Plant and Fungal Meta -bolites /MicroanalyticsNorbert Arnold & Jürgen Schmidt

Structural Analysis &Computational ChemistryWolfgang Brandt & Andrea Porzel

Molecular Commu ni- cation in Plant-PathogenInteractionsWolfgang Knogge

MolecularPhysiology of MycorrhizaMichael H. Walter

AccountingHuman ResourcesGeneral Administration

Library ServicesStockroomGraphic Arts

Facilities ManagementComputer SupportHorticulture

Hydroxycinnamic AcidsDieter Strack

Cell Biology of MycorrhizaBettina Hause

Biochemistryof MycorrhizaWillibald Schliemann

Glycosyltransferases &MethyltransferasesThomas Vogt

Cellular SignalingDierk Scheel & Justin Lee

Induced Pathogen DefenseDierk Scheel & SabineRosahl

Metal HomeostasisStephan Clemens

Bioinformatics & Mass SpectrometryDierk Scheel

IsoprenoidsLudger Wessjohann

Synthesis and Method DevelopmentLudger Wessjohann &Bernhard Westermann

Opium PoppyBiotechnologySusanne Frick

Mode of Action of JasmonatesClaus Wasternack & Otto Miersch

Papaver Gene ExpressionAnalysisJörg Ziegler

Metabolite Profiling in Arabidopsis and Crop PlantsPart of the German Plant Genome ProjectStephan Clemens, Jürgen Schmidt, Ludger Wessjohann & DierkScheel

Board of TrusteesMinisterialrat T. Reitmann

Chairman

Ministerialrat J. Roemer-Mähler, Dr.Vice Chairman

Scientific Advisory BoardW. Boland, Professor

Chairman

A. Gierl, ProfessorVice ChairmanBoard of Directors

D. Scheel, ProfessorManaging Director until 31.10.2004

T. M. Kutchan, ProfessorManaging Director since 01.11.2004

L. FranzenAdministratitive DirectorD. Strack, Professor

L. Wessjohann, Professor

Public RelationsS. Pieplow

Personal Assistent to the Managing Director

Four thematically, methodologically and organisationally overlapping re search pri-orities form the basis of the research mission statement of the Institute of Plant

Biochemistry - plant natural products, molecular in teractions, information technolo-gy and metabolic engineering.

The large manifold of plant species is reflected in the enormous diversity of their nat-ural products. This content of natural compounds is made more complex by thechange in metabolite patterns during development as well as when a plant isresponding to its environment. Knowledge of the structure and function of naturalproducts is requisite to understanding plant diversity, developmental and adaptationprocesses. New resources can then become available for innovative application inplant production, plant protection, biotechnology and in the development of biolog-ically active compounds. Furthermore, the realization of genome sequencing and thegrowing availability of expressed sequence tags of various species is of fundamentalimportance to functional genome analysis.

The comprehensive analysis of plant and fungal natural products is a priority in theresearch mission of the Institute of Plant Biochemistry. Struc ture analysis, synthesisand derivatization of natural products contribute to an understanding of their func-tion and to an increase in their structural di versity. This also forms the basis forinvestigation of their biosynthesis and for discovering new biologically active com-pounds. A qualitative and quantitative analysis of natural products in biological mate-rials requires the de velopment of suitable analytical methods. Subsequent identifica-tion and isolation of biosynthetic enzymes can provide access to the encoding genes,which in turn enables study of the regulation of the biosynthesis. The use of mutantsand transgenic plants ultimately makes possible the analysis of biological function aswell as the generation of plants with al tered natural product profiles.

Molecular interactions form the basis of cellular function. An interdisciplinary analy-sis of these interactions is therefore of central importance to the research missionof the Institute of Plant Biochemistry. The optimal adaptation of plants to their habi-tat depends upon receptor-mediated perception of biotic and abiotic environmentalparameters. External signals are evaluated, compared and converted into physiologi-cal responses via al tered gene expression patterns that are controlled by cellular andsystemic signal transduction networks. The molecular basis of these processes, re -ceptor-ligand, enzyme-substrate and protein-protein interactions, have ap plication inthe development of new biologically active agents. From this perspective, the mech-anisms of communication between plants and their symbionts and pathogens areinvestigated as are biosynthetic and signal transduction pathways. Chemical struc-tures of these interacting components are also modified using gene technologicalmethods, directed evolution and chemical derivatization.The effects of these changescan be mo nitored in model systems or with activity screens until a molecule withthe desired characteristics (e.g. a drug, signal compound or an enzyme) is achieved.The development of new syntheses, screening tests, assays and analytical methods issupported by visualization of molecular interactions via computer modeling. A nexus of natural product research and the study of molecular interactions is the

Research Mission Statement

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storage and evaluation of the large amount of data that is ge nerated. In particular,high throughput processes used in metabolome and proteome analysis and in theproduction of combinatorial libraries make necessary the development of new me-thods in information technology. To this end, a new junior group in information tech-nology is being established at the Institute of Plant Biochemistry.

Metabolic engineering is an overlapping priority in three areas of basic research - nat-ural products, molecular interactions and information technology. Model plants aregenerated that have potential for various types of application. More specifically,designer plants with tailored natural product profiles, containing new health-promo-ting metabolites or showing im proved adaptation to habitat are being developed.Plants with these cha racteristics could serve for the sustainable production of valu-able chemicals, as biological test systems or could be of importance to plant bree -ders.

Within four departments with distinct, but complementary research directions andstate-of-the-art equipment, the Institute of Plant Biochemistry provides optimal con-ditions with which to execute multidisciplinary re search in the areas of chemistry,physiology, cell biology, biochemistry, molecular biology and genetics.The analysis oftopics central to modern plant biology and chemistry using this wide array ofmethodologies enables a meaningful interpretation of the complex interactions inplant development and diversity that would otherwise not be possible. The ultimatetransfer of these results to practical applications could make ecologically compatibleuses of plant biotechnology a reality.

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Department of Natural Product BiotechnologyHead: Toni M. Kutchan, ProfessorSecretary: Christine Dietel

Department of

Natural Product Biotechnology

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For much of human history, plant extracts have been used as ingredientsin potions and poisons. In the eastern Mediterranean, use of opium

poppy latex (Papaver somniferum) can be traced back at least to 1400 to1200 B.C. Ancient folks used medicinal plant ex tracts as purgatives, anti-tussives, sedatives and treatments for a wide range of ailments from sna -kebites to fever. Use of medicinal plants then spread westward across Eu -rope. Over the centuries, one of the most important medicinals wasopium. Analysis of the individual components of opium led to the identifi-cation of morphine. The isolation of morphine in 1806 by the Germanpharmacist Friedrich Sertürner gave rise to the study of alkaloids. Theterm "alkaloid" was, in fact, coined by another pharmacist, Carl Meissner, in1819 in Halle, Germany. Alkaloids were originally defined as pharmacolog-ically active, nitrogen-containing basic compounds of plant origin. Al ka loid-containing plants were mankind's original materia medica. Ma ny alkaloidsare still in use today as prescription drugs; one of the best known andwidely used is the antitussive codeine from the opium poppy. After 198years of alkaloid research, this class of natural products still finds signifi-cance in the medical field in the treatment of diseases ranging from cancerpotentially through to malaria.

The Department of Natural Product Biotechnology emphasizes researchon the elucidation of formation of plant natural products at the moleculargenetic level. Our research centers on understanding the formation of se -lected natural products at several levels and then exploiting this knowledgeusing ge netic engineering.We are just beginning to metabolically engineeralkaloid metabolism in plants and in in vitro culture. Multicellular compart-mentation of alkaloid pathways and transport of intermediates must beconsidered if meaningful metabolic engineering experiments are to bedesigned.We also need to analyze and use promoters that drive transgeneexpression in the correct cell types to effect biosynthesis. Re gulation ofthese pathways at the gene and enzyme level is complex and there is stillalso much to be learned about metabolite levels and pathway interconnec-tion as we systematically over-express and suppress gene transcription.Today, pathway engineering in plants re mains highly variable.When we per-turb cellular physiology, metabolitehomeostasis and intra- and intercel-lular partitioning can be effec ted inunpredictable ways. Another aspectthat needs attention is the develop-ment of efficient transformation andrege neration protocols for alkaloid-producing plant outside of the So la -naceae. All told, there is still much tobe done in this field of research.

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Research Group: Alkaloid BiosynthesisHead: Toni M. Kutchan

The opium poppy Papaver somnife-rum is the source of the narcoticanalgesics codeine and morphine,which accumulate in specialized in -ternal secretory cells called latici-fers. In the aerial parts of the plant,the laticifer cells are anastomosed,forming an articulated network. La -ticifers are found associated withthe vascular bundle in all plant parts.The morphinan alkaloids morphine,codeine and thebaine are foundboth in roots and in aerial plantparts and specifically accumulate invesicles within laticifers. The ben zo -[c]phenanthridine alkaloid sanguina-rine is found in root tissue.The syn-theses of sanguinarine and of thetetrahydrobenzylisoquinoline alka-loid laudanine are completely un -derstood at the enzyme level. Near -ly all enzymes of morphine biosyn-thesis have also been described. Inmore recent years, cDNAs enco-ding ten enzymes of alkaloid biosyn-thesis in P. somniferum have beenisolated and characterized. The

pathway starting from the first tetrahydrobenzy-lisoquinoline alkaloidal intermediate (S)-norco-clau-rine to the central isoquinoline alkaloid bio-synthetic intermediate (S)-reticuline is under-stood at both the enzyme and gene level. (S)-

Nor coclaurine is O-methylated by (R,S)-norco-clau-rine 6-O-methyltransferase. (S)-Coclaurineis next N-methylated by (R,S)-coclaurine N-methyltransferase and the cDNA encoding thisenzyme has been characterized. (S)-N-Methylcoclaurine is then hydroxylated by thecytochrome P-450 de pendent monooxygenaseCYP80B1 [(S)-N-methylcoclaurine 3'-hydroxlya-se].The cDNA en coding the corresponding cyto-chrome P-450 re ductase has been isolated aswell. (S)-3'-Hydroxy-N-methylcoclaurine ismethylated to (S)-reticu-line by (R,S)-3'-hydroxy-N-methylcoclaurine 4'-O-methyltransferase(4'OMT).The cDNA 4'omt has been isolated and

Group Members

Kum-Boo ChoiHumboldt-Fellow until September 2003

Maria Luisa Diaz-ChavezDAAD PhD Student since October 2003

Nils GünnewichPhD Student since December 2003

Aphacha JindaprasertDAAD PhD Student since December2004

Robert KramellPostdoctoral Position

Monika KrohnTechnician until October 2004

Kristin KrukenbergFulbright-Fellow until July 2003

Tobias KurzPhD Student

Alfonso LaraDAAD PhD Student since April 2003

Natsajee NualkaevDAAD PhD Student until December2004

Christin RichterPhD Student since August 2004

Khaled SabarnaPhD Student

Karin SpringobPostdoctoral Position since July 2003

Marion WeidPhD Student until March 2004

The opium poppy Papaver somniferum is one of mankind's oldest medicinal plants. Opium poppytoday is the commercial source of the narcotic analgesics morphine and codeine. Along with these twomorphinans, opium poppy produces approximately 80 alkaloids belonging to various tetrahydroben-zylisoquinoline-derived classes. It has been known for over a century that morphinan alkaloids accu-mulate in the latex of opium poppy.With identification of many of the enzymes of alkaloid biosynthesisin this plant, biochemical data suggested involvement of multiple cell types in alkaloid biosynthesis inpoppy. It has also been suggested, based upon the failure to produce morphine in undifferentiated P.somniferum cell cultures, that morphine accumulation is related to cytodifferentiation - noting thatlaticifers are absent from these cultures, but present in tissue cultures. We now have addressed thequestion of cell-specific localization of the enzymes of alkaloid biosynthesis and the site of gene tran-script accumulation.We have carried through the immunolocalization of five enzymes of alkaloid for-mation in opium poppy: (R,S)-3'-hydroxy-N-methylcoclaurine 4'-O-methyltransferase central to thebiosynthesis of tetrahydroisoquinoline-derived alkaloids, the berberine bridge enzyme of the sangui-narine pathway, (R,S)-reticuline 7-O-methyltransferase specific to laudanosine formation, and salutari-dinol 7-O-acetyltransferase and codeinone reductase that lead to morphine.

Immunolabeling of of SalAT (salutaridinol 7-O-acetyltransfe-rase) of morphine biosynthesis in a cross-section of cap-sule of P. somniferum. The panels show a cross-sectionthrough a small vascular bundle of the capsule wall. Thegreen arrows indicate the position of anti-biosyntheticenzyme antibody; the red arrows indicate the po sition ofanti-MLP 15 antibody and is indicative of laticifer cells.

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characterized from P. somniferum. (S)-Reticulineis central intermediate of isoquinoline alkaloidbiosynthesis, which leads to a plethora of alkaloi-dal structures. In P. somniferum, (R,S)-reticulinecan be methylated by (R,S)-reticuline 7-O-methyltransferase, for which the cDNA 7omt hasbeen described, to the tetrahydrobenzylisoquino-line laudanine. The N-methyl group of (S)-reticu-line can alternatively be oxidatively cyclized bythe berberine bridge enzyme (BBE) to C-8 of (S)-scoulerine. (S)-Scoulerine is then fur-ther conver-ted in these plants to the antimicrobialbenzo[c]phenanthridine alkaloids sanguina-rine.Along the pathway on which (S)-reticuline is spe-cifically converted to morphine, cDNAs encodingtwo biosynthetic enzymes have been identified.Salutaridinol 7-O-acetyltransferase, encoded bysalat, transfers an acetyl moiety from acetyl-CoAto the 7-hydroxyl group of salutaridinol.Codeinone reductase is encoded by cor andcatalyzes the penultimate step in morphine bio-synthesis, the NADPH-dependent reduction ofthe keto moiety of codeinone to the 6-hydroxylgroup of codeine.

Fluorescence immunocytological localization wascarried out with the 4’OMT (reticuline- ), the7OMT (laudanine- ), BBE (sanguinarine- ), SalATand COR (morphine-pathways) with P. somnife-rum capsule, stem and root tissue sections. In thismanner, the spatial organization of enzymes oc -

curring before and after a centralbranch point is analyzed. In additi-on, in situ localization of 7omtand cor1 was performed to cor-relate the site of gene transcripti-on to enzyme ac cumulation. Incapsule and stem, both O-methyl-transferases and the O-acetyl-transferase are found predomi-nantly in parenchyma cells withinthe vascular bundle and co -deinone reductase is localized tolaticifers, the site of morphinan al -kaloid accumulation. In developingroot tip, both O-me thyltrans fera -ses and the O-acetyltransferaseare found in the pericycle of the stele and the ber-berine bridge enzyme is localized to parenchymacells of the root cortex. Laticifers are not foundin developing root tip and, likewise, codeinonereductase was not detected. These results provi-de cell-specific localization that gives a co herentpicture of the spa-tial distribution of alkaloid bio-synthesis in opium poppy.

In summary, two cell ty -pes, parenchyma within thevascular bundle and la tici -fers, are sites of the bio-synthesis of isoquinolineal kaloids in P. somniferum.This implies spatial regula-tion of the biosynthesis ofthe different classes of P.somniferum alkaloids inthat the early stages ofmor-phine biosynthesisoccur in parenchyma cellssurrounding laticifers andthen at late stages, possibly at the level of eithersalutaridinol-7-O-acetate or thebaine, moves intothe laticifer, which is the storage site of the mor-phinans thebaine, codeine and morphine. The roleof intercellular transporters of alkaloidal inter-mediates as well as intracellular transport intovesicles within laticifers adds an additional poten-tial level of regulation to morphine biosynthesisthat still needs to be investigated.

Collaborators

Gerhard BringmannUniversity of Würzburg, Germany

Wanchai De-EknamkulChulalongkorn University, Bangkok,Thailand

Tony FistTasmanian Alkaloids, Tasmania, Australia

Phil LarkinsScientific and Industrial ResearchOrganisation Plant Industry, Canberra,Australia

Friedrich LottspeichMax Planck Institute for Biochemistry,Martinsried, Germany

Markus PiotrowskiUniversity of Bochum, Germany

Joachim StöckigtUniversity of Mainz, Germany

Dissected capsule of P. somniferum, white arrow indi-cates the position of the large vascular bundle; the smallvascular bundle is located closer to the capsule wall.

Schematic drawing of a longitudinal section through thevascular bundle of P. somniferum stem. xy, xylem; cc, compa-nion cell; la, laticifer; ph, phloem; sp, sieve plate.

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Research Group: Opium Poppy BiotechnologyHead: Susanne Frick

During the last years several genesfrom the biosynthetic pathways forreticuline, sanguinarine and morphi-ne have been cloned (Fig. 1). Al -though the biosynthesis is well un -derstood at the enzyme level, themolecular and biochemical mecha-nisms that regulate these pathwaysare not known. A major goal of thisproject was the development of astable transformation and regenera-tion method for opium poppy, whichwould make the metabolic enginee-ring of the above-mentioned com-pounds possible. Poppy seed oil findsuse in chemical industry for the pro -duc tion of pigments and lacquer, butits residual morphine levels preventsmore widespread applications. Be -

cause opi um is the raw ma terial for the illicit pro-duction of heroin, cultivation of poppy is re stric -ted. By completely sup pressing morphine biosyn-thesis, opium pop py could be come a "harmless"crop plant. So far there has been no success withbreeding programs and mu tations to obtain analkaloid-free poppy. In the best case a re ductionof alkaloid biosynthesis has been achieved. Thetransforma tion of opium poppy could be an alter-native to circumvent these problems.

We have used an Agrobac te rium-mediated ap -proach to introduce different cDNAs encodingenzymes of reticuline, morphine and sanguinarinebiosynthesis as sense-, antisense- or RNAi-con-structs into explants to attempt to alter the alka-loid profile (Fig. 2, A-E). Al ka loid-free plants de -veloped in this manner will be used to test the

chemical -ecological func tion of morphinan andbenzophenanthridine alkaloids in plants.After theregeneration of plants, the qualitative and quanti-tative determination of alkaloids is analyzed byHPLC and LC-MS in latex, leaves and roots ofopium poppy. Finally the heredity of the alkaloidconcentration/pattern is confirmed.With a trans-genic cell line expressing the antisense constructof berberine bridge enzyme (BBE) we hoped toreduce the metabolic flux through sanguinarinepathway and to enhance the concentration ofmorphine instead. We are interested whether atransgenic cell line over expressing codeinone re -ductase (COR) leads to a poppy plant that con-tains more morphine or where the concentrati-on of morphine is lowered due to a possible

Group Members

Kathleen GutezeitTechnician

Stefanie HaasePhD Student

Elke HillertTechnician

Katja KempePhD Student

Heike RieglerDiploma Student since October 2004

CollaboratorsTony Fist, Tasmanian Alkaloids, Westbury, Australia

Philip Larkin, Scientific and Industrial ResearchOrganisation Plant Industry, Canberra,Australia

Jürgen Schmidt, Institute of Plant Biochemistry, Halle,Germany

Opium poppy (Papaver somniferum L.), which contains more than 80 different alkaloids, remains oneof the most important industrial medicinal plants. Poppy serves as a renew able resource of a numberof medicinally relevant alkaloids. These include the analgesic and narcotic drug morphine, the coughsuppressant codeine, as well as the muscle relaxant papaverine, the antitumoric agent noscapine andthe antimicrobial sanguinarine.We have developed a transformation system for opium poppy that willallow us:

to investigate the regulation and ecological function of these alkaloids in plants

and to alter the alkaloid metabolism in commercial poppy varieties in order to obtain varieties lacking alkaloids or with tailored alkaloid profiles for industrial and pharmaceutical use.

Figure1: Biosynthetic pathway from L-tyrosine to sanguinarine, lau-danosine and morphine in P. sonmniferum. Enzymes are highlightedin red.

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feedback inhibition of this pathway. We havealso produced poppy transformants, in whichwe potentially influenced all cytochromeP450 enzymes of the benzylisoquinolinepathways by introducing a NADPH:cyto-chrome P450-oxidoreductase (CPR). Finallywe are trying to reduce or silence the com-plete alkaloid biosynthetic pathway with atransgenic cell line containing the antisenseconstruct of (S)-N-methylcoclaurine hydro-xylase (CYP80B1). With a cell line overex-pressing cyp80b1 we are trying to stimulateall three pathways to gether.

With the antisense expression of bbe cDNAwe have been able to produce transgenicopium poppy plants with altered alkaloid profi-les. The transformation of these plants was eva-luated by PCR, northern and Sou thern hybri-disation. The transgenic plants con tained oneadditional copy of the bbe gene.We observedan in creased concentration of several pathwayin termediates from all biosynthetic branchese.g. reticuline, laudanine, laudanosine, dehydro-reticuline, salutaridine and (S)-scoulerine. Thetransformation altered the ratio of morphinanand tetrahydrobenzylisoquinoline alkaloids inlatex but not the benzophenanthridine alka-loids in roots. The altered alkaloid profile washeritable at least to the T2 generation.

The overexpression and antisense expressi-on of cyp80b1 cDNA produced poppyplants with altered alkaloid concentrations.The in crease or decrease of alkaloid concen-tration in the transgenic cell lines was herita-ble at least to the T3 generation. Comparedto cyp80b1 the overexpression of cpr cDNAdid not give rise to an increased alkaloidconcentration in transgenic opium poppy.

Because we have not been able to silencebenzylisoquinoline biosynthesis in transgenicpoppy plants containing S4S4:antiBBE orS4S4:antiCYP80B1, we constructed plasmidscontaining partial sequences that are poten -tially able to trigger RNA interference in Pa -pa ver somniferum. Explants of opium poppywere transformed with different constructs.Meanwhile we have been able to produce se -veral T0 plants. The analysis of these plants isin progress.

To study the cell- and tissue specific expres-sion of benzylisoquinoline biosynthetic ge nes,we started to isolate their endogenous pro-moters. With promoter/reporter gene con-structs we want to analyze how biosynthesisand accumulation of alkaloids are regulated.

A

B

C

D

E

Figure 2: Somatic re generationof P. somniferum. Ex plants firstgive rise to a type I callus (A),which starts to differentiate aftera certain time (B). After thetransfer to a hormone freemedium this type II callus deve-lops small em bryos (C) and fi nal -ly little plant lets (D). The wholeregeneration process is shownin picture E.

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Research Group: Mode of Action of JasmonatesHeads: Claus Wasternack & Otto Miersch

The role of JA biosynthesis isanalyzed in tomato and Arabi -dopsis. Preferentially, the AOC-catalyzed step and the profile ofjasmonates and other oxylipinsare studied to elucidate specificfunctions in plant stress respon-ses and development.

The first AOC was cloned fromtomato (Ziegler et al. 2000, pa -tent No. DE 1000 4468.9). Thissingle copy gene is specifically ex -pressed in ovules of young flo-wers and all vascular bundles. Inleaves, the vascular bundle-speci-fic occurrence of AOC attributesto a preferential ge neration ofjas mo nates in main veins, whichled together with data from va -rious transgenic tomato plants toa proposed amp lification modelin wound signaling with preferen-tial signaling properties of JA.Thisis supported by de tection of JA

bio synthetic enzymes in clu ding AOC in sieve e -lements of to ma to leaf veins (Fig. 1).

Using transgenic tomato plants o ver-expressing orre pres sing con sti tutively the AOC or expressing areporter gene dri ven by the AOC promoter (coop. T.Roitsch, Wue r z burg, Germany), role of AOC andgeneration and func tion of jasmonates could be in -spected.Tissue specific expression of AOC in seed-ling development, preferentially in roots (Fig. 3) andduring flower and em bryo development points tonew functions of JA and its metabolites. This AOCgene activity and JA function differs from that ofArabidopsis suggesting the plasticity of differentplant species to use signaling compounds. These dif-ferences together with data from the literaturepoint to role of JA in female fertility of tomato, whe-reas in Arabidopsis male fertility is strictly JA depen-dent.

Interestingly, the vascular bundle- and ovule-specificoccurrence of AOC protein in flowers is accompa-nied with a specific profile of octadecanoids and jas-monates as well as JA amino acid conjugates beingdifferent in the various flower organs. Constitutiveover-expression of AOC led to qualitative and quan-titative changes in these profiles (Fig. 2) as well asthat of the free and esterified fatty acids and theirperoxidation products (coop. I. Feussner, Universityof Goettingen). The data suggest different regulationof JA biosynthesis in flowers and leaves and indicatethat the different branches of the lipoxygenase path-way can be altered depending of the terminal pro-ducts.

Jasmonates (JA) and their precursors, the octadecanoids, are signals in plant stressresponses and in plant development. A mechanistic analysis of the mode of action ofjasmonates is performed by a reverse genetic approach using the allene oxide cycla-se (AOC)-catalyzed step in jasmonate biosynthesis. "Gain of function" and "Loss offunction" studies with transgenic tomato plants revealed modulation of jasmonatesand allowed to inspect role of jasmonates in response to biotic and abiotic stressesas well as flower and seed development. Functional analysis of AOC using geneticapproaches is also performed in Arabidopsis to ask on specific versus redundantfunctions of the four AOCs in this plant. These data are linked to metabolite profileson jasmonates and octadecanoids under various stress conditions and in differentdevelopmental stages. In addition to this analytical work, chemical synthesis of stan-dards and labeled substrates is an essential part of this work

Group Members

Carolin DelkerPhD Student since June 2002

Conrad DorerDiploma Student until February 2004

Yvonne FreyerDiploma Student until October 2004

Lydia MüllerDiploma Student until July 2003

Jana NeumerkelDiploma Student until November 2003,afterwards PhD Student

Birgit OrtelTechnician

Katrin RehagelDiploma Student since October 2004

Irene StenzelPostdoctoral Position

Carola UhligTechnician

Sabine VorkefeldTechnician until May 2004

Collaborators

Guillermina AbdalaUniversidad Nacional de Rio Cuarto, Argentina

Wilhelm BolandMax Planck Institute of Chemical Ecology, Jena,Germany

Udo ConradInstitute of Plant Genetics and Crop PlantResearch, Gatersleben, Germany

Ivo FeussnerUniversity of Goettingen, Germany

Bettina Hause, Sabine Rosahl,Dierk Scheel, Jürgen SchmidtInstitute of Plant Biochemistry, Halle, Germany

Gerd HauseUniversity of Halle, Germany

Gregg HoweUniversity of Michigan, East Lansing, USA

Harry KleeUniversity of Florida, Gainesville, USA

Thomas RoitschUniversity of Würzburg, Germany

Luc VarinConcordia University, Montreal, Canada

Figure1: AOC protein occurs in companion cells (CC) and sieve elements (SE), but AOC mRNA is exclusivelylocated in companion cells. AOC protein is detected by immunolabeling indicated by green fluorescence(left). In situ hybridisation to detect AOC mRNA was performed by probing with an AOCantisense probe(middle), note that the triangle-shaped sieve elements are free of label. As a control an AOCsense probe wasused indicating no cell-specific labelling (right) (taken from Hause et al., Plant Cell Physiol. 44, 643-648, 2003).

15

Data on JA and 12-oxo-phytodienoic acid (OPDA)level in tomato mutants and in our 35S::AOC anti -sense tomato plants following infection with Xan -tho monas campestris cv. vesicatori supported amodel of sequential action of jasmonate, ethyleneand salicylate in this plant pathogen interaction(coop. H. Klee, Gainesville, USA). Also in collabora-tive work (M. Köck, Biocenter, University of Halle,Germany), data were found which support a new JAsignaling pathway. In tomato the wound-responsiveRNaseLE gene is expressed only locally and in a JA-and systemin-independent manner.

In Arabidopsis the AOC is encoded by four genes.Inspection of transgenic Arabidopsis plants expres-sing a reporter gene driven by each AOC promoter,revealed non-redundant functions of the AOCs. Pre -ferentially, in root growth and seedling developmentas well as in flower development (Fig. 4) there aretissue and organ specific promoter activities, non-redundantly for AOC1, AOC2, AOC3 and AOC4.These data and analyses of knockout-lines indicatethat Ara bi dopsis is able to perform a fine-tuning of

regulation by spatialand temporaldifferen ces in theexpression of thefour AOCs. In con-trast, the fourrecombinant AOCsexhibited similar en -zymatic propertiesand substrate speci-ficities.

Previously, 12-hydroxy-JA was only known as tuber-inducing compound in So la naceae species. We couldidentify 12-hydroxy-JA and its sulfated derivative inA. thaliana and tomato. From Arabi dop sis a sulfo-transferase (AtST2a) was cloned which convertsspecifically 12-hydroxy-JA (coop. L. Varin, Montreal,Canada). In transgenic plants over-expressing or re -pressing constitutively AtST2a a shift in floweringtime depending on the 12-OH-JA level was detec-ted. In tomato 12-OH-JA is a permanent constitu-ent of flower organs. In to mato leaves 12-OH-JA isformed upon wounding in a JA-dependent manner.The role of 12-OH-JA in a photoperiod-dependentplant and a day-neutral plant is in the focus of futurework.

National and international cooperation is done in theabove-mentioned aspects of tomato and Ara bidopsis.The know-how of the group on JA and oxylipin analy-tics is used and requested by many groups. A minorac tivity is linked to collaboration with the ProbiodrugAG in Halle.The company is working on role of gluta-minylcyclases (QCs) of human, animal and plant sour -ces. QCs attributein mammalian sys -tems to a functiona-lization of peptidehormones and pro-teins by pyrogluta-mate formation atthe N-ter minus. Oureffort is to cloneplant ho mo lo gues ofthe QC as a tool forde sign of potent QCinhibitors.Figure 2: Constitutive over-expression of AOC in tomato leads to elevat-

ed levels of jasmonates and octadecanoids in flowers but not in leaves. Theratio among the compounds being specific for each flower organ is diffe-rentially altered by over-expression of AOC. The diameter of each circleis indicative for the total amount of all compounds, the sector within a cir-cle is indicative for a distinct compound given in % of the total amount(taken from Miersch et al., Phytochem. 65, 847-856, 2004).

Figure 3: AOC promoter activity in tomato seedling development. Seedlingsof a trans genic AOC(1000 bp)::uidA line expressing the b-glucorunidase,whose activity is indicated by blue staining, were inspected after 3 and 10days, respectively, after sawing (das). Note the AOC promoter activity in theroot tip and hypocotyl 3 das (left) and the root elongation zone as well asthe vascular bundles of cotyledons 10 das (right). AOC promoter activitycorresponds to location of AOC protein probed with an anti-AOC antibodyand visualized by violet staining (left /right) (photos by I. Stenzel and B.Hause).

Figure 4: Promoter activity of the AOC4 gene of Arabidop sis thaliana. 14-d-oldseed lings (above) and flowers (below) of transgenic lines carrying a AOC4::uidA con-struct were inspected on b-glucuronidase activity visualized by the blue staining.Note AOC4 promoter activity in root tips, side root primordia and vascular bund-les of the seedlings, as well as in sepals, petals, filaments and stigmas of open flo-wers (photos by I. Stenzel).

anti AOC

3-d-old seedling 10-d-old seedling

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Research Group: Papaver-Gene Expression AnalysisHead: Jörg Ziegler

Currently, more than 70 differentpoppy species belonging to thegenus Papaver have been described.Roughly, they are able to synthesiseabout 2,500 different benzylisoqui-nolines, which can be grouped intonine classes. The profile of benzyl -isoquinolines produced by the plants

are species specific; it is, however also dependenton growth conditions. The same holds true forgene expression. This variability requires sensi-tive methods to re cord all needed parameters inone individual plant. HPLC methods were em -ployed to detect the main compounds and LC-MS coupling will be used for the low abundantcompounds. For gene expression analysis, a pro-tocol for macroarray production was developed.These methods are sensitive enough to recordthe alkaloid profile and the gene expression pat-tern from one individual plant. For ten Papaverspecies and five varieties and mutants of theopium poppy Papaver somniferum, the alkaloidprofile was recorded by HPLC and HPLC-MS.Some of the major compounds of each plantcould be identified by MS-MS. The MS-MS data of

all other alkaloids are available now and their in -terpretation with respect to their identification isin progress.

As probes for the arrays we use PCR fragmentsderived from two EST-projects of Papaver somni-ferum stems and seedlings. Among all Papaverspecies, this plant synthesizes the largest numberof different benzylisoquinolines. Up to now wesequenced about 3,700 ESTs and obtained 2,000UniGenes. About 50 percent either code for pro-teins with unknown function or have no homolo-gy to entries in the databases. The largest groupsof cDNAs coding for proteins with known func-tion are involved in transcriptional and translatio-nal control, in responses to stress, and in redoxcontrol. Another highly represented group codesfor proteins participating in metabolism, mainlyprimary metabolism. To 20 sequences, a possiblerole in secondary metabolism could be ascribed.Six sequences code for proteins with knownfunction in the benzylisoquinoline pathway. Up tonow, the expression of these cDNAs was exa-mined in nine Papaver species and P. somniferumvarieties, respectively. These expression profiles

Group Members

Andreas Gesell PhD Student

Susan VoigtländerDiploma Student since November2004

Silvia Wegener Technician

Poppies of the genus Papaver produce a large variety of benzylisoquinoline alkaloids. Some of themare of pharmaceutical importance such as the analgesic morphine, the antitussive noscapine or thevasodilator papaverine. The biosynthesis to (S)-reticuline, the central intermediate to all monomericbenzylisoquinoline alkaloids is well understood on the molecular level, the knowledge of the latersteps, which lead to the diversity of this class of compounds, is still incomplete. Similarly, the regulatorysteps leading to the accumulation of these substances, is unknown. To approach cDNA clones codingfor these processes we make use of the close genetic relationship but the diversity in the alkaloid pro-file between Papaver species or varieties.We examine and correlate the gene expression profiles onEST-arrays with specific alkaloid profiles. By the combination of many different datasets of alkaloidprofile -gene expression correlations, we could re-duce the number of candidate cDNAs with possiblefunctions in benzylisoquinoline accumulation to eleven.

Functional classification of ESTs from P. somniferum stems and seedlings. Abbreviations: PS: photosynthesis, MLP: major latex proteins, protein syn/deg: proteinsynthesis and degradation, G-prot: G-proteins, AA: amino acids, alk: alkaloids, phenyl: phenylpropanoids, isopr: isoprenoids.

17

were compared with the alkaloid profile of eachplant. In a first approach, we looked at cDNAsshowing differential expression in morphinan al -kaloid producing P. somniferum plants comparedto other Papaver species, which do not producethis class of benzylisoquinolines. From 2,000 ESTsexamined, only eleven cDNAs showed morphi-nan specific expression. Mostly, these showed nohomology at all or homology to proteins withunknown function or to putative proteins whencompared to the NCBI database. One of the dif-ferentially expressed cDNAs showed homologyto an O-methyltransferase.The full-length cDNAwas isolated and the protein was overexpressedin bacteria. Functional characterization of the re -combinant protein identified this cDNA as 3 -hydroxy-N-methylcoclaurine 4 -O-methyltrans-ferase. The enzyme showed a strict substratespecificity in that it only accepted tetrahydroben-zyl-isoquinolines as substrates, and not, as otherclass II-O-methyltransferases, phenolic substan-ces.

A second approach to identify cDNAs implicatedin the biosynthesis of benzylisoquinolines and theregulation of alkaloid accumulation implies thefunctional characterization of ESTs showing ho -mology to proteins, where such a role can be as -sumed.

The enzyme family of the P450-monooxygenasesare known to catalyze many reactions in the struc-tural modification of the benzylisoquinoline back-

bone. Up to now, 33 ESTscoding for these enzymescould be isolated. One of theseshowed high homology to apreviously characterized P-450enzyme, which catalyses theformation of a me thyl ene dioxybridge in benzylisoquinolinebiosynthesis. The full-lengthcDNA of this EST was isolatedand its substrate specificity is under investigation.In order to a chie ve high expression levels of thisen zyme, an overexpression sys tem in Nicotianabenthamiana using viral vectors is currentlyestab lished.

Recent localization studies indicated, that the en -zymes of benzylisoquinoline biosynthesis are lo -cated in different cell types, suggesting that in ter -mediates have to be transported between cells.The large group of ABC-transporters have beenshown to transport many secondary metabolites,among them also alkaloids. In our EST da tabase,we found ten sequences coding for ABC-trans-porters. Currently, we are isolating the full-lengthcDNAs for three of these transporters. For func -tional characterization of the transporters, com-plementation of yeast strains deficient in ABC-transporter will be used. In contrast to the wild-type yeast, the mu tants are not able to transportbenzylisoquinolines. Overexpression of an ABC-transporter with specificity towards benzyliso-quinolines in the mutants should restore wild

Collaborators

Christian AmmerInstitute of Plant Biochemistry, Halle,Germany

Birgit DrägerUniversity of Halle, Germany

Sylvestre MarillonnetIcon Genetics AG, Biozentrum Halle,Germany

Jürgen SchmidtLeibniz Institute of Plant Biochemistry, Halle,Germany

Expression level of selected cDNAs in different Papaver species.

PublicationsAbdala, G., Miersch, O., Kramell, R., Vi g li oc -co, A., Agostini, E., Forchetti, G. & Alemano,S. Jasmonate and octadecanoid occurrencein tomato hairy roots. Endogenous levelchan ges in response to NaCl. PlantGrowth Regul. 40, 21-27.

Bailey, N. J. C., Oven, M., Holmes, E., Ni -chol son, J. K. & Zenk, M. H. Meta bolo micanalysis of the consequences of cadmiumexposure in Silene cucubalus cell culturesvia 1H NMR spectroscopy and chemome-trics. Phytoche mis try 62, 851-858.

Färber, K., Schumann, B., Miersch, O. &Roos, W. Selective desensitization of jas-monate- and pH-dependent signalling inthe induction of benzophenanthridine bio-synthesis in cells of Eschscholzia californica.Phytochemistry 62, 491-500.

Gidda, K. S., Miersch, O., Schmidt, J., Was ter -nack, C. & Varin, L. Biochemical and molecu-lar characterization of a hydroxy-jasmo natesulfotransferase from Arabidop sis thaliana. J.Biol. Chem. 278, 17895-17900.

Hause, B., Hause, G., Kutter, C., Miersch, O. &Wasternack, C. Enzymes of jasmonate bio-synthesis occur in tomato sieve elements.Plant Cell Physiol. 44, 643-648.

Hause, B., Stenzel, I., Miersch, O. & Waster -nack, C. Occurrence of the allene oxide cyc -lase in different organs and tissues of Arabi -dopsis thaliana. Phytoche mistry 64, 971-980.

Monostori, T., Schulze, J., Sharma, V. K., Mau -cher, H., Wasternack, C. & Hause, B. No velplasmid vectors for homologous transfor-mation on barley (Hordeum vulgare L.) withthe JIP23 cDNA in sense and antisenseorienta tion. Cereal Res. Comm. 31, 17-24.

O'Donnell, P. J., Schmelz, E., Block,A., Miersch,O., Wasternack, C., Jones, J. B. & Klee, H. J.Multiple hormones cooperatively control asusceptible tomato pa tho gen de fense re -sponse. Plant Physiol. 133, 1181-1189.

Ounaroon, A., Decker, G., Schmidt, J., Lott -speich, F. & Kutchan, T. M. (R,S)-Re ti culine7-O-methyltransferase and (R,S)-nor co -clau rine 6-O-methyltransferase of Papaversomniferum-cDNA cloning and characte-rization of methyl transfer enzymes ofalkaloid biosynthesis in opium pop py. PlantJ. 36, 808-819.

Pedranzani, H., Racagni, G., Alemano, S.,Miersch, O., Ramírez, I., Peña-Cortés, H., Ta -leisnik, E., Machado-Domenech, E. & Abda la,G. Salt tolerant tomato plants show increa-sed levels of jasmonic acid. Plant GrowthRe gul. 41, 149-158.

Samappito, S., Page, J.E., Schmidt, J., De-Ek -namkul, W. & Kutchan, T. M. Aromatic andpyrone polyketides synthesized by a stilbe-ne synthase from Rheum tataricum. Phyto -che mistry 62, 313-323.

Schilling, S., Manhart, S., Hoffmann, T., Lud - wig, H.-H., Wasternack, C. & De muth, H.-U. Sub strate specificity of glutaminyl cyc -lases from plants and animals. Biol. Chem.384, 1583-1592.

Schilling, S., Niestroj, A. J., Rahfeld, J.-U.,Hoffmann, T., Wermann, M., Zunkel, K.,Was ternack, C. & Demuth, H.-U. Iden ti fi -ca tion of human glutaminyl cyclase as ametalloenzyme - Potent inhibition by imida-zole derivatives and heterocyclic chela-tors. J. Biol. Chem. 278, 49773-49779.

Stenzel, I., Hause, B., Maucher, H., Pitzsch -ke, A., Miersch, O., Ziegler, J., Ryan, C. A. &Waster nack, C. Allene oxide cyc lase de -pendence of the wound res ponse and vas-cular bundle spe cific generation of jasmo-nates in tomato - amplification in wound-signalling. Plant J. 33, 577-589.

Stenzel, I., Hause, B., Miersch, O., Kurz, T.,Maucher, H., Weichert, H., Ziegler, J., Feuss -ner, I. & Wasternack, C. Jasmonate biosyn-thesis and the allene oxide cyclase familyof Ara bidopsis thaliana. Plant Mol. Biol. 51,895-911.

Stenzel, I., Ziehte, K., Schurath, J., Hertel, S.C., Bosse, D. & Köck, M. Differential ex -pression of PSI14, a phosphatase gene fa -mily, in response to phosphate availability,pathogen infection and during develop-ment. Physiol. Plant 118, 138-146.

Books and Book ChaptersStenzel, I., Hause, B., Feussner, I. & Was ter -nack, C. Transcriptional activation of jas-monate biosynthesis enzymes is not re -flected at protein level. In: Advanced Re -search on Plant Lipids (Murata, N., Ya mada,M., Nishida, I., Okuyama, H., Se ki ja, J. & Ha -jime, W., eds.) Kluwer Aca demic Pu bli -shers, Dordrecht, pp. 267-270.

Stumpe, M., Stenzel, I, Weichert, H., Hau se,B. & Feussner, I. The lipoxygenase path wayin mycorrhizal roots of Medica go trunca-tula. In: Advanced Research on Plant Lipids(Murata, N., Yamada, M., Ni shi da, I., Oku ya -ma, H., Sekija, J. & Hajime, W., eds.) KluwerAcademic Publishers, Dordrecht, pp. 287-290.

Weichert, H., Maucher, H., Hornung, E., Was -ternack, C. & Feussner, I. Shift in fat ty acidand oxylipin pattern of tomato leaves follo-wing overexpression of the alle ne oxide cyc -lase. In: Advanced Research on Plant Lipids(Mu rata, N., Yama da, M., Nishida, I., Okuyama,H., Sekija, J. & Hajime, W., eds.) Kluwer Aca -demic Pub lishers, Dordrecht, pp. 275-278.

PatentsKutchan, T. M., Ounaroon, A., Haase, S. &Frick, S. O-Methyltransferases of tetrahy-drobenzylisoquinoline alkaloid bio syn the -sis in Papaver somniferum. Euro peanPatent Application No. 03020023.2.

Varin, L., Miersch, O., Maucher, H. & Was -ternack, C. Methods and genetic se quen cesfor producing male sterile plants, and plantsgenetically modified to alter anther deve-lopment. USA/Canada Patent Ap pli cationNo. N/R: 004807-0007.

Diploma ThesesDorer, Conrad: Enzymatische Eigen schaf -ten der vier Allenoxidcyclasen aus Arabi -dopsis thaliana und Untersuchungen zurAOC-Aktivität transgener Pflanzen sowieArabidopsis-Mutanten, University of Halle-Wittenberg, Department of Bio chemist ry /Biotechnology, 24 /3/2004.

Günnewich, Nils: Expression und Cha rak -terisierung von Terpensynthasen und Pre -nyltransferasen aus Cannabis sativa L., Uni -versity of Halle-Wittenberg, Depart mentof Bioche mistry / Biotech no logy, 23/09/2003.

Kempe, Katja: Herstellung von Vektoren zur"RNA interference" für Gene der Ben zyliso -chinolin-Alkaloid-Biosynthese aus Pa pa versomniferum L. Technical Uni ver sity, Berg aka -demie Freiberg, 18/02/2003.

Müller, Lydia: Untersuchungen zur Regu la -tion der Promotoren von AOC1, AOC2,AOC3 und AOC4 in Arabidopsis thaliana,University of Halle-Wittenberg, De part -ment of Biochemistry /Biotech nology,16/07/2003.

Neumerkel, Jana: Gewebsspezifische Ana -lyse der Jasmonatwirkungsweise in der lo -kalen Wundantwort der Tomate, Univer si -ty of Halle-Wittenberg, Depart ment of Bio -chemistry/Biotechnology, 26/11/2003.

18

Publications and Other Scientific Activities 2003

Department of

Natural Product Biotechnology

19

PublicationsBücking, H., Förster, H., Stenzel, I., MierschO., & Hause, B. Applied jasmonates accu-mulate extracellularly in to ma to, but intra-cellularly in barley. FEBS Lett. 562, 45-50.

Frick, S., Chitty, J. A., Kramell, R., Schmidt, J.,Allen, R. S., Larkin, P. J. & Kutchan, T .M.Transformation of opium poppy (Papa versomniferum L.) with antisense berberinebridge enzyme 1 (anti-bbe1) via so ma ticem bryogenesis results in an altered ratio ofal kaloids in latex but not in roots.Transgenic Res. 13, 607-613.

Groß, N., Wasternack, C. & Köck, M.Wound induced RNaseLE expression isjasmonate and systemin independent andoc curs only locally in tomato (Lyco -persicon esculentum cv. Lukullus). Phyto -che mistry 65, 1343-1350.

Halitschke, R., Ziegler, J., Keinänen, M. &Baldwin, I. T. Silencing of hydroperoxidelyase and allene oxide synthase revealssub strate and defense signaling crosstalk inNicotiana attenuata. Plant J. 40, 35-46.

Köck, M., Groß, N., Stenzel, I., & Hause, G.Phloem-specific expression of the wound-inducible ribonuclease LE from tomato (Ly -copersicon esculentum cv. Lu kul lus). Planta219, 233-242.

Ma, X.-Y., Koepke, J., Fritzsch, G., Diem, R.,Kutchan, T. M., Michel, H. & Stöckigt, I. Cry s -talization and preliminary X-ray crystallo-graphic analysis of strictosidine synthasefrom Rauwolfia - the first member of a no -vel enzyme family. Biochim. Biophys. Acta1702, 121-124.

Maucher, H., Stenzel, I., Miersch, O., Stein,N., Prasad, M., Zierold, U., Schwei zer, P.,Dorer, C., Hause, B. & Waster nack, C. Theallene oxide cyclase of barley (Hordeumvulgare L.) - cloning and organ-specific ex -pression. Phytochemis try 65, 801-811.

Miersch, O., Weichert, H., Stenzel, I., Hau se,B., Maucher, H., Feussner, I. & Was ternack, C.Constitutive overexpression of allene oxidecyclase in tomato (Lyco persicon esculen-tum cv. Lukullus) elevates levels of jasmona-tes and octadecanoids in flower organs butnot in lea ves. Phyto chemistry 65, 847-856.

Millgate, A. G., Pogson, B.J., Wilson, I. W,Kut chan, T. M., Zenk, M. H., Gerlach, W. L.,Fist, A. J. & Larkin, P. J. Morphine pathwayblock in top1 poppies. Nature 431, 413-414.

Page, J. E., Hause, G., Raschke, M., Gao, W.,Schmidt, J., Zenk, M. H. & Kutchan, T. M.Functional analysis of the final steps of the1-deoxy-D-xylulose 5-phosphate (DXP)pathway to isoprenoids in plant using vi -

rus -induced gene silencing. Plant Physiol.134, 1401-1413.

Schüler, G., Mithöfer, A., Baldwin, I.T., Ber -ger, S., Ebel, S., Santos, J. G., Herrmann, G.,Hölscher, D., Kramell, R., Kutchan, T. M., Mau -cher, H., Schneider, B., Stenzel, I., Was ternack,C., & Boland, W. Co ro nalon: a pow erful toolin plant stress physiology. FEBS Lett. 563, 17-22.

Weid, M., Ziegler, J. & Kutchan, T .M.The rolesof latex and the vascular bundle in morphinebiosynthesis in the opium poppy, Papaversomniferum. Proc. Nat. Acad. Sci. USA 101,13957-13962.

Publications in PressFrick, S., Kramell, R., Larkin, P. J. & Kut chan,T. M. Studying morphine biosynthesis usingtransgenic opium poppy (Papa ver somnife-rum L.). Acta Hortic.

Frick, S., Kramell, R., Schmidt, J., Fist, A. J. &Kutchan, T. M. Comparative qualitative andquantitative determination of alkaloids innarcotic and condiment Papaver somnife-rum cultivars. J. Nat. Prod.

Gerhard, B., Fischer, K., Balkenhohl, T. J.,Pohnert, G., Kühn, H., Wasternack, C. &Feussner, I. Lipoxygenase-mediated me ta -bolism of storage lipids in germinating sun-flower cotyledons and beta-oxidation of(9Z, 11E, 13S)-13-hydroxy-octadeca-9, 11-dienoicacid by the cotyledonary glyoxyso-mes. Planta.

Kutchan, T. M. A role for intra and intercel-lular translocation in natural product bio-synthesis. Curr. Opin. Plant Biol.

Kutchan, T. M. Predicitve plant metabolicengineering - still full of surprises. TrendBiotechnol.

Ounaroon, A., Frick, S. & Kutchan T. M. Mo -le cu lar genetic analysis of an O-methyl-transferase of the opium poppy Papaversomniferum. Acta Horticulturae.

Rudus, I., Kepczynska, E. Kepczynski J., Was -ternack C. & Miersch, O. Distinct changesin jasmonate and 12-oxophytodienoic acidcontent of Medicago sativa L. du ring soma-tic embryogenesis. Acta Physiol. Plant.

Wasternack, C. Jasmonates - Overview onbiosynthesis and diversity in actions. In: Jas -monates, Special Issue of J. Plant GrowthRe gul. (Wasternack, C., ed.).

Ziegler, J., Diaz-Chávez, M. L., Kramell, R.,Ammer, C., & Kutchan, T. M. Comparativema croarray analysis of morphine contai-ning Papa ver somniferum and eight mor -phine free Papaver species identifies an O-methyltransferase involved in benzyliso-

quinoline biosynthesis. Planta.

Ziegler, J & Kutchan, T. M. Differential geneexpression in Papaver species in compari-son with alkaloid profiles. Acta Horti cul -turae.

Book ChapterWasternack, C. Jasmonates - Biosynthe sisand role in stress responses and de ve lop -mental processes. In Programmed CellDeath and Related Processes in Plants.(Noo den, L.D., ed.) Academic Press, NewYork, pp. 143-154.

Books and Book Chapters in pressKutchan, T. M., Frick, S. & Weid, M. En gi nee -ring plant alkaloid biosynthetic pathways -Progress and prospects. In: Ad van ces inPlant Biochemistry and Mole cular Biology(Le wis, N. & Nes, D. W., eds.) Vol. 1, Bio en -gi neering and Molecular Biology of PlantPathways (Bohnert, H.J. & Ngu yen, H.T.,eds.) Elsevier Science Ltd., Ox ford.

Wasternack, C. & Abel, S. Plant hormones.In: Molecular Plant Physiology. Chap ter 15,(Sharma, R., ed.) Harward Press, Bing ham -ton.

PatentKutchan, T. M., Frick, S. & Kempe, K. Mo du -lation of alkaloid biosynthesis in plants andplants having altered alkaloid bio synthesis. European Patent Application No. 04019154.6.

Diploma ThesesFreyer, Yvonne: Phänotypische Charak te ri -sierung einer Allenoxidcyclase-Knock out-Linie (AOC3-KO) von Arabidopsis tha liana.University of Halle-Wittenberg, Depart -ment of Biochemistry /Biotech no logy,24/11/2004.

Pienkny, Silke: Jasmonatbiosynthese durchAllenoxidcyclasen (AOCs) in Ara bidopsisthaliana -Unterdrückung der AOC2-Ex -pression durch RNA-Inter fe renz. Univer si -ty of Halle-Wittenberg, De partment ofBiochemistry/Biotech no logy, 17/09/2004.

Doctoral ThesesSchilling, Stephan: Charakterisierung derhumanen Glutaminylcyclase im Vergleichmit dem analogen Enzym aus Carica pa pa -ya. University of Halle-Wittenberg, De -partment of Biochemistry/Biotech no logy,05/05/2004.

Weid, Marion: Gewebe- und zellspezifischeLokalisation der Alkaloidbiosyn the se in Pa -paver somniferum L., University of Halle-Wittenberg, Department of Bio che mis -try/Biotechnology, 15/09/2004.


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Department of Bioorganic ChemistryHead: Ludger Wessjohann, ProfessorSecretary: Elisabeth Kaydamov

Department of

Bioorganic Chemistry

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Natural products play a key role in modern drug discovery. According to a recent survey, morethan 40% of all current trade drugs are natural products, or mostly derived or inspired by them,

although they constitute less than 1% of the compounds screened for activity.This higher probabilityof success in essence has evolutionary causes. By our research we try to understand the structuraland functional principles underlying this success, theoretically by chemo- and bioinformatic analysesand modelling, and in practice by isolation, characterization, modification, biosynthetic studies, andactivity screens of secondary metabolites, and to some extent of proteins involved in their conver-sion and interaction. The analytical work is backed by an extensive synthesis program, designed toincrease compound availability and molecular diversity by combinatorial chemistry, me thod develop-ment, and de novo synthesis. The principal compound classes covered are isoprenoids and lipids, phe-nolics and small cyclopept(o)ides. Applications of this research include the use of metabolites as leadstructures for drugs, cosmetics, or as research tools, and the use of enzymes as screening targets, oras catalysts for synthesis.

The department comprises four major research units covering the following topics:

Isolation (Plant and Fungal Metabolites) Isoprenoids (Biosynthesis and Biocatalysis, Modification) Synthesis (incl. Method Development, Biocatalysis) Computational Chemistry (Cheminformatics, Modeling)

These are backed by the analytical units (NMR, MS, Screening, Databases). An excellent intradepart-mental cooperation ensures the success of projects, which are not located in one organisationalgroup only, because they require-at different times-the whole range of expertise, from isolation tosynthesis and biosynthesis competence, accompanied by analytical and computational methods.Interdepartmental research further broadens the basis of such projects. Thus, with the Departmentof Stress and Developmental Biology, a group dedicated to the profiling of secondary metabolites(metabolomics) and proteomics from Arabidopsis and ra pe seed (GABI) is continuously successful.Overall, interdepartmental research has increased considerably (Tubulinbinders and Glycosyl trans -ferases with Prof. Strack, Terpencyclases with Prof. Kutchan, Bioinformatics and Metabolomics withProf. Scheel).

Two highlights of the year 2004 where the successful start of Prof. Bernhard Westermann as suc-cessor to Dr. Brunhilde Voigt as new group leader within the synthesis unit, and the inauguration ofBuilding R with state of the art laboratories for combinatorial synthesis, biocatalysis, and specia lizedrooms such as a night/stink lab, fermentation, GC-lab, DNA-sequencing etc. The laboratory sec tionsdedicated to our department were fully occupied and functional in less than four months.

In the report period, the department hosted researchers of 17 nationalities from four continents,e.g. regularly from Brazil,Vietnam, and Hungary. The international quota of medium term non-per-manent researchers (Ph.D.s /postdoctorals) was about 44%. 27% of the co-workers had a scholar-ship, e.g. from DAAD, CAPES, Daimler-Benz Foundation etc. Apart from diploma theses, six Ph.D.-titles were granted to co-workers, two of them with distinction marks. The department was suc-cessful in competitive DFG priority programs with five projects, graduate schools, EuropeanResearch Council funds, BMBF- and four EU-projects, several DAAD-programs, HWP-program, andwith industry financed projects.

The publication record was considerably increased. In the last two years, 69 contributions in re -viewed journals and books were published. But not only numbers increased, the publications werealso placed in higher valued journals (averaged sum of impact points: 2001: 134, 2002: 152, 2003: 206,2004: 292). The first patent applications based on IPB-research highlighted extremely potent anti-bacterial, antifungal and anticancer compounds.

The upcoming period will have to see improvements in the database-, IT- and screening facilities ofthe department as well as a further concentration and integration of research efforts along themajor lines outlined hereafter.

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Research Group: Plant and Fungal Metabolites/MicroanalyticsHeads: Norbert Arnold & Jürgen Schmidt

Floral Oils - Evolution,Analysis and Biological Significance

This project aims at the better under-standing of the chemical composition offloral oils, of their biosynthesis and evo-lutionary origin. The oils serve as at trac -tant for pollinators, for which they havedifferent functions, most prominently nu -tritional ones.The analysis of floral oils ofsix plant families (Orchidaceae, Malpi -ghiaceae, Krameriaceae, Iridaceae, Scro -phulariaceae and Solanaceae) re vealedacylated (3R)-hydroxy and dihydroxyla-ted fatty acids, in free form and / orbound as acylglycerols. The hydroxyfunction is always located at odd numbe-red carbon atoms. The substitution pat-terns of the hydroxylated fatty acids arein agreement with recently discove redhydroxylated aldehydes and alkyldiolsfrom epicuticular waxes. This fact sug-gests a close relationship between thebiosynthesis of plant epicuticular waxespossessing a protective function for theplant and floral oils. The microanalyticalmethods were refined further, based onGC/EI-TOFMS of different derivatives(e.g. methyl es ters, trimethylsilyl estersor ethers, pyrrolidides, dimethyldisulfideadducts) and ESI-MS(/MS) of the non-derivatized se cretions. This is used toextend the species basis. We could alsorevise that previous reports onOncidinae flower oils (Orchidaceae).

Constituents of Higher Fungi

Fungi probably comprise the largest group of higherorganisms in our world with estimated 106 speci-men. Therefore, fungi are an important resource ofnew metabolites. Our research is mainly focused onthe isola tion and characterization of fungal metabo-

lites from collected fruit bodies of higher fungi (Basi -diomycetes), preferentially mycorrhizal ones. In sim-ple bioassays, the isolated substances are tested forfungicidal and bactericidal activity.

Species in Hygrophorus are rarely attacked by para-sitic fungi, insects, or snails. 20 new 5-(hydroxyalkyl)-2-cyclopentenone derivatives (hygrophorones) couldbe isolated from Hygrophorus latitabundus, H. oliva-ceoalbus, H. persoonii, and H. pustulatus. Thesehygrophorones have structural similarities to theantibiotic pentenomycin. Chemically, hygrophoronesare 2-cy clo pentenones with hydroxy or acetoxysubstituents at C-4 and/or C-5. An odd-numbered1'oxidized alkyl chain (C11, C13, C15, or C17) is attachedat C-5. In addition, from H. persoonii the new g-buty-rolactone derivative (5-(E)-2-hydroxytetradexylide-ne-5H-fu ran-2-one) could be isolated. Some hygro-phorones are responsible for the color reaction ofthe stipes of these fungi upon treatment with potas-sium hy droxide solution. In our bioassays, we coulddemonstrate fungicidal activity as well as a remarka-ble bactericidal activity against gram-positive bacte-ria. Strains of multiresistant Staphylococcus aureus(MRSA) and vancomycin-resistant Enterococcus fae-cium (VRE) were inhibited in low concentration. Apa tent has been filed.

From fruit bodies of the basidiomyceteHygrophorus eburneus (Tricholomataceae) eightfatty acids (C16, C18) with g-oxocrotonate partialstructure could be isolated. Initial tests demonstratetheir significant bactericidal and fungicidal activity.

Cortinarius bolaris (subgenusLeprocybe) is described in theliterature as poisonous.The capand stem staining yellow whenbruised. The underlying prin -ciple likely is a new benzofuranglycoside, which can be isola-

The isolation and structure elucidation of plant and fungal metabolites with modern analy-tical techniques is the basis for investigations of both the native function of natural productsas well as for applications. The evolutionary selection undergone by these compounds,favour them as leads for drug candidates as well as for the identification of new targets,which might be derived from un derstanding the native function. Also, ecological relationsor new enzymatic transformations can be deducted from the knowledge of an organism’sconstituents.

Hygrophorus olivaceoalbus, theShea thed Waxy Cap is growing inthe Harz mountains.

Group Members

Sanela Bacinovic Diploma Student since November2004

Danston BarazaNAPRECA-DAAD Fellow untilNovember 2004

Kanchana Dumri DAAD-Leibniz Fellow since March2004

Katrin Franke Postdoctoral Position

Gudrun Hahn Technician

Myint Myint Khine D.-B. Fellow

Christine Kuhnt Technician

Monika Kummer Technician

Martina Lerbs Technician

Tilo Lübken PhD Student

Jana Mühlenberg PhD Student until December 2003

Lars Seipold PhD Student until May 2004

Axel Teichert Diploma Student until May 2004

Nguyen Hong Thi Van Visiting Scientist until April 2003

Alexander Voss Diploma Student since December2004

Collaborators

Joe AmmiratiUniversity of Washington, USA

Helmut BeslUniversity of Regensburg, Germany

Günter Gerlach Botanical Garden Munich, Germany

Lutz HeideUniversity of Tübingen, Germany

Matthias H. Hoffmann Botanical Garden Halle, Germany

Christoffel SpenglerBionorica AG Neumarkt, Germany

Wolfgang Steglich University of Munich, Germany

Tran Van Sung Institute of Chemistry Hanoi, Vietnam

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ted from freeze-dried samples. This compoundturns yellow after exposure to HCl or upon heating.The stereochemistry could not yet be established.Surprisingly, fresh specimen of C. bolaris do notcontain the benzofuran glycoside. Instead, a new yel-low-colored phthalimide derivative exhibiting abright blue fluorescence appears. Its biosynthesis isassumed to originate from a benzofuran glycoside.

Constituents of traditional medicinal plants

HEATOS - A Vietnamese opiate detoxification sym-ptom medicationThe project HEATOS was originally coordinated bythe United Nations (UNESCO and UNOPS). Dr. Danfrom Vietnam developed the original herbal aid foropioid detoxification and called it HEATOS (heat ofsun). HEATOS consists of ca. 13 components and wasconstantly improved in composition, also with the aidof our findings. In cooperation with Vietnamese part-ners our research is focused on the isolation and cha-racterization of the constituents of these plants.Most species have been identified. So far some 180compounds are isolated from eight plants. A sub-stance based overview and extensive literature sur-vey was conducted.

Traditional South East Asian medicinal plantsThe aim of this project is to isolate and characte-rize the phytoconstituents of the medicinal plantsStreptocaulon tomentosum, Vitis repens, Curcumacomosa, Aristolochia tagala and Spermacoce hispidafrom Myanmar, and to test their bioactivity.

The roots of Streptocaulon species are used medici -nally for the treatment of dysentery and stomach -ache, and the leaves are used externally for thetreatment of snake poisoning and abscesses. Cur cu -ma comosa is a member of the economically impor-tant plant family Zingiberaceae. The extract of C.co mosa is reported to be nematocidal and cholere-tic in rats. As well as Aristolochia tagala this speciesis used against malaria. Rhizomes of Vitis repens areused against different cancers. Spermacoce hispida isan effective natural drug by treatment of hyperten-sion.

From S. tomentosum several triterpenoids, cardeno-lides, and oleanane-type saponins have been identi-fied. A new cardenolide (1) was isolated from theroots. Some cardenolides had remarkable an tipro life -

rative ac tivity pro -files. Apo lar ex tractsof three plants (C.co mo sa, A. tagala, V.re pens) also showsignificant antifungalactivities.

Microanalytics

In collaboration with the University of Halle themass spectral fragmentation of morphine and co -deine was studied indetail using LC-electrospray MS/MSand high resolutionFT-ICR-MS combinedwith an IRMPD. La -beled codeine wasisolated and pu rifiedfrom Papaver somni-ferum seedlings, which were grown in the presence of[ring-13C6]-L-tyrosine, [ring-13C6]-tyramine and [1,2-13C2], [6-O-methyl 13C]-(R,S)-coclaurine. Using theselabeled derivatives it was possible todetermine the origin of the carbonsin the fragment ions. In collabora -tion with the Department of Na -tural Pro duct Biotechnology and theBiocenter of the Uni versity the LC-ESI-MS/MS investigations of morphi-nan-type and benzylisoquinoline al -kaloids are continued.

Furthermore, a se ries of new aminocoumarin anti-biotics (2) produced in vi tro and in vivo by a combi-ned biochemical, genetic and synthetic ap proach froma coo pera tion of the NWC-Isoprenoids Group andthe University of Tübingen were structurally elucida-ted by using LC-ESI-SRM both under positive andnegative ionization.

The high-resolution FT-ICR-MS was successfully ap -plied for the determination of the exact mass mea-surements of rifamycin derivatives including theirfragments obtained by varying the capillary exit vol-tage (3).

In addition, the group is intensely involved in themetabolic profiling and IT-projects at the IPB.

1

2

3

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Research Group: Structural Analysis & Computational ChemistryHeads: Wolfgang Brandt & Andrea Porzel

Cheminformatics and NMR

Analysis of macrocycles abundant in natural pro-ductsA database containing the structures ofmore than 120,000 compounds was ana-lyzed and data mining was performed formacrocycles with more than 13 atomsin unbridged rings. The resulting data -base was analyzed for molecular weightdistribution, the frequency of commonsubstructural motifs, and biosyntheticorigin in relation to ring size.The underly -ing principles of chemical diversity werereviewed in terms na ture's strategies fordiversity and complexity generation andwith respect to biosynthetic origin. Fi -nally, it was suggested that synthetic che-mists should not only use nature's mole-cules, but also its strategies as a sourceof inspiration. To illustrate this, the bio-synthesis of macrocycles by terpene andpolyketide cyclases and nonribosomal

peptide synthetases, as well as recent advances inemploying these strategies in an integrated synthe-sis /biotechnology approach were briefly reviewed.

In house database, cheminformatic tools and spectros-copyIn collaboration withVolkmar Vill, Univer sityof Hamburg, the deve-lopment of the in housedatabase project Phyto -base has been continued.Se ve ral hundred struc-tural formulas have beenadded and in put /outputprogram options andsearch tools were im -proved in order to de -sign a user-friendlier sur -

face.In continuation of our cheminformatic activities, adatabase of more than 500 experimental as well ascalculated 1H/13C 1J shift correlated 2D NMR spectrawas designed. In collaboration with the Institute ofComputer Science, University of Halle, a databasemanagement system was developed which providessimilarity search and mixture analyses based on the2D NMR database content.

Modern NMR spectroscopy experiments were usedfor the structural elucidation of bioactive naturalproducts isolated from plants and higher fungi. Also,the synthetic studies in the department were sup-ported by NMR investigations. Routine spectra(1H/13C/19F/31P NMR, IR, CD, ORD, UV) were recor-ded as a service for the Department of BioorganicChemistry and all research groups of the IPB.

Further steps towards an unified ACD-based plat-form of spectroscopy data were taken and the IT-coordination within the IPB was improved.

Modelling and Quantum Mechanical Calcu lations

The functional role of selenocysteine (Sec) in thecatalysis mechanism of large thioredoxin reductasesWe investigated the redox mechanism of thiored -oxin reductases containing a selenocysteine in theca talytically active site. Thioredoxin reductases ca -ta lyze the reduction of thioredoxin disulfide andsome other oxidized cell constituents. They are ho -modimeric proteins containing one FAD and accep-ting one NADPH per subunit as essential cofactors.Ba sed on the X-ray structure of rat thioredoxin re -ductase, homology models of human thioredoxinre ductase were created, and subsequently dockedto thioredoxin to model the active complex.

The formation of a new type of a swapping catalytictriad between selenocysteine (Sec), histidine, and a

Group Members

Susanne AustGuest Research Scientist since March2003

Monika BögelResearch Scientist until December2003

Lars BräuerPhD Student

Frank BrodaSystem Administrator since January2004

Stefanie GuldeDiploma Student since October 2004

Jana Selent PhD Student, Guest, until April 2004

Annett SiebenhünerDiploma Student since November2004

Maritta SüßeTechnician

Bianca Wachsmuth Diploma Student until December 2004

The research group is investigating three-dimensional molecular structures of small molecules pro-teins and reaction mechanisms (e.g. enzyme catalysis) in the field of bioorganic chemistry by means ofmolecular modelling techniques, semi-empirical and ab initio calculations, bio- and cheminformatics,theoretical chemistry and NMR spectroscopy.The group is also involved in the development of mole-cular databases. The collected information together with data mining and new data from the otherresearch groups form the basis for cheminformatic analyses which enables new insights into the bio-logical significance of plant and fungal metabolites, and also in drug design.

Figure1:The catalytically active site of thioredoxin reductasein complex with the substrate thioredoxin. The red arrowsindicate the proton transfer from Sec498 to His472 forming thecatalytic triad together with Glu477. Carbon atoms of thesubstrate are colored in magenta.

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glutamate could be detected in the protein struc -ture. By means of DFT calculations we could showthat the formation of such a triad is essential to sup-port the proton transfer from selenol to a histidineto stabilize a selenolate anion, which is able to inter-act with the disulfide of thioredoxin and catalysesthe reductive disulfide opening (Fig. 1).

Whereas a simple proton transfer from seleno cys -teine to histidine is thermodynamically disfa vo red itbecomes favored when the carboxylic acid group ofa glutamate stabilizes the formed imida zole cation.An identical process with a cysteine instead of sele-nocysteine will require 4 kcal/mol more energy,which corresponds to a calculated equilibrium shiftof ~ 1000:1 or a 103 rate acceleration a value closeto the experimental one of about 102-times.

These results give new insights into the catalysismechanism of thioredoxin reductase and for thefirst time explain the advantage of the incorpora tionof a selenocysteine instead of a cysteine residue in aprotein.

The modeled protein structures of both the cytoso-lic and mitochondrial thioredoxin reductase in com-plex with thioredoxin have been accepted by andstored in the Protein Data Bank.

Protein 3D-homology modeling of UDP-glucosedependent betanidin 5-O-glucosyl-transferase fromDorothean thus bellidiformisThis project was based on a close cooperation withthe Department of Secondary Metabolism of ourinstitute. The betanidin 5-O-glucosyltransferase(UGT73A5) from Livingstone daisy(Dorotheanthus bellidiformis) is involved in regio-specific glucosyla tion of betanidin and various fla-vonols. A three -dimensional model of this regio-specific betanidin and flavo noid glucosyltransferasehas been constructed and the active site has beenmodeled (Fig. 2). To explain the observed inversionin the configuration of the bound sugar, semiempi-rical calculations favor an SN1-type reaction as aplausible alternative to the generally proposed SN2-mechanisms discussed for plant glucosyltransfera-ses. The calculated structural data do not onlyexplain the ab straction of a proton from the ac -ceptor be tanidin, but also further imply that the re -action mechanism may involve a ca talytic triad with

si milarities to the serine proteasefa mily. Selected amino acids suggested tobe involved in substrate binding andturnover, were substituted via site-directed mutagenesis of the functio-nally expressed protein. Substi tu -tion of two highly conserved aminoacids, Glu378, located in the proposedUDP-glucose binding site, and His22,located close to the N-terminus,both led to a complete loss of en -zyme activity, which considerablysupports the principal correctnessof the model.

Prenyltransferases

A main project of the department isthe investiga tion of C-C-coupling prenyltransfera-ses, located in the research group Isoprenoids. Thiswork is ac com pa nied by bioinformatic studies,homology mo delling, and quan tum mechanical calcu-lations. Com putational chemistry work on p-hydroxy-benzoate oligoprenyltransferase (ubi a) has been finis-hed for the mo ment, and re sults were pub lished.Recently, homology protein modeling of other prenyl-transferases of other families was started.

Collaborators

APOGEPHA GmbHDresden, Germany

Horst BögelUniversity of Halle, Germany

Susanna FürstUniversity of Budapest, Hungary

Andris KreicbergsKarolinska Institute, Stockholm, Sweden

ProBioDrug AGHalle, Germany

Ryszard PrzewlockiPolish Academy of Sciences, Krakow,Poland

Helmut SchmidhammerUniversity of Innsbruck, Austria

Volkmar VillUniversity of Hamburg, Germany

Wella AGDarmstadt, Germany

Figure 2: Stereo representation of the active site of the UGT73A5 from D. bellidiformis with beta-nidin (magenta highlighted). Amino acids proposed to be involved in substrate binding and cata-lytic sugar transfer are displayed. Red-labeled residues belong to the PSPG-box, magenta ones areessential for the catalysis and blue labeled ones for binding of the ligands. Hydrogen bonds arerepresented by green dotted lines. The red arrow indicates the initial step of the suggested cata-lysis mechanism.

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Research Group: IsoprenoidsHead: Ludger Wessjohann

Synthesis of isoprenoid substrates, inhibitors andnatural products

A large set of oligoprenyl diphos -phates was synthesized as substratesand discovery tools, especially non-natural ones like homo-, nor- or hete-roderivatives, or dye labeled ones, toprobe substrate specificity. Interme -diates to the non-mevalonate MEP-pathway, e.g. epoxidized and hydroxy-lated isopentenyl and dimethylallyldi-phosphate (DMAPP), were synthe -sized in iso tope labeled form. Furtherchain-mo dified derivatives were syn-thesized as part of a biocatalytic totalsynthesis program to wards prenyla-ted aromatic natural products.

In addition, several farnesyl and geranyl analo-gues with mimics of the pyrophosphate unithave been prepared for an examination of theirability to in hibit prenyl transfer. 7,11-Dimethyl-3-oxododeca-6,10-dienoic acid, 3-hydroxy-7,11-

dimethyldodeca-6,10-dienoic acid,2-hydroxy-4,8-dimethyl-3,7-nona-dienylphosphonic acid, and [[(4E)-5,9-dimethyldeca-4,8-dienyl] phos -phina to](difluo ro) me thyl phospho -nate were synthesized from gera-niol. w-2,w-1-dihydroxy lated farnesyldiphosphate was prepared fromtrans,trans-farnesol.

8-Prenylnaringenin (8-PN, 1) is thestrongest natural phytoestrogen wit-hin the flavonoids. Recently the com-

pound triggered considerable commer cial inte-rest as mild menopause medication. However, thetotal synthesis is lengthy, complicated and expen-sive. Also, totally "artificial" synthetic compoundsare less acceptable to the anticipated customerbase. Based on the abundant na tural hop com-pound xanthohumol, we could de vise a simple,protection group free two step synthesis of 8-PNin over 90% yield, in which the natural carbonisotope distribution is re tained 100%. The mostdifficult task was the se lective cleavage of a me -thyl ether in the presence of the acid sensitiveprenyl unit, which so far was only achieved with afew percent yield and conside rable byproductformation.

ubi A Prenyltransferase4-Hydroxybenzoate oligoprenyltransferase of E.coli, encoded in the gene ubiA, is an importantkey enzyme in the biosynthetic pathway toubiqui none, an essential component of aerobiclife. It catalyzes the prenylation of 4-hydroxyben-zoic acid in the 3-position using an oligoprenyldiphosphate as second substrate. An X-ray struc-ture is not available, not even of a related enzy-me. The knowledge of the tertiary structure andpossible active sites is, however, essential for theunderstanding of the catalysis mechanism and thesubstrate specificity.

The substrate model known for the aromaticpart was extended. For the first time a minima -lized model for the prenyl substrate could bedefined based on experiments with increasinglytruncated GGPP-, FPP-, and GPP-derivatives (G =Geranyl, F = Farnesyl).

Group Members

Lars BräuerPhD Student

Marco DessoyPhD Student until July 2003

Michael Fulhorst PhD Student until September 2003

Gudrun HahnTechnician

Roman WeberPhD Student since September 2004

Heike WilhelmPostdoctoral Position since August2003

Svetlana ZakharovaPostdoctoral Position

Isoprenoid units can be detected in some 70,000 natural products, including steroids, terpenoids andconjugates with molecules of other major pathways. The study of isoprenoid compounds and meta-bolism provides insight into the predominant mechanisms and routes that nature uses to build up car-bon skeletons. Understanding these, will provide new enzymes for in vitro C-C-coupling reactions asbiocatalysts for the production of prenylated and terpenoid compounds, as well as new targets forinhibitors of important metabolic processes in plants, most pathogenic bacteria, and many parasites.Of special interest to us is the transfer of prenyldiphosphates onto sp2-carbons (aromatic and vinylicsubstrates). We hope to elucidate mechanistic and structural details, provide better access to probesand substrates, develop mechanism-based inhibitors and finally achieve access to a set of enzymesenabling a multitude of enzymatic C-C-coupling reactions. In addition, synthetic modifications of iso-prenoids and diphosphates to increase the compound base are part of the program.

8-Prenylnaringenin (8-PN) is the strongestnatural phytoestrogen within the flavonoids.

1

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With respect to mechanistic studies, throughmodeling techniques, secondary structure pre-diction tools, molecular dynamics simulations,and energy optimizations, an acceptable homolo-gy model could be created. Multiple alignments ofa multitude of related proteins clearly showed100% conservation of the amino acid residuesforming the putative active site. An additionalhighly conserved region could be detected, whichcurrently is considered to be a rudimentary ac -tive site, giving some hints regarding the evolutio-nary origin of ubiA-enzyme. Semi-empiric quan-tum mechanical PM3 calculations have been per-formed to investigate the thermodynamics andkinetics of the catalysis mechanism. These resultssuggest a mechanism closer to SN1- than to SN2-type for the cleavage of the diphosphate ion fromthe isoprenyl unit. Most interesting is the depro -tonation behavior of enzyme bound 4-hydroxy-benzoic acid, of which formation of the pheno lateanion is favored to that of a benzoate anion. Thisideally aids the attack of the isoprenyl cation,which appeared to be the rate-limiting step ofthe whole process according to quantum chemi-cal calculations. Probing this model by site direc-ted mutagenesis failed so far, it proved extremelydifficult to obtain functional mutated protein.

Finally, several analogues and mimics of farnesyland geranyl pyrophosphate were tested for en -zyme inhibition in a competitive assay with GPP.The effect of these compounds on ubiA-prenyl-transferase activity varied substantially, rangingfrom almost full competitive inhibition to, surpri-singly, enhanced enzymatic activity at low concen-trations by some compounds. The activity enhan-cing activity of some diphosphate mimics couldbe related to their magnesium ion complexingproperties and is only effective at high ion/ lowenzyme concentration. It can be mimicked byEDTA-addition.

Non-mevalonate (dxp- or mep-) Pathway in PlantsExperimental and theoretical investigations wereperformed concerning the two last steps of thedxp/mep-pathway in isoprenoid biosynthesis inplants. The proposed intrinsic or late interme -diates, 4-oxo- and 4-hydroxy-DMAPP were syn-thesized in deuterium or tritium labeled form ac -

cording to new protocol especial-ly adapted to work without pro-tection of the diphosphate moie-ty. In cooperation with Prof. Mein -hart Zenk, these as well as the la -beled cyc lic precursor MEcPPwere ap plied to chromoplast cul-tures. For the first time, 4-hydro-xy-DMAPP was identified as thelast intermediate towards DMAPPand IPP in plastids of higherplants: Its isotope labeled formwas incorporated into the caro -tene precursor phytoene in vitro.

This finding is in agreement withour mechanistic and structuralmodel of the fragment L271 to A375 of the enzymeGcpE of Streptomy ces coelicolor (transformingMEcPP to 4-hydroxy-DMAPP), including NADPH,the Fe4S4-cluster and MEcPP as ligand. Based onthis homology model, semi-empirical PM3 cal -cula tions were used to analyze the likely catalysisme chanism of the reductive ring opening ofMEcPP. Our suggested mechanism is characte -rized by a proton transfer (presumably from theconserved arginine 286) to the substrate, accom-panied by a ring opening without high energy bar-riers, followed by the transfer of two electronsdelivered from the Fe4S4-cluster and finally pro-ton transfer from a carboxylic acid side chain tothe hydroxyl group to be removed from the li -gand as water. The proposed mechanism is inagreement with all known experimental findingsand the arrangement of the ligand within theputative enzyme model. It differs especially fromproposals suggesting epoxide intermedia-tes which, however, are highly unfavorablein energy and could not be supported bytheoretical or by experimental data. Aprincipally si milar mechanism is also ex -pected for the reductive dehydroxylationof 4-hydroxy-DMAPP in the last step ofthis basic isoprenoid pathway.

Collaborators

Martin BiendlHallertauer Hopfenveredelungs GmbH,Mainburg, Germany

Uwe BornscheuerUniversity of Greifswald

Lutz Heide, Shuming LiUniversity of Tübingen Germany

György HorvathUniversity of Antwer, Belgium

Toni M. Kutchan, Jon PageIPB Halle, Germany

Markus Pietzsch, Meinhart ZenkUniversity of Halle, Germany

SunGene GmbH & Co. KGaAGatersleben, Germany

Hop cones

MacrocyclesEven functionally very simple mole -cules like host-molecules already re -quire considerable size and design.Thus, in order to minimize entropyloss upon binding, a reduction of con-formational freedom compared to li -near mo lecules is required. At thesame time, some flexibility needs tobe retained in order to allow adap tiveproperties or different stages within afunctional process, mostly a bindingprocess. This can be achieved by self -organization and folding (as in en -zymes), rigidifying elements (e.g. turninducers), or (macro-)cyclization. In na -ture, selec tive hosts or ligands of a MW-range below that of enzymes or me -

dium peptides often are macrocycles with rigidi-fying and flexible elements placed in the appro-priate positions, often of polyketide or peptideori gin. Another important element for nature-like selection and interaction properties, recogni-tion, and function is asymmetry. Besides asymme-try, the overall functional group density and varia-bility distinguishes most natural hosts from e.g.crown ethers or calixarenes, which are rather ru -dimentary hosts from a biomimetic viewpointand in terms of selectivity.

Thus macrocycles, ideally even with adjustableconformational bias, are good candidates to mi -mic natural binders in a minimal fashion, e.g. toachieve hosts selective for binding in water, or forthe design of protein-protein-interaction tools.Synthetic approaches thus must have the flexibi-lity to introduce a large variety of structural ele-ments of rigidity, flexibility, lipophilicity, and direc-tional functionality.

Access to libraries of such macrocycles requiresthe development of efficient, flexible, diversity-oriented synthesis routes. This still is a big chal-lenge for organic synthesis because of the excep-tionally stringent requirements for reaction effi-ciency, chemoselectivity, functional group tole-rance, and stereoselectivity. Thus, diversity-orien -ted synthesis also provides an important en ginefor innovation in organic chemistry. Natural pro-duct like macrocycles can combine rigidity andflexibility at the same time, contradicting the pa -radigm that compounds of pharmaceutical in te -rest have to be flat, and nitrogen containing.Therefore, work in the group has been largely fo -cussed on the target- and diversity-oriented syn-thesis of macrocycles and the development ofnew macrocyclization reactions.

In order to obtain compounds of high biological relevance, the most recent developmentin organic synthesis favors the combination of (natural product) target -oriented and diver-sity -oriented synthesis. The synthetic group is ideally placed in this context. Efforts areundertaken to synthesize small natural products (i. a. such as provided by the Plant andFungal Metabolites Group) and mimics thereof, focussed natural product-like libraries, anddesigner mole cules. The biggest problem with respect to natural product like and functio-nal structures is their complexity, which usually requires complicated multistep syntheses.New synthetic tools, methods and concepts to improve the speed and selectivity of syn-thetic reactions are the basis to obtain the desired chemical diversity of modified naturalproducts or mimetics with improved or even new biological activity profiles, e. g., by multicomponent reaction (MCR) strategies. One focus is on macrocyclic compounds, whichcombine conformational preorganization with flexibility. Combinatorial approaches inliquid as well as on solid phase have been used to obtain higher selectivity, improve reac-tion conditions, and to form small focussed libraries demonstrating the power of our pro-cedures. Applications in the chemical field include reactions in water, selective separation,and catalysis; in the pharmaceutical field, compounds with anticancer, antibiotic, phytoestro-genic and cosmetic properties have been studied.

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Research Group: Synthesis and Method DevelopmentHeads: Ludger Wessjohann & Bernhard Westermann

Group Members

Muhammad AbbasPostdoctoral Position since May 2004

John Bethke Postdoctoral Position until April 2004

Viktor DickPhD Student since April 2004

Simon DörnerPhD Student since April 2004

Tobias DrägerPhD Student since October 2004

Uwe EichelbergerPostdoctoral Position

Daniel Garcia-RiveraPhD Student since October 2003

Gergely GulyasPhD Student since August 2004

Alexander GutscheDiploma Student since August 2004

Nicole HüneckeTrainee since August 2004

Oliver KreyePhD Student since July 2004

Christiane NeuhausPhD Student since April 2004

Eelco RuijterPhD Student until June 2004

Angela SchaksTechnician

Gisela SchmidtTechnician

Alex SchneiderPhD Student since June 2004

Henri SchrekkerPhD Student until February 2004

Tran Thi Phuong ThaoPhD Student

Tran Van ChienPhD Student until September 2003

Katharina WolfTrainee since August 2004

Mingzhao ZhuPhD Student

Target-oriented synthesis of macrocyclesEpothilones have pronounced biological profiles,which make them interesting drugs for cancertreatment now explored in phase II / III clinical tri-als. All four diastereomers of the new epothiloneD5-series, e.g. isomer 1, were synthesized, andcompared to the natural isolate. In this context,the chromium-mediated Reformatsky reactiondeveloped by us proved to be the method ofchoice for the construction of the southern part.

In order to produce third generation epothiloneswith enhanced accessibility and pharmacologicalproperties, thio-derived congeners of epothiloneD lacking one stereocenter, like in compound 2,have been synthesized. They showed low nano-molar activity against some cancer cell lines andpatent applications have been filed.

The compounds synthesized proved useful in thestudy of plant cell cycles in cooperation withBettina Hause in the Department of SecondaryMetabolism.

Diversity-oriented synthesis of macrocyclesFor the synthesis of macrocycle libraries we devi-sed three strategies. Strategy I utilizes commercially available ma -crocycles like rifamycin, tylosin or erythromycin.These are modified by selective reactions, inclu-ding biocatalytic ones in solution or on solidphase, to produce libraries of rim-modified deri-vatives, usually without any protective group che-mistry. This allows a fast chemical screen for ac -cessibility and biologically importance of rapidlymodifiable positions. A new enzyme penetrableresin-linker combination was designed for thispurpose.

29

Scheme 1: Ugi-type MiB combining a steroid and biarylether moiety (MiB = multiple multicomponent macrocyclization inclu-ding bifunctional building blocks)

1

2

30

Research Group: Synthesis and Method DevelopmentHeads: Ludger Wessjohann & Bernhard Westermann

Strategy II utilizes multicomponentreactions (MCRs) like the Ugi-reaction (Scheme 1), which isextremely atom-economic andprovides ra pid access to highly di -verse peptoid compounds. Utili -zing specially designed bifunctionalstarting ma terials, the synthesis ofmacro cycles can be achieved in aone step multi reaction procedure(MiBs = Multiple MulticomponentMacrocyclization including Bifunc -tional Building Blocks). Initially, wehave concentrated on the synthe-sis of steroid- and (vancomycin in -spired) biarylether containing pro-ducts like compounds 3 and 4. E.g.,some MiBs in a one-pot re actioncombine twelfe building blocks as -sembled in 16 reactions to give aconstitutionally defined 60-mem-bered macrocycle with water as theonly formal byproduct. For the firsttime, complex macrocycles withnon-identical subunits (cf. the repe-titive calixarenes and cyclodex -trines) become available in an ex -tremely efficient combinatorial fa -

shion in infinite number and quantity with almostunlimited size, shape, and functionality, thus ope-ning the door to a much improved host-guest che-mistry, organocatalysis and artificial enzymes, cy -clopept(o)id mimics, and protein-protein interac-tion studies.

Strategy III is a combination of a linear precursorsynthesis and a cyclizing reaction like the Grubbs-olefin metathesis and, e.g., the azide/al kyne-"click"-approach to achieve the synthesis of car-bohydrate moiety containing macrocycles as in 5.Such compounds are of particular interest due totheir resemblance of aminoglycosides, a class ofnatural compounds exhibiting antibiotic activity.Method development

In the report period, several selective methodsutilizing, e.g., biocatalysis towards asymmetricacyloins and novel hetero-Diels-Alder reactionsto form polyketide fragments, Ugi-MCRs, transi -tion metals and transition metal catalysts [Cr(II),Co, Pd] have been developed and utilized in thesyntheses discussed above. Selenium-based andorganocatalytic processes are a more current de -velopment.

Selenium chemistrySelenium compounds are little studied in chemi-stry, especially in combination with their catalyticand biological significance. We set out to studythe properties of selenium derivatives in compa-rison to the usually well-studied sulfur analogues,with a special focus on selenocystein (Sec) andSec-containing peptides (cf. also the ResearchGroup of Computational Chemistry). A reason -able study should be performed under (close to)physiological conditions and not be limited to asingle compound. This required a synthetic route

Collaborators

Uwe Bornscheuer University of Greifswald, Germany

Antonio Luiz BragaFederal University of Santa Maria, Brazil

Alexander Dömling, Wolfgang Richter R&D Biopharmaceuticals GmbH Munich,Germany

Sabine FlitschUniversity of Edinburgh, UK

Lucia GardossiUniversity of Trieste, Italy

Bettina HauseIPB Halle, Germany

Udo KraglUniversity of Rostock, Germany

Rob Leurs, Martine SmitFree University of Amsterdam,Netherlands

Francisco Coll ManchadoUniversity of Havanna, Cuba

Karoly MicskeiUniversity of Debrecen, Hungary

MorphoChem AG MunichMunich, Germany

Tamas PatonayUniversity of Debrecen, Hungary

Romano OrruFree University of Amsterdam,Netherlands

Polymer Laboratories Ltd Shropshire, UK

3

4

5

31

to libraries of (water soluble) Sec-peptide deriva-tives. In principle, the Ugi-MCR can be utilized forthis purpose again, but a difficult adaptation to in -corporate the initially problematic selenium con-taining building blocks had to be made.With theright combination of solvent, catalyst, and buildingblocks, we are now able to produce libraries ofselenopept(o)ids in water, including, e.g., com-pound 6 with the biologically important Cys-Sec-moiety. Furthermore, new syntheses of seleno-and tellurocystein and related sulfur, seleniumand tellurium compounds were devised and uti -lized, e.g., for catalysis.

Organocatalysis Organocatalysis with proline 7, offers the possibi-lity to carry out asymmetric Mannich reactions

leading to the appropriate Mannich type bases,like 8, in high yields and selectivity. While usingprotected dihydroxy acetone derivatives, this re -action can be seen as a surrogate of aldolase ca -talyzed reactions found in nature. Or gano cata -lytic processes not only provide the products inhigh stereoselectivity, they are also "green pro-cesses" due to the absence of any me tal ion. Inaddition, a couple of these reactions can be car-ried out in aqueous solutions decreasing theamount of organic solvents.

6 7 8

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PublicationsBethke, J., Karaghiosoff, K. & Wessjohann,L. A. Synthesis of N,N-disubstituted sele-noamides by O/Se-exchange with sele -nium-Lawesson's reagent. TetrahedronLett. 44, 6911-6913.

Braga, L. A., Paixao, M. W., Lüdtke, D. S.,Silveira, C. C. & Rodrigues, O. E. D. Syn the -sis of new chiral aliphatic amino disele -nides and their application as catalysts forthe enantioselective addition of diethylzincto aldehydes. Org. Lett. 5 (15), 2635-2638.

Braga, A. L., Rubim, R. M., Schrekker, H. S.,Wessjohann, L. A., de Bolster, M. W. , Zeni,G. & Sehnem, J. A. The facile synthesis ofchiral oxazoline for the diethylzinc addi-tion to aldehydes. Tetrahedron Asymm. 14,3291-3295.

Black, S. L., Jales, A. R., Brandt, W., Lewis, J.W. & Husbands, S. M. The role of the sidechain in determining relative d- and k-affi-nity in C5'-substituted analogues of nal tr in -dole. J. Med. Chem. 46, 314-317.

Cacace, S., Schröder, G., Wehinger, E.,Strack, D., Schmidt, J. & Schröder, J. A fla-vonol O-methyltransferase fromCatharan thus roseus performing twosequential me thylations. Phytochemistry62, 127-137.

Degenkolb, T., Berg, A., Gams, W., Schlegel,B. & Gräfe, U. The occurrence of pep tai -bols and structurally related peptaibioticsin fungi and their mass spectrometric iden-tification via diagnostic fragment ions. J.Peptide Sci. 9, 666-678.

Eckermann, C., Schröder, G., Eckermann,S., Strack, D., Schmidt, J., Schneider, B. &Schröder, J. Stilbenecarboxylate biosynthe-sis: A new function in the family of chal -cone synthase related proteins. Phyto che -mis try 62, 271-286.

Gidda, S. K., Miersch, O., Levitin, A., Schmidt,J., Wasternack, C. & Varin, L. Bio chemical andmolecular characterization of a hydroxyjas-monate sulfotransferase from Arabidopsisthaliana. J. Biol. Chem. 278, 17895-17900.

Holzgrabe, U. & Brandt, W. Mechanism ofaction of the diazabicyclononanone-typek-Agonists. J. Med. Chem. 46, 1383-1389.

Ibdah, M., Xing-Hai Zhang, J. Schmidt, J. &Vogt, T. A novel Mg++-dependent O-me thyl -transferase in the phenylpropanoid meta-bolism of Mesembryanthemum crystalli-num. J. Biol. Chem. 278, 43961- 43972.

Jarry, H., Spengler, B., Porzel, A., Schmidt, J.,Wuttke, W. & Christoffel, V. Evidence for

estrogen b-selective activity of Vitexagnuscastus and isolated constituents. PlantaMed. 69, 947-950.

Kamperdick, C., Nguyen Minh Phuong,Adam, G. & Tran Van Sung Guaiane dimersfrom Xylopia vielana. Phytochemistry 64,811-816.

Kolbe, A., Porzel, A., Schmidt, J. & Adam, G.A new synthesis of [26,28-2H6] brassino -lide and [26,28-2H6] castasterone via anun usual methyl migration. J. Label Compd.Radiopharm. 46, 231-242.

Kuhn-Wache, K., Hoffmann, T., Manhart, S.,Brandt, W. & Demuth, H. U. The specificityof DP IV for natural substrates is peptidestructure determined. Adv. Exp. Med. Biol.24, 57-63.

Lorey, S., Stöckel-Maschek, A., Faust, J.,Brandt, W., Stiebitz, B., Gorrell, M. D., Käh -ne, T., Mrestani-Klaus, C., Wrenger, S., Rein -hold, D., Ansorge, S. & Neubert, K. Dif fe -rent modes of dipeptidyl peptidase IV(CD26) inhibition by oligopeptides de -rived from the N-terminus of HIV-1 Tatindicate at least two inhibitor binding sites.EJB 270, 2147-2156.

Luu Duc Huy & Porzel, A. Heptazoline - acarbazol alkaloid extracted from Clausenaheptaphilla (Roxb.) W. & ARN. Tap ChiDuoc Hoc (Pharmaceutical Journal of VietNam), 11, 11-12.

Mrestani-Klaus, C., Brandt, W., Faust, J.,Wrenger, S., Reinhold, D., Ansorge, S. &Neubert, K. New results on the conforma-tions of potent DP IV (CD26) inhibitorsbearing the N-terminal MWP structuralmotif. Adv. Exp. Med. Biol. 524, 65-68.

Münzenberger, B., Hammer, E., Wray, V.,Schauer, F., Schmidt, J. & Strack, D. Deto xi -fication of ferulic acid by ectomycorrhizalfungi. Mycorrhiza13, 117-121.

Nguyen Thi Hoang Anh, Tran Van Sung,Franke, K. & Wessjohann, L. Phytochemicalstudies of Rehmannia glutinosa rhizomes.Pharmazie 58, 593-595.

Nguyen Thi Hoang Anh, Tran Van Sung,Porzel, A., Franke, K. & Wessjohann, L. A.Homoisoflavonoids from Ophiopogonjaponicus Ker-Gawler. Phytochemistry 62,1153 - 1158.

Nguyen Thi Hoang Anh, Tran Van Sung &Wessjohann, L. A. Some homoisoflavonoi-dal compounds from Ophiopogon japoni-cus Ker-Gawler. Tap chí Hóa hoc (Vietnam.Journal of Chemistry) 41, 114-118.

Nguyen Thi Hoang Anh, Tran Van Sung &Wessjohann, L. A. Steroidal glycosides from Ophiopogon japonicus. Tap chí Hóa hoc(Vietnam. Journal of Chemistry) 41, 135-141.

Ounaroon, A., Decker, G., Schmidt, J., Lott -speich, F. & Kutchan, T. M. (R,S)-Reticuline7-O-methyltransferase and (R,S)-norco-claurine 6-O-methyltransferase of Papaversomniferum - cDNA cloning and characte-rization of methyl transfer enzymes of al -kaloid biosynthesis in opium poppy. PlantJ., 36, 808-819.

Raith, K., Neubert, R., Poeaknapo, C.Boettcher, C., Zenk, M. H. & Schmidt, J.Electrospray tandem mass spectrometricinvestigations of morphinans. J. Am. MassSpectrom. 14,1262-1269.

Richter W. G., Cappi M. W., Henkel B., HessS., Surivet J. P., Baeschlin D., HubschwerlenC., Höfle G., Wessjohann L. & EichelbergerU. Second generation epothilones andtheir biological properties. Clin. Canc. Res.(Part 2, Suppl. S) 9 (16), 6137S-6137S.

Samappito, S., Page, J., Schmidt, J., De-Ek -namkul, W. & Kutchan, T. M. Aromatic andpyrone polyketides synthesized by a stil -bene synthase from Rheum tataricum. Phy -to chemistry 62, 313-323.

Schmutz, E., Steffensky, M., Schmidt, J., Por -zel, A., Shu-Ming Li & Heide, L. An unusualamide synthetase (CouL) from the cou-mermycin A1 biosynthetic gene clusterfrom Streptomyces rishiriensis DSM40489. EJB. 270, 4413-4419.

Schütz, J., Brandt, W., Spetea, M., Wurst, K.,Wunder, G. & Schmidhammer, H. Synthesisof 6-amino acid substituted derivatives ofthe highly potent analgesic 14-O-methyl -oxymorphone. Helv. Chim. Acta 86, 2142-2148.

Spiteller, P., Arnold, N., Spiteller, M. & Steg -lich, W. Lilacinone, a red aminobenzoqui -none pigment from Lactarius lilacinus. J.Nat. Prod. 66, 1402-1403.

Trinh Thi Thuy, Tran Van Sung & Wess jo -hann, L. Chemical constituents of the rootsof Codonopsis Pilosula. Vietnam. J. Chem.41, 119-123.

Wessjohann, L. A., Brandt, W., Thiemann, T.Biosynthesis and metabolism of cyclopro-pane-rings in natural compounds. Chem.Rev.103, 1625-1648.

Westermann, B. Organokatalyse oder dieSuche nach dem kleinsten Enzym. Nachr.


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Chem. Tech. Lab., 802-806.PatentsBuske, A., Adam, G., Häring, A., Kekulé, A.Arzneimittel enthalten Derviate von Tetra -chinolin. Brazilian Patent PI 0211026-1.

Feußner, I., Hornung, E., Krüger, C. & Por -zel, A. Verfahren zur Herstellung von kon-jugierten mehrfach ungesättigten Fett säu -ren mit mindestens drei Doppelbindungenin Pflanzen. DE 10308850.4

Wessjohann, L. A., Braun, H.-J., Oberson, S.& Umbricht, G. Two-component hair dyecompositions including triacetoxy-naph-thalene derivatives and a hydrolase. DE10226061

Wessjohann, L. A., Eichelberger, U. & TranThi Phuong, Thao Neue Makrocyclen zurBehandlung von Krebserkrankungen. DE

10355223.5Wessjohann, L. A., Scheid, G., Bornscheuer,U., Henke, E., Kuit, W. & Orru, R. V. A. Epo -thilone-Synthesebausteine IV: Unsymme -trisch substituierte Acyloine und Acyloin -derivate und Verfahren zu deren Herstel -lung. DE 0010134172A1

Wessjohann, L. A., Schmitt, M., Allwohn, J.,Umbricht, G. & Braun, H.-J. Hair dyes andmethod for dying hair with 1,2,4-trihydr oxy -naphthalene glycosides and glycosidases. DE10205500

Doctoral ThesesMarco Aurelio Dessoy: Synthesis and enzy-matic coupling of prenyldiphosphates andbenzoates. University of Halle-Witten berg,Department of Chemistry, 10/07/2003.

Michael Fulhorst: Prenyltransferases as po -

tential biocatalysts - substrate and inhibi-tor studies on p-hydroxybenzoate oligo-prenyl transferase from E. coli. Universityof Halle-Wittenberg, Department of Che -mistry, 22 / 09 / 2003.

Mieke Toorneman: (-)-Myrtenol based trig-ger system for the controlled formation ofenediynes. Vrije Universiteit Amsterdam,Fakulteit Exakte Wetenschappen 30/10/2003.

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PublicationsBräuer, L., Brandt, W. & Wessjohann, L. A.Mo deling of the E. coli - 4-hydroxybenzoicacid oligoprenyl-transferase (ubiA-transfe-rase) and cha racterization of potentialactive sites. J. Mol. Model. 10, 317-327.

Brandt, W., Dessoy, M. A., Fulhorst, M., Gao,W., Zenk, M. H. & Wessjohann, L. A. A pro-posed me chanism for the reductive ringopening of the cyclodiphosphate MEcPP, acrucial transforma tion in the new DXP/MEP-pathway to isoprenoids based onmodeling studies and feeding ex periments.ChemBioChem 5, 311-323.

Franke, K., Nasher, A. K. & Schmidt, J. Con -sti tuents of Jatropha unicostata. Biochem.Syst. Ecol. 32, 219-220.

Frick, S., Chitty, J. A., Kramell, R., Schmidt, J.,Allen, R. S., Larkin, P. J. & Kutchan, T. M.Trans -formation of opium poppy (Papaver somni-ferum L.) with antisense berberine bridgeenzyme gene (anti-bbe) via somatic em -bryogenesis re sults in an altered ratio ofalkaloids in latex but not in roots. Trans -genic Research 13, 607-613.

Galm, U., Dessoy, M. A., Schmidt, J., Shu-Ming Li, Wessjohann, L. A. & Heide, L. Invitro and in vivo production of new ami-nocoumarins by a combined biochemical,ge netic and synthetic ap proach. Chemistry& Biology 11, 173-183.

Hans, J., Brandt, W. & Vogt, T. Site-directedmutagenesis and protein 3D-homologymodelling suggest a catalytic mechanismfor UDP-glucose-dependent betanidin 5-O-glucosyltransferase from Dorothean -thus bellidiformis. Plant J. 39, 319-333.

Hirata, K., Poeaknapo, C., Schmidt, J. & Zenk,M. H. 1,2-Dehydroreticuline synthase, thebranch point enzyme opening the morphi-nan biosynthetic pathway. Phytochemistry65, 1039-1046.

Horvath,G., Wessjohann, L., Guissez, Y., Bie -baut, E., Caubergs, R. J. & Horemans, N.Seeds of grapes of Vitis vinfera var.Alphonse Lavallée (Royal) - a possiblemodel tissue for studying tocotrienol bio-synthesis. Acta Hortic. 652, 415-424.

Huneck, S., Feige, G. B. & Schmidt, J. Che -mie von Cladonia furcata und Cladoniarangiformis. He r zogia 17, 51-58.

Krelaus, R. & Westermann, B. Preparationof pep tide-like bicyclic lactams via a se -quential Ugi reaction-olefin metathesis ap -proach. Tetra he dron Lett. 45, 5987-5990.

Lübken, T., Schmidt, J., Porzel, A., Arnold, N.& Wessjohann, L. Hygrophorones A-G: Fun -

gicidal cyclopentenones from Hygro pho -rus species (Ba sidiomycetes).Phytochemistry 65, 1061-1071.

Micskei, K., Hajdu, C., Wessjohann, L., Mercs,L., Kiss-Szikszai, A. & Patonay, T. Enantio se -lective reduction of prochiral ketones bychromium(II) amino acid complexes.Tetra hedron: Asymm. 15, 1735-1744.

Myint Myint Khine, Franke, K., Arnold, N.,Por zel, A., Schmidt, J. & Wessjohann, L. Anew cardenolide from the roots of Strep -tocaulon to men tosum. Fi to te rapia 75,779-781.

Nguyen Thi Hoang Anh, Tran Van Sung &Wess johann, L. Further constituents fromOphiopo gon japonicus. Vietnam. J. Chem.42, 261 - 264.

Page, J., Hause, G., Wenyun Gao, Raschke,M., Schmidt, J., Zenk, M. H. & Kutchan, T. M.Func tional analysis of the final steps of the1-deoxy-D-xylulose 5-phosphate (DXP)pathway to isoprenoids in plants using vi -rus-induced gene si lencing. Plant Physiol.134, 1401-1413.

Poeaknapo, C., Fisinger, U., Zenk, M. H. &Schmidt, J. Evaluation of the mass spectro-metric fragmentation of codeine and mor-phine after 13C-isotope biosynthetic labe-ling. Phytoche mistry. 65, 1413-1420.

Poeaknapo, C., Schmidt, J., Brandsch, M.,Dräger, B. & Zenk, M. H. Endogenous for-mation of morphine in human cells. Proc.Natl. Acad. Sci. USA, 101, 14091-14096.

Scheid, G., Kuit, W., Ruijter, E., Orru, R. V. A,Henke, E., Bornscheuer, U. & Wessjohann,L. A new route to protected acyloins andtheir en zymatic resolution with lipases.Eur. J. Org. Chem. 5, 1063-1074.

Scheid, G., Ruijter, E., Konarzycka-Bessler,M., Born scheuer, U.T. & Wessjohann, L. Syn -thesis and resolution of a key buildingblock for epo th ilones: A comparison ofasymmetric synthesis, chemical and enzy-matic resolution. Tetra hedron Asymm. 15,2861-2869.

Schneider, P. H., Schrekker, H. S., Silveira, C.C., Wessjohann, L. A. & Braga, A. L. First ge -neration of cysteine- and methionine-de -rived oxazoli dine and thiazolidine ligandsfor palladium-catalyzed asymmetric allyla-tions. Eur. J. Org. Chem. 12, 2715-2722.

Schrekker, H. S., Micskei, K., Hajdu, C., Pa -tonay, T., de Bolster, M. W. G. & Wessjohann,L. A. Involvement of an oxidation-reducti-on equilibrium in chromium-mediatedenantioselective Nozaki-Hiyama reactions.Adv. Synth. Catal. 346, 731-736.

Seipold, L., Gerlach, G. & Wessjohann, L. Anew type of floral oil from Malpighia coc-cigera (Mal pighiadeae) and chemical consi-derations on the evolution of oil flowers.Chemistry & Biodiversity 1, 1519-1528.

Trinh Thi Thuy, Kamperdick, C., Pham ThyNinh, Trinh Phuong Lien, Tran Thi PhuongThao & Tran Van Sung. Immunosuppressiveauronol glycosides from Arto carpus tonki-nensis. Die Pharmazie 59, 297-300.

Trinh Thi Thuy, Tran Van Sung & Adam, G.Study on chemical constituents of Viet na -mese Mi cro melum hirsutum. Vietnam. J.Chem. 42, 177-181.

von Roepenack-Lahaye, E., Degenkolb, T.,Zer jeski, M., Franz, M., Roth, U., Wess jo -hann, L., Schmidt, J., Scheel, D. & Clemens,S. Profiling of Arabidopsis secondary meta-bolites by capillary liquid chromatographycoupled to electrospray ionization qua-drupole time-of flight mass spectrometry.Plant Physiol. 134, 548-559.

Wessjohann, L., Wild, H. & Schrekker, H.Chro mium-mediated aldol and homoaldolreactions on solid support directed to -wards an iterative polyol strategy. Tetra he -dron Lett. 45, 9073-9078.

Westermann, B, Diedrichs, N., Krelaus, R.,Wal ter, A. & Gedrath, I. Diastereoselectivesynthesis of homologous bicyclic lactams-potential building blocks for peptide mi -mics. Tetrahedron Lett. 45, 5983-5986.

Zakharova, S., Fulhorst, M., Luczak, L. &Wess johann, L. A. Synthesis, inhibitory andactivation properties of prenyldiphosphatemimics for aromatic prenylations withubiA-prenyl transferase. Arkivoc 13, 79-96.

Zhu, M., Ruijter, E. & Wessjohann, L. Newscavenger resin for the reversible linkingand mo noprotection of functionalized aro-matic alde hydes. Org. Lett. 6, 3921-3924.

Books and Book ChaptersHuneck, S., Lumbsch, H. T., Porzel, A. &Schmidt, J. Die Verteilung von Flechten in -haltsstoffen in Lecanora muralis und Leci -dea inops und die Ab hängigkeit derUsninsäure-Konzentration vom Substratund von den Jahreszeiten bei Lecanoramuralis. In: Contri butions to Lichenology.Fest schrift in Ho nour of Hannes Hertel(Döbbeler, P. & Ram bold, G., eds.) Bibl.Lichenol. 88, 211-221.

Lendeckel, U., Bukowska, A., Lättig, J. H. &Brandt, W. Alanyl-Aminopeptidase in hu -man T cells: Structure and function. In:Ami no pepti da ses in Biology and Disease(Hop per, N. M. & Len deckel, U., eds.)


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Kluwer Academic /Plenum Pub lis hers,New York, pp 210-227.Publications in PressAbbas, M., Neuhaus, C., Krebs, B. & Wes -ter mann, B. Synthesis of g-amino acids viacatalytic asymmetric Homo-Mannich reac-tions. Synlett.

Braga,A. L., Lüdtke, D. S., Wessjohann, L. A.,Paixão, M. W. & Schneider, P. H. A chiral di -sulfide derived from (R)-cysteine in theenantioselective addition of diethylzinc toaldehydes: Loading effect and asymmetricamplification. Synthesis.

Braga, A. L., Alves, E. F., Silveira, C. C., Zeni,G., Appelt, H. R. & Wessjohann, L. A. A newcysteine derived ligand as catalyst for theaddition of diethylzinc to aldehydes: Theimportance of a "free" sulfide site forenantioselectivity. Synthesis.

Brandt, W. & Wessjohann, L. A.The functio-nal role of selenocysteine (Sec) in the ca -talysis mechanism of large thioredoxin re -ductases: Proposition of a swapping cataly-tic triad including a Sec-His-Glu state.ChemBioChem. 6.

Nguyen Thi Hong Van, Nguyen Thi HoangAnh, Tran Van Sung, Franke, K. & Wessjo -hann, L. Stil bene ferulic acide and its deri-vatives from the roots of Angelica sinensis.Vietnam. J. Chem. 42.

Teichert, A., Lübken, T., Schmidt, J., Porzel,A., Arnold, N. & Wessjohann, L. Unusual

bioactive 4-oxo-2-alkenoic fatty acids fromHygrophorus eburneus. Z. Naturforsch. B.Rodrigues, O. E. D., Perottoni, J., Paixao, M.W., Zeni, G., Lobato, L. P., Braga, A. L., Ro -cha, J. B. T. & Emanuelli, T. Renal and hepaticALA-D activity and selected oxidativestress parameters of rats exposed to inor-ganic mercury and organoselenium com-pounds. Fd. Chem. Toxic. 42, 17-28.

Ruijter, E., Schültingkemper, H. & Wess jo -hann, L. A. Highly substituted tetrahydro -pyrones from hetero-diels-alder reac tionsof 2-alkenals with stereochemical inducti-on from chiral dienes. J. Org. Chem.

Wessjohann, L.A. & Ruijter, E. Macrocyclesrapidly produced by multiple multicompo-nent reactions including bifunctional buil-ding blocks (MiBs). Molecular Diversity 9.

Wessjohann, L. A., Ruijter, E., Garcia-Rivera,D. & Brandt, W. What can a chemist learnfrom nature s macrocycles? - A brief, con-ceptual view. Molecular Diversity 9.

Books and Book Chapters in PressMrestani-Klaus, C., Faust, J., Golbik, R.,Brandt, W., Wrenger, S., Reinhold, D. &Neubert, K. De tection of PPII-helix-likestructural features in short proline-contai-ning peptide inhibitors of the cell-surfaceprotease dipeptidyl peptidase IV. In: Pep ti -des 2004 - Proceedings of the 3rd Inter na -tional and 28th European Peptid Sym po -sium, Prag.

Nuhn, P. & Wessjohann, L. A. Natur stoff -chemie - mikrobielle, pflanzliche und tieri-sche Natur stoffe. 4th edition, S. HirzelVerlag, Stuttgart (Germany).

Wessjohann, L. A. & Ruijter, E. Strategiesfor total and diversity-oriented synthesisof Na tural product (-like) macrocycles. In:Top. Curr. Chem. Vol. 243 (J. H. Mulzer,ed.), Springer, Heidelberg.

PatentsLübken, T., Arnold, N., Wessjohann, L., Fa.Morphochem. Hygrophorone und derenDerivate. German Patent 10 2004 015566.6.

Wessjohann, L., Eichelberger, U., Tran ThiPhuong, T. Neue Makrocyclen zur Be hand -lung von Krebserkrankungen. InternationalPatent PCT/EP 2004/013451.

Doctoral ThesesHenri Stephan Schrekker: Selective orga -nic reactions promoted by chromium(II)ions. Vrije Universiteit Amsterdam,20/02/2004.

Lars Seipold: Blütenöle - Chemische Analy -se, Biosyn these und Betrachtungen zurEnt stehung von Ölblumen. University ofHalle-Wittenberg, Department of Che -mistry, 24/05/2004.

Diploma TheseAxel Teichert: Fungitoxische Inhaltsstoffeaus Fruchtkörpern der Gattung Hygro -pho rus (Basidiomycetes). University of Leip -

Department of

Bioorganic Chemistry

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Department of Stress and Developmental BiologyHead: Dierk Scheel, ProfessorSecretary: Ruth Laue until March 2004, afterwards Susanne Berlin

Department of Stress and

Developmental Biology

37

Plant development, although genetically determined, is largely modulatedby biotic and abiotic environmental factors. In this way, developmental

programs are adapted to specific local conditions and protective as well asdefense reac tions are initiated during stress situations - an advantageoussituation for sedentary living plants.

The basis for those processes is the ability of plants to perceive environ-mental factors and initiate signal transduction networks that modify geneexpression patterns. The investigation of the molecular mechanisms under-lying this course of events is the main topic of the Department of Stressand Developmental Bio logy.

Plant pathogens play a major role in biotic stress. The work of severalresearch groups of the department focuses on the analysis of recognition,signal transduction and gene activation processes in plant-pathogen inter-actions. The work on abiotic environmental factors centers around metalhomeostasis in plants, using hyperaccumulating model organisms.

Plant responses to biotic and abiotic environmental factors finally result inaltered patterns of proteins and metabolites. In order to be able to directlymonitor such alterations, mass spectrometry-based methods have beenestablished allowing the comprehensive profiling of proteins and secondarymetabolites. Both me thods are also being employed for biochemical phe-notyping of mutants.

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Research Group: Molecular Communication in Plant-Pathogen InteractionsHead: Wolfgang Knogge

To identify pathogen genes whose pro-ducts are involved in the communica -tion with the host and, hence, play arole during pathogenesis, mutagenesisstrategies can be applied. Loss-of-func -tion mutations in genes whose productscontribute to optimal pathogenesis (vi -ru lence genes) or that are even essen -tial for microbial penetration and deve-lopment in planta (pathogenicity genes)can be presumed to generate a pheno-

type that deviates more or less obviously from thewild type situation. Isolation and detailed molecularanalysis of the affected genes and their protein pro-ducts will lead to the unraveling of the processesthat are important for the interaction. An alterna-tive strategy aims at identifying the proteins thatare secreted by the pathogen ("secretome"). Theseproteins are either required for the synthesis ofmi crobial extracellular structures or candidates forfactors involved in the interaction with the host.

Rhynchosporium secalis, the causal agent of a leafspot disease on several grasses, is of particular eco-nomic importance as a pathogen of barley. Afterpe netration of host leaves the fungus grows in theextracellular region beneath the leaf cuticle and theepidermal cells of susceptible plants. During earlystages of pathogenesis epidermal cells collapse,whereas necrotic lesions only become visible du -ring later stages when the mesophyll cells are affec-ted as well. Small proteins that are secreted by thefungus are involved in the expression of diseasesymptoms, as was shown using fungal knock-outmutants. In the presence of resistance gene Rrs1 in

the host plant, one of these virulence factors, NIP1,becomes a specific recognition signal that serves asthe trigger of plant resistance reactions. A specificNIP1-binding site was detected on membranesfrom resistant and susceptible barley cultivars aswell as from other cereals. This indicates that theNIP1 receptor is not encoded by the Rrs1 geneand that (an) additional component(s) are requiredfor resistance-related signal transduction.

Recently, two different mutagenesis approaches, in -sertion mutagenesis and promoter trapping, led tothe identification of several fungal pathogenicityand virulence genes. The deduced functions of the

Group Members

Barbara DegnerTechnician

Claudia MönchmeierDiploma Student since September2004

Sylvia SierslebenPhD Student since April 2004

Marina WibeDiploma Student since April 2004

Many phytopathogenic microorganisms colonize the intercellular spaces of their host plants. This regionis relatively poor in nutrients and in order to optimize their life style pathogens therefore needed to deve-lop strategies that aim at improving the nutrient supply by the host cells. To prevent just this, plants ontheir part evolved mechanisms that enable the efficient recognition of invaders as the prerequisite fortheir rejection. These communication processes are crucial for the outcome of an interaction. They aremediated by membrane-loca lized or intracellular receptors in the plant, which are the targets of patho-gen-secreted mole cules, frequently of proteinaceous nature. As the consequence of these protein-proteininteractions two scenarios can be envisaged. The plant metabolism is redirected in favor of the pathogenand disease ensues. Alternatively, upon recognition of the pathogen through secreted compounds theplant defense is induced leading to the expression of plant resistance. Furthermore, in particular in com-patible interactions pathogen receptors may be involved that sense stimuli of plant origin and, hence, allowthe adaptation of the pathogen physiology to the specific environment of the host.

Schematic depiction of the communication between R. secalisand barley. (1) NIP1, NIP2, NIP3; (2) transcription factor; (3)permease; (4) P450 protein; (5) histidine protein kinase; (6)intramembrane protease. (C) cuticle, (CW) cell wall, (H) fun-gal hypha, (N) nucleus, (PC) plant cytoplasm, (PM) plasmamembrane, (T) tonoplast, (V) vacuole.

39

gene products, although biochemically not yetconfirmed, suggest their role in various pathoge-nic processes. Thus, a protein with sequence ho -mology to a class of transcription factors is li kelyto control fungal genes that are only ex pressedduring fungal growth in the plant. A g-aminobuty-ric acid /ami no acid permease is supposed to me -diate fungal uptake of specific plant nutrients,where as a P450 protein may be involved in theoxidation of substrate with an as yet unknownfunction. One of the genes encodes a histidineprotein kinase. These enzymes are part of intra-cellular information processing systems that linksig nals from the environment with specific cellu-lar adaptive processes. Al though the im mediatefunction of most of these enzymes is not known,their high number in phytopathogenic fungi is in -

terpreted as indicating an impor-tant role in microbial adaptation totheir respective host plants. In thiscontext the question arises as tothe existence and na ture of (a)plant factor(s) that may affect fun-gal development through these en -zyme sys tems. Finally, a gene wasidentified that encodes an intra-membrane protease of the rhom-boid type. These enzymes ca ta lyzethe release of ex tracellular signals from trans-membrane precursor proteins. However, neitherthe identity of the sig nal generated by R. secalisnor its target or the role of "regulated intramem-brane proteolysis" in fun gus-plant interactions ingeneral are known to date.

Collaborators

James BrownJohn Innes Centre, Norwich, UK

Barbara HowlettUniversity of Melbourne, Australia

Bruce McDonaldSwiss Federal Institute of Technology,Zurich, Switzerland

Dirk WarneckeUniversity of Hamburg, Germany

40

Research Group: Cellular SignalingHeads: Dierk Scheel & Justin Lee

The interaction of parsley with thesoybean pathogen, Phytophthorasojae, constitutes the main model forstudies of plant non-host disease resi-stance in our group. This form of resi-stance is comparable to the basal orinnate immunity of animal systems.Pep-13, an oligopeptide derived from aP. sojae cell wall transglutaminase, ser-ves as a pathogen-associated mo -lecular pattern (PAMP) to trigger de -fense reactions in parsley cells. Uponrecognition of the Pep-13 elicitor by aplasma membrane-localized receptor,a network of cellular signaling eventsis triggered that culminates in the ap -propriate defense response. The sig -naling events elucidated in the parsley-Pep-13 system include ion fluxes atthe plasma membrane, activation ofmitogen-activated protein kinases(MAPK) and the accumulation of sig -naling molecules such as reactive oxy-gen species, phospholipids and jasmo-nates. In particular, studies with thecalcium-sensitive reporter, aqueorin,have shown that calcium influx, with acharacteristic signature of sustained

cytosolic Ca2+ increase, is an early and key stepthat is essential for all other downstream eventsstudied. Thus, molecular characterization of thecalcium channel involved will be pur sued.

Parsley genes encoding MAPKs and their up -stream MAPK kinases (MKK) have been isolated.The genes designated PcMPK3a/b, PcMPK6 andPcMKK5 have been identified as being the Pep-13-activated components in the parsley MAPKcascade. Immunofluorescence studies indicatethat the activated MAPKs are rapidly transloca-ted in to the nucleus while the upstream PcMKK5is seen to accumulate at the perinuclear regionupon elicitation. It appears that MKK might serve

as a cytoplasmic anchor for MAPK prior to elici-tor stimulus. Phosphorylation of MAPK by MKKleads to its release from a MAPK-containing pro-tein complex, possibly allowing the activatedMAPKs to interact with their substrates.The spa-tial redistribution into the nucleus indicates thatsome of its substrates are likely to be nuclearproteins, presumably factors that control genetranscription. Transient expression of dominantinactive versions of PcMPK3a and/or PcMPK6and/or PcMKK5 interfered with the expressionof co-transfected pathogenesis-related (PR1 orPR2) gene promoter-reporter constructs. Expe -riments with inactive mutants of other, non-Pep-13-activated, MAPK or MKK do not show thiseffect. In contrast, the inhibitor of the NADPHoxidase, DPI, did not affect expression of this par-ticular subset of defense genes but blocked phy-toalexin accumulation in parsley. Hence, the oxi-dative burst and the MAPK cascade are requiredin the induction of distinct subsets of defensegenes by Pep-13.

A constitutive active version of PcMKK5 was suf-ficient to activate PcMPK3/6 and drive PR-geneexpression in parsley. He terolo gous expressionof this constitutive active PcMKK5 in transgenicArabidop sis, similarly led to ac ti vation of the Ara -bi dop sis MPK3/6or thologs and themRNA ac cumu la -tion of many de -fense-related ge -nes. Binding sitesfor WRKY trans -cription factors areover-representedin the promotersof these ge nes;suggesting that ac -tiva ted MPK3/6might target WRKYfactors directly or

Elicitation of parsley cells leads to translocation ofthe activated MAPK (PcMPK6, upper panel) intothe nucleus while the corresponding upstreamMAP kinase kinase (PcMKK5, lower panel) remainscytoplasmic or is localized in perinuclear region de -spite activation. Right panels are the correspondingDAPI staining (blue) to visualize nuclei position.

The sessile lifestyle of plants compels the development of a wide spectrum of defense reac-tions to counteract and deter potential pathogens, insects and parasites. Our researchemphasis is on plant defense signal transduction pathways - in non-host plant- pa thogeninteractions as well as the so-called gene-for-gene type of disease resistance. In addition,infection strategies of Cuscuta sp., a parasitic plant, are also being investigated.

Group Members

Reetta AhlforsGuest Scientist until April 2003

Gerit BethkeDiploma Student since February 2004

Mandy BirschwilksPostdoctoral Position since April 2003

Jutta ElsterTechnician

Anja HalbauerDiploma Student since April 2004

Franziska HandmannPhD Student since March 2003

Jan Heise Postdoctoral Position since March2004

Nora Köster-EiserfunkeDiploma Student since February 2004

Sylvia KrügerTechnician

Violetta MacioszekPostdoctoral Position until April 2004

Kai NaumannPostdoctoral Position since November2004

Anja NickstadtPhD Student

Jason RuddPostdoctoral Position until February2004

Christel Rülke Technician

Rita Schlichting PhD Student

Claudia Spielau Diploma Student until September2004, afterwards PhD Student

Christiane UngerDiploma Student until January 2004

Ivy WidjajaPhD Student since April 2004

41

indirectly.We are currently attempting to identifyMAPK substrates using different biochemical,proteomic and molecular techniques. The com-pleted genome sequence and the extensive post-genomic tools available for the model plant, Ara -bidopsis thaliana, render it the plant of choice forsuch studies.

Monospecific antibodies to AtMPK3, AtMPK6and AtMPK4 have been generated. As observedfor the parsley-Pep-13 system, activation ofMAPKs, e.g. by ozone treatment, also led to theirnuclear translocation. Furthermore, diverse gene-ral elicitors and abiotic stresses were shown toactivate these kinases. The convergence of va -rious stress factors at the same MAPK activationalso introduces the question on how specificity insignaling is maintained. One possibility is the exis -tence of defined protein complexes (signalo -somes). Indeed, size-exclusion chromatographyshowed that MAPKs in protein extracts, albeitonly a small portion, migrate as larger molecularcomplexes.Transgenic plants with tandem-affinitytagged versions of the elicitor-activated MAPKshave been generated to facilitate the isolation ofsuch protein complexes.

We have established high-resolution two-dimen-sional PAGE analysis of proteins from transgenicArabidopsis plants with inducible expression ofthe bacterial avirulence gene, AvrRPM1, in thepresence or absence of the corresponding resi-

stance RPM1gene. Pro teinspots that are dif-ferentially ac cu -mulated are iden-tified via MALDI-TOF spectrome-try. In order to un -derstand the dif -

ferences/similarities between non- host and the ge ne -for-gene type ofdisease resistance, the analysis willbe extended to transgenic plantsthat ex press a Phy toph tho ra sojaeNecrosis-Inducing Pro tein (NIP),which represents a ge neral elicitorof defense in the non-host Ara bi -dopsis plant.

The interaction of the holoparasi-tic plant, Cuscuta sp., with Ara bi -dopsis was established for study-ing angiosperm parasites. Not unli-ke other phytopathological sys -tems, successful invasion requiresthe transfer of nutrients and waterfrom the host to the parasite; inthis case via a multicellular hausto-rial structure. Specific xylem-mo -bile or phloem-mobile dyes de -mon stra t ed a functional connec -tion of the invading haustorium tothe vascular system of the host.The high efficiency of nu trienttransfer to the parasite can even-tually result in death of the host.In com patibility between Cus cutaand its host has occasionally beenobserved, where the functionalhaustoria is terminated at variousdevelopmental stages. A screen invari ous ecotypes and defense-re -lated mu tants has been initiated toelucidate the components responsible for incom-patibility.

Collaborators

Thomas BollerUniversity of Basel, Switzerland

Jeff DanglUniversity of North Carolina, Chapel Hill,USA

Jerome GiraudatInstitut des Sciences du Végétal, CNRS,Gif-sûr-Yvette, France

Minna Haapalainen, Suvi Taira, Martin RomantschukUniversity of Helsinki, Finland

Bettina HauseLeibniz Institute of Plant Biochemistry,Halle, Germany

Heribert HirtUniversity of Vienna, Austria

Jonathan JonesThe Sainsbury Laboratory, Norwich, UK

Sophien KamounOhio State University, Wooster, USA

Birgit KerstenMax Delbrück Center for MolecularMedicine, Berlin, Germany

Chris LambJohn Innes Centre, Norwich, UK

John MundyUniversity of Copenhagen, Denmark

Teun MunnikUniversity of Amsterdam, The Netherlands

Karsten NiehausUniversity of Bielefeld, Germany

Thorsten NürnbergerUniversity of Tübingen, Germany

Jane Parker, Imre Somssich, Joachim UhrigMax Planck Institute of Plant BreedingResearch, Cologne, Germany

Jose Sanchez-SerranoAutonomous University, Madrid, Spain

Two-dimensional polyacrylamide gel electrophore-sis analysis of proteins in transgenic Arabidopsisplants that are induced to express the bacterialAvrRPM1 gene. The ar rows point to protein spotsthat are more abundant in these plants as compa-red to isogenic plants lackingg the RPM1 resistancegene, that is required for percep tion/transducingthe AvrRPM1 signal.

Cuscuta twining around the stem of an Ara bidop sis inflo-rescence. The close-up shows an en larged area where thehaus torial region is visible. Inset is a transverse section illu-strating the development of a haustorium towards the vas-cular systems of the host.

42

Research Group: Induced Pathogen DefenseHeads: Dierk Scheel & Sabine Rosahl

The Phytophthora sojae-derived oli-gopeptide elicitor, Pep-13, originallyidentified as an inducer of plant de-fense in parsley and shown to act as apathogen-associated molecular pat-tern (PAMP) in evoking innate im-mune responses, triggers defense re -sponses in potato, such as the oxida-tive burst, defense gene activation andthe accumulation of the signaling com-pound salicylic acid. Interestingly, deri-vatives of Pep-13 show similar elicitoractivity in parsley and potato, sugge-sting a receptor-mediated inductionof defense responses in potato similarto that observed in parsley. However,unlike in parsley, infiltration of Pep-13

into leaves leads to hypersensitive-like lesions inpotato, which show characteristics of pro gram -med cell death. Analyses of transgenic potatoplants, which are unable to accumulate salicylicacid, due to the expression of a salicylate hy -droxylase gene, suggest that a subset of the Pep-13-induced de fense responses in potato leaves issalicylate-de pendent.

In addition to the local accumulation of salicylicacid, infiltration of Pep-13 or the phytopathoge-nic bacteria, Pseudomonas syringae pv. maculico-la, induces increases in the levels of the signalingcompounds, jasmonic acid, as well as its biosyn-thetic precursor, 12-oxophytodienoic acid.Interestingly, 12-oxophytodienoic acid, but notjasmonic acid, accumulates also systemically,which correlates with the expression of systemicacquired resistance. To analyze the role of jasmo-nic and 12-oxophytodienoic acid for both localand syste mic defense responses, transgenic plantswere ge nerated, which express RNA interfe-rence constructs targeted at several genes enco-ding en zymes of the jasmonic acid biosyntheticpathway. Significant reduction in jasmonic and 12-

oxophytodienoic acid levels were achieved inplants with reduced expression of genes enco-ding allene oxide cyclase and 12-oxophytodienoicacid re ductase.

Oxylipins synthesized via the lipoxygenase path-way possess antimicrobial activity and can act assignaling molecules.We tested 49 different oxyli-pins for their inhibitory effects against P. infestansin an in vitro assay. 18 were able to significantlyreduce mycelial growth of the oomycete or inhi-bit spore germination. Among those were pro-ducts of the 9-lipoxygenase pathway, which alsoaccumulate in potato leaves in response to Pep-13, as well as after pathogen infection. To analyzethe role of 9-lipoxygenase-derived oxylipins forthe response to pathogens, transgenic potatoplants expressing RNA interference constructstargeted at the pathogen-induced 9-lipoxygenaseand the 9-divinyl ether synthase were generated.Significantly reduced levels of 9-lipoxygenase pro-ducts and of the divinyl ethers colneleic and col-nelenic acids were observed in several transgenicplants. Lipid peroxidation during the hypersensi -tive cell death was not significantly altered in theabsence of 9-lipoxygenase activity. Real time PCRassays to determine pathogen biomass also didnot reveal significant differences in the growth ofP. infestans on a susceptible host plant with re -duced levels of 9-lipoxygenase activity. Whetheroxylipins from solanaceous plants like potato can

Group Members

Simone AltmannDiploma Student since October 2004

Lennart Eschen-LippoldDiploma Student since January 2004

Vincentius A. HalimPhD Student

Jörn LandtagPhD Student

Grit RothePostdoctoral Position

Angelika WeinelTechnician

Lore WestphalPostdoctoral Position

Dorothea WolfDiploma Student since June 2004

The oomycete, Phytophthora infestans, is the causal agent of late blight disease, one of the most deva-stating diseases of potato. To characterize defense mechanisms against this pa thogen, we are studyingthe interaction of P. infestans with its host plant potato and with the non-host plant Arabidopsis tha-liana. For potato, our major interest is the analysis of pathogen recognition, signal transduction andcharacterization of the defense reactions. On the other hand, we are identifying and characterizingmutants of the non-host plant Arabidopsis, which are compromised in non-host resistance to P. infe-stans.

Oxylipins inhibit germination of Phytophthora infestans invi tro. When added to a zoospore suspension of P. infestans,oxygenated polyunsaturated fatty acids are able to efficient-ly inhibit germination (right panel) compared to untreatedcontrols (left panel).

43

also be effective against pathogens in other plantsis being tested by transferring the respectivegenes from potato into A. thaliana.

Arabidopsis is not a host plant for P. infestans.Therefore, the identification of genes involved inthis non-host resistance should elucidate mecha-nisms of defense against the infectious agent oflate blight disease. P. infestans spores germinateon Arabidopsis leaves and attempt to penetratecells which react with the deposition of callose,

accumulation of autofluorescentmaterial and hypersensitive celldeath. Analysis of more than 40Arabidopsis ecotypes revealedthat, despite differences in theirhypersensitive response, none al -lows significant growth or multi-plication of the oomycete. TheArabidopsis mutant, pen2, whichwas isolated by Volker Lipka andPaul Schulze-Lefert (MPI Köln) asallowing enhanced penetration ofthe non-host pathogen barleypow dery mildew (Blumeria gram-nis f. sp. hordei), shows a highernumber of penetrated cells andin creased cell death in responseto infection by P. infestans. Seedsof the pen2 mutant were mutage-nized and approximately 70,000lines were screened for altera -tions in their response to infec -tion by P. infestans. Several mu -tants were isolated, which show increased hyper-sensitive cell death, or which allow enhancedpenetration of epidermal cells as well as en -hanced growth of P. infestans. Mapping of theposition of the affected genes is in progress.

Collaborators

Rod Casey John Innes Institute Norwich, UK

Ivo Feussner University of Göttingen, Germany

Markus Frank BASF Plant Science Ludwigshafen,Germany

Joëlle Fournier, Marie-Thé rèse Esquerré-Tugayé Université Paul Sabatier Toulouse, France

Christiane Gebhardt, Paul Schulze-Lefert Max Planck Institute for Breeding ResearchKöln, Germany

Mats Hamberg Karolinska Institute Stockholm, Sweden

Volker Lipka University of Tübingen, Germany

William Albert Rensink The Institute for Genomic Research(TIGR), Rockville, USA

Jose Sanchez-Serrano Universidad Autonoma de Madrid, Spain

Claus WasternackLeibniz Institute of Plant Biochemistry,Halle, Germany

Arabidopsis mutants impaired in non-host resistance to Phytophthora infestans. In contrastto wild type Arabidopsis (left leaf), two mutants react to infection with P. infes tans with en -hanced cell death, indicated by stronger staining with trypan blue (middle and right leaves).

Elicitation of the oxidative burst by Pep-13 is dependent onsalicylic acid. The oligopeptide elicitor Pep-13 induces theaccumulation of hydrogen peroxide in potato plants as vi -sualized by staining with diaminobenzidine (left). However,in plants unable to accumulate the signaling compound sali-cylic acid, accumulation of reactive oxygen species is im -paired (right).

44

Research Group: Metal HomeostasisHead: Stephan Clemens

Comparative transcript profiling withA. thaliana and A. halleri has led toim portant insights into the molecularmechanisms of metal hyperaccumula-tion. A. halleri is a hyperaccumulatorgrowing on metal-contaminated sitesin Central Europe. We found (pre-vious cDNA-AFLP re sults) that thesimilarity of coding se quences of A.thaliana and A. halleri is high enoughto use A. thaliana GeneChips also forA. halleri.We performed a number ofmicroarray experiments to find con-stitutive differences in the root tran-scriptomes of the two species and toidentify transcriptional changes in re -sponse to various doses of both es -sential and non-essential metal ions.The constitutive comparison revealedse veral genes that are expressed at amuch higher level in A. halleri. In te -res tingly, some of these encodeknown metal homeostasis factors.The two genes showing highest ex -pression in A. halleri roots relative toA. thaliana roots encode a nicotian-amine synthase (NAS) and a putativeZn2+ uptake system (ZIP). The signifi-

cantly high er activity of these and other genes in -volved in metal homeostasis could be confirmed.A. halleri roots also show higher NAS protein le -vels. Fur ther more, we developed a CapLC-ESI-QTOF-MS-based nicotianamine (NA) analysis pro-cedure and found higher NA levels in roots of A.halleri. Ex pression of a NAS in a Zn2+-hypersensi -tive Schi zo saccharomyces pombe mutant demon-strated that formation of NA can confer Zn2+ tole-rance. Taken together, these observations impli -cate NA in plant Zn homeostasis and in thehyperaccumula tion of Zn by A. halleri.

The identified genes are still responsive to eleva-ted metal levels in A. halleri albeit at concentra-tions orders of magnitude higher than in A. thalia-na. This led us to hypothesize that one molecularmechanism behind metal hyperaccumulation mightbe a de-regulation of metal deficiency re sponses.Additional microarray experiments with the "full-genome" ATH1 chip confirmed this pattern. It isan emerging picture that plant adaptation to ex -treme environments is mostly a regulatory pheno-menon and our microarray data lend further sup-port to this. It is now of paramount importance toelucidate the molecular ba sis for this altered regu-lation. We therefore cloned the promoters of se -veral metal homeostasis genes from both Ara -bidopsis species and generated various reporterlines. Preliminary results indicate that the A. halleripromoters retain most of their activity in A. tha-liana. The immediate goal of these experiments isthe identification of cis elements responsible forthe de-regulated ex pres sion in A. halleri.

The results of our GeneChip studies are also ofgeneral importance.They demonstrate the feasibi-lity of comparative microarray analysis of clo selyrelated species and their potential to become anextremely valuable tool for the molecular elucida-tion of biodiversity.

The involvement of nicotianamine in plant metalhomeostasis and Zn hyperaccumulation is studiedin more detail. All known isoforms from A. halleriwere cloned and functionally characte rized. Re gu -lation of the genes in response to varying suppliesof essential micronutrients is being analyzed byreal-time PCR. In addition, field samples from sitesin the Harz mountains that differ in degree of me -tal contamination are under investigation. Trans -genic approaches are aiming at modulating NAlevels in both Arabidopsis species.

Group Members

Annegret BähreckeDiploma Student until March 2004, after-wards PhD Student

Frank BretschneiderDiploma Student until September 2004

Sophie BundtzenDiploma Student until November 2004

Sandra FranzDiploma Student until September 2004

Thomas FritscheDiploma Student until September 2004,afterwards PhD Student

Marina HäußlerTechnician

Emiko HaradaPostdoctoral Position until January 2004,Guest Scientist since September 2004

Claudia SimmPhD Student until March 2004

Pierre TennstedtPhD Student

Aleksandra TrampczynskaPostdoctoral Position since December2003

Tino UnthanDiploma Student until September 2004

Christoph VeßPhD Student until June 2003

Susan WasserslebenPhD Student until August 2004

Michael WeberPhD student

Plants - like all other organisms - are able to tightly regulate the intracellular concentration and thedistribution of essential metals such as zinc and copper. Also, the cytosolic concentrations of non-essential toxic metals (e.g. cadmium, lead) have to be minimized. Some plant species (so-called metal-lophytes) can tolerate otherwise toxic concentra tions and grow on metal-contaminated soil. The pro-jects of our group are aiming at elucidating the molecular mechanisms underlying plant metal homeo-stasis, metal tole rance and metal hyperaccumulation. Plants under investigation are Arabidopsis thalia-na and its close relative Arabidopsis halleri, which grows, for instance, on medieval mining sites in theHarz mountains. In addition, we are working with Schizosaccharomyces pombe as a cellular model formetal homeostasis.

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Microarray data on the metal responses providedclues as to the modes of toxicity and helped iden-tifying common as well as species-specific tran-scriptional changes that will guide functional stu-dies. In particular, "core" response genes, genes re -sponsive specifically to excess of one particularmetal and putative signal transduction compo-nents were identified. A collection of T-DNA in -sertion lines is being established for putative sig naltransduction components that, based on their me -tal responsiveness, are hypothesized to be in -volved in the transcriptional regulation of genesupon changes in external metal concentrations.Homozygous lines are being characterized withrespect to tolerance, accumulation and transcrip-tome changes.

The synthesis of phytochelatins (PCs), small me tal-binding peptides derived from glutathione, is es -sential for the detoxification of Cd and As ions inplants, some fungi and certain animals. A numberof questions concerning the function of the ubiqui-tously present PC synthase encoding genes (PCSgenes) remain unanswered. Foremost, it is notclear, how the sporadic need to detoxify non-essential metal ions could have provided the se -lective pressure to account for the presence ofPCS genes throughout the plant kingdom (withthe exception of some mosses). We found twolines of evidence that might help to solve this my -stery: (i) studies on two PC-deficient A. thalianamutants clearly showed a key role of PC forma tionalso in buffering excess es sential metal ions; (ii)characterization of one of the bacterial proteinswith similarity to the PCS Pfam domain, alr0975from Nostoc spec., revealed that this protein cata-lyzes only the first formal step in PC synthesis, thecleavage of the C-terminal glycine from glutathione.This finding provides a clue as to the evolutionaryorigin of PC synthases and points to possible addi-tional roles in glutathione metabolism, e.g. the cata-bolism of GS-conjugates.

With respect to additional PCS functions, we arealso interested in the physiological role of the se -cond PCS gene in Arabidopsis, AtPCS2.We show-ed partial complementation of the PC-deficient

cad1-3 mutant with AtPCS2-HA.A homozygous insertion line forAtPCS2 is being studied. Compa -ra tive studies both in vitro and invivo on the activities of PCS1 and 2from A. thaliana and A. halleri wereinitiated.

A second major question we areaddressing concerning PC syntha-ses is their mode of activation.They are constitutively expressedyet PC synthesis occurs only in thepresence of elevated metal ion le -vels. Several metal ions are knownto elicit PC synthesis both in vivoand in vitro. Peptide scanningexperiments helped to identify pu -tative metal binding sites. Also, wehave identified differences in there sponse range among various PCsynthases and are now trying toestablish the structure-function relationshipsunderlying these differences. This might help togain insight into the interaction of metal ions andmetal-ligand complexes with proteins - a researchtopic that has come into focus with the realizationthat aberrant metal-binding might be involved inthe pathogenesis of various human disorders,including Alzheimer s and prion diseases.

S. pombe cells are being used as tools for the func -tional characterization of plant proteins and in or -der to learn more about the mechanisms of cellu-lar metal homeostasis in an easily workable sys -tem, that can serve as a model for PC-formingcells. The analysis of putative metal homeostasisfactors in S. pombe was extended. We construc-ted and characterized additional S. pombe knock-out mutants - especially for the transporters ofthe ZIP family. In order to help understand the ho -meo static function of the sole metallothionein inS. pombe and to develop an additional tool for theanalysis of the "metal status" of S. pombe cells, a"metal array" was established that covers knownmetal homeostasis genes as well as general stressresponse and sulphur metabolism genes.

Collaborators

Sebastian FettigUniversity of Bayreuth, Germany

Rüdiger HellUniversity of Heidelberg, Germany

Ute KrämerMax Planck Insituteof Molecular Plant Physiology, Golm,Germany

Youngsook LeePOSTECH, Pohang, South Korea

Andreas MeyerUniversity of Heidelberg, Germany

Dietrich H. NiesUniversity of Halle, Germany

Julian I. SchroederUniversity of California San Diego, La Jolla,USA

Sébastien ThomineInstitute des Sciences du Végétal, CNRSGif-sûr-Yvette, France

Nathalie VerbruggenUniversité de Bruxelles, Brussels, Belgium

Kuo-Chen YehInstitute of BioAgricultural Sciences,Academia Sinica, Taipeh, Taiwan

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Interdepartmental Research Group: Metabolite Profiling in Arabidopsis and Crop PlantsHeads: Stephan Clemens, Jürgen Schmidt, Ludger Wessjohann & Dierk Scheel

The main emphasis of the group hasbeen on developing a novel and com-petitive metabolite profiling platformexploiting the potential of hybrid massspectrometers developed in the pastfew years. The chemical diversity ofthe metabolome necessitates the useof different analytical techniques to co -ver the wide range of polarities foundamong the metabolites occurring in acell. Thus, we decided to complementexisting GC-MS approa ches with aprofiling scheme using li quid chroma-tography (LC). We established an ex -perimental system based on ca pillaryLC coupled to electrospray io nizationquadrupole time-of-flight mass spec-

trometry (CapLC-ESI-Qq-TOF). Major questionmarks concerning this approach were the robust-ness of the analysis, its reproducibility, and its suita-bility for high-throughput and quantitative analysis.We performed an extensive evaluation of the pro-cedures we had developed earlier. Our results de -monstrated, that CapLC-ESI-Qq-TOF-MS princi -pally meets all the necessary criteria for powerfulmetabolic analysis. Retention times of compoundseluting from the CapLC column were found toshow only little variation. Evaluation of the mass ac -curacy showed that the target value in the range offive ppm error can be reached for many signalseven in a complex mixture of compounds allowing,as discussed below, the calculation of elementalcompositions and thereby the tentative identifica -tion of compounds. Robustness of the analysis wasil lustrated by the fact that the average deviation fora range of mass signals was no higher than elevenpercent when the same extract was run severaltimes. Reproducibility of retention time and massaccuracy provided a sufficient basis for the unequi-vocal detection and identification of mass signals.Quantification is possible because a satisfactory li -

near relationship between compound abundanceand signal strength of the CapLC-ESI-Qq-TOF-MSanalysis was found for most of the signals tested.

The raw data generated by CapLC-ESI-Qq-TOF-MSare very complex and require deconvolution. Majorefforts therefore had to be devoted to developingand optimizing tools for the automatic data decon-volution and data processing. These achievementsgreatly increased the depth of analysis and thethroughput. Reliability of data was found to becomparable to manual data processing. On averageabout 1,400 signals are detected in leaf extractsand about 800 signals in root extracts. Given thelimited overlap between root and leaf metabolitesa total of about 2,000 different signals can be de -tected in extracts from Arabidopsis plants grownunder control conditions. Most likely the numberswill be significantly higher once the analysis is ex -tended to other tissues such as flowers, to differentdevelopmental stages and to plants exposed to en -vironmental stimuli.

Thus, this technique is suitable for detecting subtlemetabolic changes in response to environmentalstimuli or metabolic differences dependent on ge -netic background.This ability was proven already inpilot studies on Arabidopsis mutants affected inmetabolic enzymes and on Arabidopsis stress re - sponses. Furthermore, identification of metabolitesis feasible. Electrospray as a soft ionization methodcombined with the enhanced resolution and massaccuracy of the TOF instrument and tandem MS al -low to determine the elemental composition andto detect characteristic in-source fragments. Struc -tural information can also be obtained in a targetedway by performing tandem MS analysis of mass sig -nals showing interesting changes in abundance. Inconclusion, the principles of separation and massanalysis of this technique together with its sensiti-vity and resolution greatly expand the range of me -

Group Members

Christoph Boettcher Postdoctoral Position since August 2004

Thomas DegenkolbPostdoctoral Position until June 2003

Mathias FranzStudent

Kerstin Körber-FerlTechnician until March 2004

Edda v. Roepenack-LahayePostdoctoral Position

Udo RothPostdoctoral Position until September2003

Michaela WinklerTechnician since August 2004

Michael ZerjeskiStudent

Our project is aiming at contributing to the "post-genomic" analysis of the model organism Arabidopsisthaliana and of crop plants by establishing an extensive profiling of metabolites and, in addition, of proteinsand peptides. These profiles are used for the detection and identification of early stress responses. Also,they are valuable tools for the analysis of various developmental and stress-induced changes as well as forthe biochemical phenotyping of mutants and the exploration of natural diversity. Exemplary biotic andabiotic stresses un der investigation are pathogen attack and toxic metal exposure, respectively. Themetabo lite profiling is to be used also for crop plant biotechnology.

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tabolic profiling.The development of tools for theautomatic data extraction and processing make itan invaluable tool for addressing a myriad of bio-logical questions.

To date, the profiling in its current form has beenused mostly for studying metabolic changes in re -sponse to excess metals and for elucidating pa -thogen resistance by analyzing an Arabidopsismutant with partial loss of non-host resistance.Extensive analysis of different alleles has allowedto identify metabolic changes caused by loss-of-function of the gene in question.

Besides trying to answer biological questions con-siderable effort is still invested in further im pro -ving the profiling platform with regard to scope,

the establishment of data basesand the ease and speed of dataanalysis. Also, the fo cus of the me -tabolite profiling has re cently beenextended to rapeseed biotechno-logy within the frame of theGABI 2 program.

The profiling of Arabidopsis pro-teins is based on large-format high-resolution two-dimensional gel elec-trophoresis (2D-PAGE). Following the improve-ment of image analysis this technique has mostlybeen ap plied to identifying proteins that are sho-wing changes in abundance upon expression ofbacterial avirulence genes in Arabidopsis leaves.

Collaborators

Jeff DanglUniversity of North Carolina, Chapel Hill,USA

Gunhild Leck bandNorddeutsche Pflanzenzucht Hans Georg Lembke KG, Hohenlieth,Germany

Volker LipkaUniversity of Tübingen Germany

Paul Schulze-LefertMax Planck Institute for Breeding ResearchKöln, Germany

PublicationsClemens, S. & Simm, C. Schizo sac cha ro my -ces pombe as a model for metal ho me os -ta sis in plant cells: The phytochelatin-dependent pathway is the main Cd detoxi-fication mechanism. New Phytol. 159, 323-330.

Genger, R. K., Brown, A. H. D., Knogge, W.,Nesbitt, K. & Burdon, J. J. Develop ment ofSCAR markers linked to a scald resistancegene derived from wild barley. Euphytica134, 149-159.

Göbel, C., Feussner, I. & Rosahl, S. Lipidperoxidation during the hypersensitive re -sponse in potato in the absence of 9-lip-oxygenases. J. Biol. Chem. 278, 52834-52840.

Kroj, T., Rudd, J. J., Nürnberger, T., Gäbler, Y.,Lee, J. & Scheel, D. Mitogen-activated pro-tein kinases play an essential role in oxida-tive burst-independent expression of pa -thogenesis-related genes in parsley. J. Biol.Chem. 278, 2256-2264.

Legatzki, A., Franke, S., Lucke, S., Hoff mann,T., Anton, A., Neumann, D. & Nies, D. H.First step towards a quantitative modeldescribing Czc-mediated heavy metal resi-stance in Ralstonia metallidurans.Biodegra dation 14, 153-168.

Maier, T., Yu, C., Küllertz, G. & Clemens, S.Localization and functional characteriza -tion of metal-binding sites in phytochelatinsynthases. Planta 218, 300-308.

Markovic-Housley, Z., Degano, M., Lam ba, D.,von Roepenack-Lahaye, E., Cle mens, S., Su -sani, M., Ferreira, F., Scheiner, O. & Brei ten -eder, H. Crystal structure of a hypoallergenicisoform of the major birch pollen allergenBet v1 and its likely biological function as a

plant steroid carrier. J. Mol. Biol. 325, 123-133.Steiner-Lange, S., Fischer, A., Böttcher, A.,Rouhara, I., Liedgens, H., Schmelzer, E. &Knogge, W. Differential defense reactionsin leaf tissue of barley in response to infec-tion by Rhynchosporium secalis and totreatment with a fungal avirulence geneproduct. Mol. Plant-Microbe Interact. 16,893-902.

van't Slot, K. A. E, van den Burg, H. A.,Kloks, C. P. A. M., Hilbers, C. W., Knogge,W., & Papavoine, C. H. M. Solution struc -ture of the fungal disease resistance-trig-gering protein NIP1 from the fungus Rhyn -chosporium secalis shows a novel b-sheetfold. J. Biol. Chem. 278, 45730-45736.

Varet, A., Hause, B., Hause, G., Scheel, D. &Lee, J. The Arabidopsis NHL3 gene encodesa plasma membrane protein and its overex-pression correlates with increased resis -tance to Pseudomonas sy ringae pv. tomatoDC3000. Plant Physiol. 132, 2023-2033.

Wysocki, R., Clemens, S., Augustyniak, D.,Golik, P., Maciaszczyk, E., Tamás, M. J. &Dziadkowiec, D. Metalloid tolerance ba sedon phytochelatins is not functionally equi-valent to the arsenite transporter Acr3p.Biochem. Biophys. Res. Commun. 304, 293-300.

Books and Book chaptersClemens, S., Simm, C. & Maier, T. Heavymetal binding proteins and peptides. In:Bio polymers, Vol. 8 Polyamides and com-plex pro teinaceous materials II (Stein bü -chel, A. & Fahnestock, S. R., eds.) Wiley-VCH, Wein heim, pp. 255-288.

Lee, J. & Nürnberger, T. Is pore formationactivity of HrpZ required for defence acti-vation in plant cells? Pseudomonas sy -

ringae pathovars and related pathogens. In:De ve lopments in Plant Pathology 10 (Ia -cobellis, N.S. et al., eds.) Kluwer Aca de micPub lishers Dordrecht, pp. 165-173.

Neumann, D. Silicon in plants. In: Progressin Molecular and Subcellular Biology 33(Müller, W. E. G., ed.) Springer-Verlag Ber linHei del berg, pp. 149-160.

PatentsLipka, V., Schulze-Lefert, P., Scheel, D., Ro -sahl, S. & Landtag, J. Polynucleotides enco-ding a beta glucosidase. European Patent EP 2003000743388International Patent PCT/EP03/02315Canadian Patent 2,475,572Australian Patent 3227037.

Diploma TheseEngelhardt, Stefan: Struktur- und Funk -tions analyse des bakteriellen Effektor -proteins HrpZ aus Pseudomonas syringaepv. phaseolicola. University of Halle-Wit -tenberg, Department of Biochemistry /Biotechno logy, 17/07/2003.

HabilitationsClemens, Stephan: Molekulare Mecha nis -men der Aufnahme, Detoxifizierung undAkkumulation von Metallen. Univer sity ofHalle-Wittenberg, Faculty of Ma thematics,Natural Sciences and Techno logy, Depar -tment of Biology, 22/10/2003.

Nürnberger, Thorsten: Signalerkennung-und Signalumwandlungsprozesse bei derAuslösung nichtkultivar-spezifischer Pa -thogenabwehrmechanismen in Pflanzen.University of Halle-Wittenberg, Faculty ofMathematics, Natural Sciences and Tech -nology, Department of Bioche mistry/Bio -technology, 16/09/2003.


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Department of Stress and

Developmental Biology

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PublicationsAhlfors, R., Macioszek, V., Rudd, J., Bro sché,M., Schlichting, R., Scheel, D., & Kan gasa jär -vi, J. Stress hormone-independent activa -tion and nuclear translocation of mitogen-activated protein ki nases (MAPKs) in Ara -bidopsis thaliana during ozone exposure.Plant J. 40, 512-522.

Gao, L. L., Knogge, W., Delp, G., Smith, F. A.& Smith, S. E. Expression patterns of de -fense-related genes in different types ofarbuscular mycorrhizal (AM) developmentin wild-type and mycorrhiza-defective mu -tant of tomato. Mol. Plant-MicrobeInteract. 17, 1114-1125.

Halim V., Hunger A., Macioszek V., LandgrafP., Nürnberger T., Scheel D., & Rosahl S. Theoligopeptide elicitor Pep-13 induces salicy-lic acid-dependent and -independent de -fense reactions in potato. Physiol. Mol.Plant Pathol. 64, 311-318.

Harada, E., von Roepenack-Lahaye, E. &Clemens, S. A cyanobacterial protein withsimilarity to phytochelatin synthases cata-lyzes the conversion of glutathione to g-glutamylcysteine and lacks phytochelatinsynthase activity. Phy to chemistry 65, 3179-3185.

Lee, J., Rudd, J. J., Macioszek, V. K. & Scheel,D. Dynamic changes in the localization ofMAP kinase cascade components con trol -ling pathogenesis-related (PR) gene ex -pression during innate immunity in parsley.J. Biol. Chem. 279, 22440-22448.

Nickstadt, A., Thomma, B. P. H. J., Feuss ner,I., Kangasjärvi, J., Zeier, J., Loeffler, C.,Scheel, D. & Berger, S. The jasmonate-in -sensitive mutant jin1 shows increased re -sistance to biotrophic as well as ne cro -trophic pathogens. Mol. Plant Pathol. 5,425-434.

Schürch, S., Linde, C. C., Knogge, W., Jack -son, L. F. & McDonald, B. A. Molecular po -pulation genetic analysis differentiates twovirulence mechanisms of the fungal aviru-lence gene NIP1. Mol. Plant-MicrobeInteract. 17, 1103-1113.

Takeda, S., Tadele, Z., Hofmann, I., Probst A.V., Angelis, K. J., Kaya, H., Araki, T., Men giste,T., Mittelsten-Scheid, O., Shibahara, K.,Scheel D. & Paszkowski, J. BRU1, a novellink between responses to DNA damageand epigenetic gene silencing in Arabi -dopsis. Genes & Development 18, 782-793.

von Roepenack-Lahaye, E., Degenkolb, T.,Zerjeski, M., Franz, M., Roth, U., Wess jo -hann, L., Schmidt, J., Scheel, D. & Cle mens,S. Profiling of Arabidopsis secondary me -

tabolites by capillary liquid chromatogra-phy coupled to electrospray io nizationquadrupole time-of-flight mass spec tro -metry. Plant Physiol. 134, 548-559.

Weber, M., Harada, E., Vess, C., von Roe -penack-Lahaye, E. & Clemens, S. Com para -tive microarray analysis of Ara bi dop sisthaliana and Arabidopsis halleri rootsidentifies nicotianamine synthase, a ZIPtransporter and other genes as potentialmetal hyperaccumulation factors. Plant J.37, 269-281.

Publications in PressLi, C.-M., Haapalainen, M., Lee, J., Nürn ber -ger, T., Romantschuk, M. & Taira, S. Harpinof Pseudomonas syringae pv. phaseolicolaharbors a protein binding site. Mol. Plant-Microbe Interact.

Books and Book chaptersScheel, D. & Nürnberger, T. Signal transduc-tion in plant defense responses. In: FungalDisease Resistance in Plants. Bio che mistry,Molecular Biology, and Genetic Engi -neering. (Punja, Z. K., ed.) The HaworthPress, Inc., Binghamton, U.S.A., pp. 1-30.

Rosahl, S. & Feussner, I. Oxylipins. In: PlantLipids: Biology, Utilisation and Mani pu -lation. (Murphy, D., ed) Blackwell Pub -lishing, Ox ford, pp. 329-354.

Diploma ThesesBährecke, Annegret: Funktionelle Cha rak -terisierung von ZIP-Transportern aus Ara -bidopsis thaliana und Arabidopsis halleri.Uni versity of Halle-Wittenberg, Depart -ment of Biology, 19/05/2004.

Bethke, Gerit: Analyse der Protein-Pro -tein-Interaktionen zwischen Kompo nen -ten aus Arabidopsis MAP-Kinase-Kas ka -den mittels Fluoreszenz-Reso nanz-Ener -gietransfer (FRET). University of Halle-Wittenberg, Department of Biology,13/12/2004.

Bretschneider, Frank: Zellbiochemie derPhytochelatinsynthase aus Schizosac cha ro -myces pombe. University of Halle-Wit ten -berg, Department of Biochemistry/ Bio -technology, 15/09/2004.

Eschen-Lippold, Lennart: Entwicklung ei -ner Realtime-PCR basierten Methode zurBestimmung des Wachstums von Phy toph -thora infestans auf Kartoffel pflan zen mitunterschiedlicher Resistenz. University ofHalle-Wittenberg, Depart ment of Biology,06/12/2004.

Franz, Sandra: Charakterisierung von Ge -nen, die in Microarray-Experimenten als pu -tative Metallakkumulations fakto ren inArabi dopsis halleri identifiziert wurden.

University of Halle-Wittenberg, Depart -ment of Bio che mistry /Biotech nology,20/09/2004.

Fritsche, Thomas: Funktionelle Cha rak te ri -sierung der MAP-Kinase MPK11 aus Ara -bi dopsis thaliana. University of Halle-Wit -tenberg, Department of Bioche mi stry /Biotechnology, 15/09/2004.

Köster-Eiserfunke, Nora: Analyse von pu -tativen Phosphoproteinen in transgenenArabidopsis-Pflanzen mit erhöhter MAP -Kinase-3 und-6 Aktivität. University of Hal -le-Wittenberg, Department of Bioche mis -try/Biotechnology, 09/12/2004.

Müller, Claudia: Struktur- und Funk tions a -nalyse einer mikrobiellen Transglutami naseaus Phytophthora sojae. University ofHalle-Wittenberg, Department of Bio che -mis try/ Biotechnology, 22/01/2004.

Schwedt, Anne: Molekulare Charak teri sie -rung einer Rezeptorkinase aus Ara -bidopsis thaliana. University of Halle-Wittenberg, Department of Bioche -mistry/Biotech no lo gy, 09/07/2004.

Spielau, Claudia: Grundlagen der Analysevon Proteinkinase-Komplexen mittels Tan -dem Affinity Purification (TAP) aus trans-genen Arabidopsis thaliana. Univer sity ofHalle-Wittenberg, Department of Bio che -mistry/Biotechnology, 15/09/2004.

Unger, Christiane: Beteiligung von Lipi denan der Pep-13-initiierten Signal trans duk -tion in Petersilie. University of Halle-Wit -tenberg, Department of Bio logy, 12/01/2004.

Unthan, Tino: Untersuchungen zur Rolleder Nicotianamin-Synthese für die Me tall -hyperakkumulation. University of Halle-Wittenberg, Department of Bio chemis try/Biotechnology, 20/09/2004.

Doctoral ThesesHofmann, Ingo: Erzeugung, Isolation undCharakterisierung von Suppressor mu -tanten für Transcriptional Gene Silen cingin Arabidopsis thaliana. University ofHalle-Wittenberg, Department of Bio -chemistry/ Biotechnology, 03/11/2004.

Hunger, Astrid: Die Rolle von Jasmonat,12-Oxophytodiensäure (OPDA) und Sa li -cylat bei der induzierten Resistenz in Kar -toffel. University of Halle-Witten berg, De -part ment of Biochemistry/ Biotechnology,20/12/2004.

Simm, Claudia: Das Metallothionein Zym1aus Schizosaccharamyces pombe - Unter -su chungen zur Funktion des Me -tallothineins in einem phytochelatin-bil-denden Orga nis mus. University of Hal le-


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Department of Secondary MetabolismHead: Dieter Strack, ProfessorSecretary: Heidemarie Stolz until October 2004, afterwards Ildikó Birkás

Department of

Secondary Metabolism

Work of the department is concerned with the molecular regulationof plant secondary metabolism, evolution of the enzymes involved

and the role of secondary products in interactions of plants with their en -vironment.

The work on metabolic regulation includes isolation and characterizationof the enzymes and the encoding genes, focusing on transferases. We cur-rently investigate malate and choline hydroxycinnamoyltransferases as wellas several hydroxycinnamate glucosyltransferases from Arabidopsis thalia-na and oilseed rape (Brassica napus) as well as flavonoid and betanidin glu-cosyltransferases from betacyanin-accumulating plants or flavonoid methyl-transferases from the ice plant (Mesem bryan themum crystallinum).

The aim of the work on glucosyl- and hydroxycinnamoyltransferases is toelucidate their evolutionary origin and structure-function relations to pre-dict substrate specificity. Glucosyltransferases involved in betacyanin bio-synthesis are considered to be oligophyletic and originate from differentclusters of flavonoid glucosyltransferases. A new class of methyltransfera-ses was identified, whose substrate specificities can be modulated by metalcations and N-terminal deletions. Hydro xy cinna moyl transfe ra ses, whichare dependent on b-acetal esters as acyldonors, are vacuolar serine carb -oxypeptidase-like (SCPL) proteins as found for the enzyme involved in theformation of si napoylmalate in Arabidopsis. This plant harbors a gene fami-ly encoding 53 SCPL proteins, of which 21 cluster to a distinct group ofSCPL acyltransferases.The existence of such acyltransferases proves a newconcept in the regulation of plant secondary metabolism.

Special emphasis is also placed on programs focusing on the molecularinteractions of plants with arbuscular mycorrhizal fungi. The work of twogroups is concerned with fungus-induced alterations in plant isoprenoidmetabolism, in particular carotenoid biosynthesis and degradation, accom-panied by a dramatic reorganization of plastid population in arbuscule-har-boring root cells. Another main objective is the analysis of the role of phy-tohormones, in particular jasmonates, in development and func tional main-tenance of mycorrhizal symbiosis. It was found that jasmonates obviouslyplay a crucial role in the esta-blishment of arbuscular my -corrhizas. These studies aresupported by comprehensiveanalysis of primary and secon-dary metabolites (metaboliteprofiling) in wild type andtransgenic mycorrhizal plants.

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Research Group: Molecular Physiology of MycorrhizaHead: Michael H. Walter

Two classes of apocarotenoids (ca -ro tenoid cleavage products) accu-mulate in the roots of various plantsupon colonization by mycorrhizal fun - gi. These are linear C14 mycorradicinand glycosylated C13 cyclohexenonederivatives, which are predicted tooriginate from a common C40-ca -rotenoid or xanthophyll precursor.The first step of plastidial carote-noid precursor biosynthesis in theMEP pathway is catalyzed by 1-de -oxy-D-xylulose 5-phosphate syn tha se(DXS). We have discovered dupli -cate genes for DXS in Medi cagotruncatula, which are distantly rela-ted and differentially regulated. Thegeneral concept of dedica-ted roles of DXS1 in thebiosynthesis of primaryiso prenoids (e.g. leaf ca ro -

te noids) and of DXS2 in secondaryisoprenoid formation holds for allplants investigated so far. The onlyexception is Arabidopsis thaliana,which has two DXS1 co pies but noDXS2 copy. Ex amples for DXS2-corre-lated biosynthesis of se condary isopre-noids are the mo no terpenes of pep -per mint leaf trichomes, the steviolditerpenes of Ste via rebaudiana leavesand the terpenoid indole alkaloids ofCatharanthus roseus cells. The geneduplication and subfunctionalizationleading to this diversification must bevery old, since we have shown thatduplicate DXS genes occur in gym -nosperms by data base mi ning of ESTs

and by isolating a spruce DXS1 cDNA clone.Future work will try to trace back this geneduplication even further in evolution.

Current experiments on the angiosperm Me di ca -go truncatula have identified an additional geneduplication of DXS2 in this system.The duplicateparalogous genes MtDXS2-1 and MtDXS2-2 arealmost identical in coding regions, similar in in -tron and proximal promoter regions and occur asa tandem repeat.These characteristics argue for arecent duplication event. Both genes are ac ti va -ted during mycorrhization. Possible early sub-functionalization features (alterations in ex pres -sion) of MtDXS2-2 are currently being investiga-ted. An RNAi strategy has been devised todown-regulate both genes concomi tantly and

Group Members

Daniela Floß PhD Student since April 2004

Joachim Hans Postdoctoral Position until October2004

Kerstin Manke Technician

Collaborators

Harro Bouwmeester Plant Research International,Wageningen, The Netherlands

Jörg DegenhardtMax Planck Institute for ChemicalEcology, Jena, Germany

Thomas Fester, BettinaHause, Willibald Schliemann,Jürgen SchmidtLeibniz Institute of Plant Biochemistry,Halle, Germany

Franziska KrajinskiUniversity of Hannover, Germany

Arbuscular mycorrhizas are mutualistic symbiotic associations between a small number of specializedfungi and the roots of most terrestrial plants. The fungi deliver mineral nutrients to the plant and recei-ve carbohydrates in return. The focus of the group is plant isoprenoid biosynthesis and its molecularregulation in mycorrhizal roots. Starting from an analysis of mycorrhiza-specific apocarotenoids speci-fic gene targets in the plastidial non-mevalonate methylerythritol phosphate (MEP) pathway and incarotenoid cleavage reactions have been selected. In the first step of the MEP pathway, catalyzed by1-deoxy-D-xylulose 5-phosphate synthase (DXS), an ancient gene duplication has evolved into specia-lized DXS genes for the generation of primary and secondary isoprenoids. The my corrhiza-regulatedDXS2 gene has duplicated again recently and occurs as a tandem repeat in the genome of the modellegume Medicago truncatula.

Changes in root color of Medicago truncatula plants upon colonization by mycorrhizal fungidue to the accumulation of apocarotenoids, one of them being the "yellow pigment" (mycor-radicin derivatives). The inset shows an RNA blot indicating elevated transcript levels of thebiosynthetic enzyme MtDXS2 exclusively in mycorrhizal roots (R, M).

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cre ate loss-of-function mutants devoid of apoca-rotenoid accumulation without interfering withbiosynthesis of primary carotenoids in leaves.TheRNAi-constructs are expressed in roots of Me -dicago truncatula after transformation via Agro -bacterium rhizogenes. Transformed roots sprea-ding from the injection site of bacteria are recog-nized by the fluorescent marker DsRed and canthus be fur ther propagated, while non-transgenicroot parts are discarded. Preliminary re sults indi-cate a significantly lower concentra tion of mycor-radicin de rivatives in mycorrhizal roots transge-nic for the RNAi construct as compared to con-trol mycorrhizal roots. In addition, MtDXS2-1promoter-GUS fusions and promoter deletion-GUS fusions are being created to eventually iden-tify promoter elements relevant to my corrhizalregulation of plant genes.

A recent gene duplication of DXS2 has occurredalso in the genome of African marigold (Tageteserecta). Amplification of genomic fragments fromT. erecta DXS2 genes indicates very strong simi-larity of two TeDXS2 genes similar to theMtDXS2 duplicates. Comparative analyses haveidentified the DXS2 duplicates in six out of sevenTagetes species and cultivars. On ly T. tenuifoliaappears to lack the second gene copy. By lowe-ring the stringency in PCR amplification experi-ments a dis tantly related DXS1 fragment couldbe amplified from this species but never a second

DXS2 copy. The second step of the MEP pathway cata -lyzed by 1-deoxy-D-xylulose 5-phosphate re -duc toisomerase (DXR) is or ga nized in amore simple fashion than DXS. At least inmaize, rice, tomato, peppermint and Catha -ranthus roseus only single genes have beenidentified for DXR. A single maize DXR geneex hibits elevated transcript levels in leaves aswell as in mycorrhizal roots. The generationof antibodies created from recombinantmaize DXR has allowed for detailed immuno-localization studies on maize DXR in arbus-cule-containing mycorrhizal roots. The resultsdemonstrate an extensive proliferation ofDXR-containing plastids during mycorrhiza -tion, which form a network of interconnec-ted plastids (stromules) around the arbus -cules in co lonized maize root cells. The DXRac cumulation reaches its maximum at orslightly af ter the peak of arbuscule matura -tion and declines thereafter indicating a closecorrelation of (apo)carotenoid formation withthe arbuscular life cycle.

Carotenoid cleavage constitutes a late step in apo -carotenoid biosynthesis requiring regiospecific di -oxygenases. A carotenoid cleavage enzyme (CCD)clone with specificity for the 9,10 (9',10') doublebond creating C13 and C14 products has been isola-ted from maize. Again, a single gene appears to beresponsible for elevated transcript levels in leavesand in mycorrhizal roots. Ex pression of the maizeCCD in E. coli strains engineered to produce caro-tenoids leads to a discoloration of bacterial colo-nies. However, product identification after incuba -tion of recombinant CCD with carotenoid substra-tes has not yet been accomplished.

M. truncatula plant lets injectedwith Agro bac te rium rhizogenesbelow the hy po cotyl develop newtrans genic hairy roots at the injectionsi te (a bo ve, plant to the right).Trans genic roots of M. truncatulaflu o resce in red due to ex pres singthe fluorescent mar ker DsRed

Highly branched fungal arbuscule in a mycorrhizal maizeroot cell visualized by WGA-TRITC staining (red). Im -munolocalization of DXR (green) reveals strong prolifera -tion of plastids around the arbuscule. DNA in plant nucleusand fungal hyphae is shown in blue by DAPI staining.

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Research Group: Cell Biology of MycorrhizaHead: Bettina Hause

A possible role of jasmonates in themycorrhizal interaction is indicatedby the following data: Jasmonic acid(JA), applied exogenously, promotescolonization and development of my -corrhizal structures, and the en do -genous JA level of mycorrhizal rootsis remarkably higher than that ofnon-mycorrhizal roots.The in creaseof JA content in barley roots uponmycorrhization is accompanied bythe expression of genes coding forenzymes of JA biosynthesis and forjasmonate-induced proteins.

To analyze the function of JA duringthe development of the AM symbio-sis, we intend to perform functionalanalyses by modulating the JA con-tent in mycorrhizal roots of M.truncatula. Two cDNAs coding forthe allene oxide cyclase (AOC), theen zyme performing the crucial stepin JA biosynthesis, were isolatedfrom M. truncatula. The AOC pro-tein was localized to plastids andfound to occur constitutively in allvascular tissues of M. truncatula. Inleaves and roots, MtAOC1 isexpressed upon JA application.Enhanced expression accompaniedby elevated JA levels was alsoobserved during mycorrhizationwith G. intraradices. A partial sup -pression of MtAOC1 expressionwas achieved in roots followingtransformation with Agrobacteriumrhizogenes harboring the MtAOC1cDNA in antisense direction under

control of the CaMV35S promoter leading tochimeric plants with wild-type shoots and trans-formed roots. In comparison to samples transfor-med with a control vector (35S::uidA), roots withsuppressed expression of MtAOC1 ex hibitedlower JA levels and a remarkable delay in the pro-cess of colonization with G. intraradices. Boththe mycorrhization rate, quantified by fungalrRNA, and the formation of arbuscules, analyzedby the expression level of the AM-specific geneMtPT4, were affected. In the future work, analy-ses of transcript and metabolite patterns bycDNA microarrays (collaboration with H. Küster,Uni versity of Bielefeld) and metabolite profiling(collaboration with W. Schliemann, IPB Halle),both in wild-type and transgenic roots, will provi-de answers to the question, which processes ofmycorrhization are mediated by JA.

Because higher levels of JA within mycorrhizalroots could be caused by the higher sink func tionof such roots, transgenic tobacco (Nicotiana taba-cum) plants specifically modulated in root su garmetabolism were studied as well. For this, tobaccoplants expressing a yeast-derived apoplast-di rec tedinvertase under the control of an alcohol-induciblepromoter (alc) were used. Soil-drenching with aqu -e ous acetaldehyde solutions resulted in a rapid andtemporary induction of the alc gene ex pressionsystem exclusively in roots. In ad dition, the split-root system allowed an activation restricted to thetreated part of the root. In duc tion of the invertaseupon treatment with acetaldehyde led to a changein the ratio of suc rose to hexoses specifically wit-hin the roots. To include in vertases targeted to thecytoplasm and vacuole in our analyses, suitablevectors were used for root transformation of to -bacco. All the change described, how ever, did notlead to alterations in AM colonization according to

Plant hormones are believed to play a role in the establishment and development of sym-biotic interactions between plants and arbuscular mycorrhizal (AM) fungi. Jasmonates,known as regulators in plant response to biotic or abiotic stress, are good candidates forsuch a role. Therefore, the functional ana lysis of jasmonates during the interaction bet-ween Glomus intraradices and barrel medic (Medicago truncatula) is the main objectiveof our group. In a second project, the massive proliferation of plastids in AM colonizedroot cortical cells is studied. Metabolic and cytological changes as well as respectivemolecular mechanisms will be elucidated. In a third project, we are analyzing the influen-ce of epothilones on the structure of plant microtubules.

Group Members

Sabine BothTrainee until November 2004

Ana Maria Cencano DAAD Fellow until November 2004

Thomas Fester Leader Junior Group

Ulrike Hintsche/HuthTechnician

Stanislav Isayenkov Postdoctoral Position until October2004

Nina Kreißig Student until June 2004

Sandra Lischewski Trainee until March 2004

Swanhild LohsePhD Student

Cornelia MroskPhD Student since July 2004

Daniel PeiskerStudent until February 2004

Silke PienknyStudent until May 2003

Sara Schaarschmidt PhD Student

Rostand Tonleu TonfackDiploma Student since November 2004

Carola TretnerResearch Scientist

Gerlinde Waiblinger Technician until July 2004

Chimeric M. truncatula plant after roottransformation with 35S::uidA. The plantwas processed for GUS staining showingthe transgenic part of the root in blue.

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microscopical, metabolite and transcript analy-ses. Plastids in colonized root cortical cells are cha-racterized by an active carotenoid biosynthesisand by a strong proliferation close to fungalstructures. In the current project we have beenlooking for further changes in plastid metabo-lism, for a functional reason for the activationof carotenoid biosynthesis and for mechanismsinvolved in changing plastid number and shape.Transcript levels of the genes referring to themain plastid biosynthetic pathways have beenanalyzed by "Electronic Northern" and real -time RT-PCR. Increased transcript levels havebeen observed in particular for genes involvedin fatty acid and asparagine biosynthesis. Thesefindings correlate with data from the researchgroup Biochemistry of Mycorrhiza, indicatingin creased levels of metabolites of the two res -pec -tive pathways. Regarding carotenoid bio-synthesis, we are currently studying a possibleconnection of the activation of this pathway tothe accumulation of reactive oxygen species(ROS) in AM roots. AM roots from transgenictobacco plants containing reduced levels ofcatalase (cooperation with Frank vanBreusegem, Uni versity of Gent) accumulateincreased amounts of apocarotenoids. A linkbetween ca rotenoid biosynthesis and the pre-sence of ROS is fur ther suggested by the fin-ding that both processes occur predominantlyin the phase of arbuscule degradation.Regarding the analysis of plas tid number andshape, we have analyzed M. truncatula after A.rhizogenes-mediated root transformation usinga construct targeting the green fluorescentprotein to plastids. A significant proliferation ofthis compartment was ob served in colonized

root cortical cells. The extent ofnetwork formation, however, see-med to be less pronounced whencompared to tobacco. Fur theranalyses re gard the characteriza-tion, lo cali za tion and functionalanalysis of molecular markers in -volved in processes like plastiddivision or plastid deformation. Inthe case of plastid division, wehave chosen the plastid divisionprotein FtsZ for such an analysis.First results from immunolocali-zation of this protein suggest thatthe division of plastids is connec-ted to a specific phase of arbuscu-lar development. An antisense ap -proach re gar ding this protein willreveal the functional significanceof plastid division for the AMsymbiosis in general and for pla-stid biosynthetic metabolism inparticular. Re garding a similar analysis of plastiddeformation, we are currently searching forsuitable proteins in volved in this process.

Our group is further interested in studying theinfluence of epothilones, macrocyclic lactonesfrom culture filtrates of the myxobacterium So -rangium cellulosum, on the structure of plantmi crotubules (MTs). Stability and dynamics ofMTs are influenced by natural compounds liketaxol, the most prominent example for inhibi-tors of MT depolymerization. Epothilone andits derivatives are known as taxol-like microtu-bular drugs in human medicine, but their effectson plants are still unknown. We have analyzedthe effect of Epo thilone D on the plant cell-cycle in cultivated cells.The treatment of toma-to cell suspension cultures resulted in a conti-nuous in crease of the mitotic index, visible al -ready after two hours. As shown by immuno-cytological methods, abnormal spindles are for-med during metaphase leading to a random dis-tribution of chromosomes in the whole cellwithout formation of a metaphase plate. Theeffects of Epo thilone D seem to be irreversible,because cells with an abnormal spindle couldnot be recovered after removal of the drug.

Confocal laser scanning mi cro graph of an arbuscule (Ni co tia na taba-cum colonized by Glomus in tra radices) after staining for reactiveoxygen spe cies using dihydrorhodamine-123. Bar represents 20 µm.

Invertase in situ staining of crosssec tions of alc::cwINV- to baccoroot tips indicates a high inver-tase activity in all root cells afterinduction of the yeast-derivedinvertase by soil-drenching with0.05 percent acetaldehyde (rightside).Wa ter-treated control plantsshow only a weak purpur staining(left side). Bars represent 25 µm.

Immunofluorescence images oftomato cells in metaphase sho-wing the microtubular patternlabelled with anti-a-tubulin an ti -body (green) and the distribu -tion of chromosomes stainedwith DAPI (blue) in non-treatedcells (a) and cells treated with1.5 µM Epothilone D for 8 h (b).Bar represents 5 µm for bothmicrographs.

Collaborators

Frank van BreusegemUniversity of Gent, Belgium

Ivo Feussner, Katharina PawlowskiUniversity of Göttingen, Germany

Philipp FrankenInstitute of Vegetables and Ornamental Crops,Großbeeren, Germany

Gerd HauseUniversity of Halle, Germany

Helge KüsterUniversity of Bielefeld, Germany

Thomas RoitschUniversity of Würzburg, Germany

Uwe SonnewaldInstitute of Plant Genetics and Crop PlantResearch, Gatersleben, Germany

Claus Wasternack, Otto Miersch, Willibald Schliemann, Ludger WessjohannLeibniz Institute of Plant Biochemistry, Halle,Germany

Victor WrayGerman Research Centre for Biotechnology,Braunschweig, Germany

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Research Group: Biochemistry of MycorrhizaHead: Willibald Schliemann

On the basis of metabolite profilingresults of preliminary mycorrhiza -tion kinetics an extended mycor-rhization experiment was per -formed including additional con-trols with plants fully provided withphos phate.This should allow to dif-ferentiate between metabolite alte-rations induced by phosphate onlyfrom those caused by mycorrhizalin teraction. Furthermore, additio-nal harvest time points and paral-lels enable a more reliable statisti-cal evaluation of the metabolite da -ta obtained by GC/MS, LC/ESI-MS and DAD-RP-HPLC.For a more substantial me -tabolite identification, theNIST MS-database was sup-plemented by the plant-spe-

cific MSRI database (collaboration withJoachim Kopka, MPI of Mole cular PlantPhysiology, Golm). The large data sets aresubjected to a detailed quantitative analy-sis requiring self-programmed tools forautomated data processing and visualiza -tion.The utilized analytical techniques andbioinformational tools are the key ele-ments of our metabolite profiling plat-form. In the course of preprocessing theraw data are a ligned, smoothed, averagedand graphically summarized for re cog ni -zing general trends. Qualitative and quan-titative differences in the chromatogramsare revealed in calculated differential plots.The quantitative (integrated) me ta bolitedata are similarly preprocessed as the rawdata. In subsequent steps the data aresubjected to various statistical me thodsto reveal correlations and interactions be -

t ween the metabolites at different inoculationstages during mycorrhization (Pearson correla -tion and network analysis, cluster, principal com-ponent and regression analysis). The re sults ob -tained up to now refer to a general activation ofmi tochondrial (tricarboxylic acid cycle) as well asplastid metabolism (lipid biosynthesis, N-assimila-tion) during mycorrhization, which correlateswith corresponding increased transcript levels (insilico and real-time RT-PCR analyses-collabora -tion with Thomas Fester, IPB Halle). Furthermore,higher Pearson correlation coefficients of meta-bolite pairs in mycorrhizal root extracts (M) indi-cate a closer metabolic relationship among them-

Besides transcriptomics and proteomics, metabolomics as a further functional genomic approach cha-racterizes a biological system at a given time. Our research focus is arbuscu lar mycorrhiza originatingfrom the interaction of fungi (Glomeromycota) with the roots of the majority of plants. In a project ofthe DFG focus program 1084 Molecular Basics of Mycorrhizal Symbioses the pattern of primary andsecondary metabolites of developing ar buscular mycorrhizal roots of Medicago truncatula colonizedwith Glomus intraradices are analyzed by metabolite profiling. The ultimate aim is the characterizationof causal relationships between mycorrhiza-specific gene expression and metabolite profiles to assignprocesses relevant for mycorrhiza development and symbiotic functioning.

Metabolite profiling platform: analytical methods and bioinformatio-nal tools

Group Members

Christian Ammer Research Scientist

Barbara Kolbe Technician

CollaboratorsHermann Bothe, Ulrich HildebrandtUniversity of Cologne, Germany

Bettina Hause, Thomas Fester, Alfred Baumert, Jürgen Schmidt,Martina LerbsLeibniz Institute of Plant Biochemistry,Halle, Germany

Joachim KopkaMax Planck Institute of Molecular PlantPhysiology, Golm, Germany

Pål Axel OlssonUniversity of Lund, Sweden

Lloyd W. SumnerSamuel Roberts Noble Foundation,Ardmore, USA

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selves than in non-mycorrhizal controls (NM).The accumulation of the fungus-specific palmit-vaccenic acid (C16:1D11) can be detected as earlyas 14 days after inoculation (dai), whereas thefungal trehalose and the my corrhiza-induced apo-carotenoids (cyclohexenones, mycorradicin-de -rived compounds and the complex "yellow pig-ment") are accumulating continuously during thecomplete analysis period. The developed ratioplot for comparison of metabolite data from ave-raged HPLC runs of different sample sets allowsto detect wavelength-dependent differences be -tween them immediately. The controls suppliedwith increased concentration of phosphate didnot show any of these mycorrhiza-characteristicmetabolite altera tions. How ever, they showedstrong reduction or dis ap pea ran ce of at least two

c omp o n e n t s .The levels of sa -ponins and es pe -cially isoflavo noids(malonylononinas the main com -po nent) are slight-ly higher in my -cor rhizal than innon-my corrhi zalroots. The me ta -bolite profiling ap -proach is comple-mented by pre-parative isola tionof secondary compounds and their structural elu-cidation by MS and NMR.

HPLC profiles and accumulation of mycorrhiza-induced apocarotenoids MtCH1,2,3 - M. truncatula cyclohexenone derivative 1,2,3; MtApo1,2 - M. truncatula mycorradicin-derived compound 1,2

Retention time-wa ve length ratio plot of averaged root HPLC dataof my cor rhi zal plants (M)/100 % phosphate controls (100 % P)

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Research Group: Glycosyl- and MethyltransferasesHead: Thomas Vogt

Based on earlier work, our groupstarted on the molecular physiologyof betalains, but lately as a result ofthe ongoing work, the major focushas shifted towards two classes ofmodifying enzymes involved in themodification of the two classes ofplant natural products under inves-tigation, the betacyanins and the fla-vonoids. By detailed sequence analy-sis and substrate specificity studiesof two UGTs, the betanidinUGT73A5 and UGT71F2 from Do -rotheanthus bellidiformis (Dbs), wewere able to demonstrate an oligo-

phyletic origin of the corresponding glucosyl-transferase genes from different clusters of flavo -noid UGTs. Several lines of evidence, includingcloning of highly homologous UGT73A5 andUGT71F1 sequences from red beet (Beta vulga-ris), suggest that these position specific enzymesinvolved in betanidin glucosylation have derivedfrom common ancestors, flavonoid and other hy -droxylated phenylpropanoid-modifying enzymes.Site-directed mutagenesis, performed with theheterologously expressed UGT73A5 indicatedthe importance of a highly conserved N-terminalhistidine and a glutamate of the conserved UDP-glucose binding site to be catalytically essential.Ba sed on a crystal structure of the microbialUGT from Amycolatopsis orientalis, the first 3D-structure of UGT73A5 was modelled, and a newSN-1-like reaction mechanism proposed, whichconfirms the essential role of the N-terminal his -tidine as the catalytic base. Docking studies alsodemonstrated the substrate preference ofUGT73A5 for betanidin.

Our second model system, the ice plant (Mes -embryanthemum crystallinum), adapts to extre-me light intensities by the rapid accumulation ofglycosylated and methylated flavonol and beta-cyanin conjugates in leaf epidermal layers. The fla-vonoid-methylating activity (PFOMT) is in volvedin the accumulation of a variety of conjugatedquercetagetin 6-3 -diOMe derivatives upon lightinduc tion. Based on sequence identities and sub-strate specificities, it defines a new subgroup ofcation-dependent OMTs, characterized by abroad substrate specificity, previously as sumed tobe involved only in the methylation of the ligninprecursor caffeoyl coenzyme A. Light- inducible

transcript accumulation and localization of the

Group Members

Judith Hans PhD Student until March 2004

Dagmar Knöfel Technician

Collaborators

Wolfgang Brandt, JürgenSchmidt Leibniz Institute of Plant Biochemistry,

Modifications of plant natural products are performed by a variety of transferases with such promi-nent families like glucosyltransferases (UGTs) and methyltransferases (OMTs), which can share thesame substrates and lead to the observed diversity of plant secondary metabolites. These enzymesmay both solubilize and deactivate bioactive low-molecular weight compounds. Sequence identitiescluster the UGTs as well as the OMTs according to regiospecificities rather than strict substrate pre-ference. Predictions imply that both superfamilies of proteins have evolved oligophyletically as one ofthe primary adaptive mechanisms of plants to meet the changing environmental conditions in a timelyand developmentally controlled manner. Recent developments have focused to elucidate the structuralproperties of both classes of enzymes with specific emphasis on substrate and position specificities aswell as the reaction mechanisms.

Predicted tertiary structure of UDP-glucose: be tanidin 5-O-glucosyltransferase (UGT73A5) from Dorothe an thus belli-diformis with the docked ligands betanidin and UDP-gluco-se.

59

en zyme in epidermal tissues correlates with itsim portant role in phenylpropanoid accumulation.

Optimized heterologous expression resulted inhigh yields of recombinant PFOMT from E. coli,after purification used for subsequent crystalliza-tion. Crystals were obtained under various preci-pitant conditions and several data sets re cordedwith resolutions down to 1.4 Å, in case of therecombinant PFOMT and 1.8 Å for its seleno-methionine-derivative, respectively. Both datasetswere sufficient to result in a preliminary 3D-crys -tal structure of this cation-dependent enzyme.Like in case of the UGT, the N-terminal part ofthe enzyme is important for position specificity.Compared to the (N-terminally trun cated) nativeplant protein, with eleven ami no acids missing,the full length sequence showed an altered pro-duct profile, when assayed with the endogenoussubstrate quercetagetin. The profile of the re -combinant PFOMT-isoform, lacking tho se elevenamino acids is consistent with the observed pro-duct profile in vivo, whereas the full lengthrecombinant enzyme catalyzes the formation of

additional fluorescent flavonol 5-OMe derivati-ves, never observed in the plant. Tests with rela-ted cation-dependent OMTs revealed that notonly the ami no acid sequence but also that thenature of bivalent cation within the active site iscrucial for the position and substrate specificityof this class of enzymes.

Potential substrates of cation-dependent OMTs. 1, caffeoyl CoA; 2, quercetin; 3, caffeic acid;4, caffeoylglucose; 5, 3,4-dihydroxybenzoic acid;

Typical crystals of PFOMT obtained withpolyethylene glycol-based precipitants.

1 2

3 4 5

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Research Group: Hydroxycinnamic AcidsHead: Dieter Strack

Based on the isolation of cDNAs en -coding the enzymes for sinapine bio-synthesis, UDPglucose:sinapate gluco-syltransferase (BnSGT1) and sinapoyl-glucose : choline sinapoyltransferase(BnSCT) we analyzed the regulation ofsinapate ester metabolism in B. napus.BnSGT1 and BnSCT activities werefound to be subject to a pronouncedtranscriptional regula tion. BnSGT1transcript level in creases throughoutearly stages of seed development untilthe early cotelydonary stage and staysconstant in later stages. In two-dayold seedlings the BnSGT1 transcriptsreach their high est abundance follo-wed by a dramatic decrease. Ex pres -sion of BnSCT is restricted to seeds.

Enzyme activities in crude extracts and metabo -lite concentrations reflect the RNA abundance ofBnSGT1 and BnSCT indi cat ing that transcriptio-nal regulation of these genes is of major im -portance for regulation of sinapine biosynthesis.Southern-blot analyses revealed that the amphi-diploid ge nome of B. na pus contains at least twogenes for SGT and SCT, each derived from itsprogenitors B. rapa and B. oleracea. As the rela-ted genome of Arabidopsis was shown to encodebesides SGT three less-specific ester-forminghydroxycinnamic acid glucosyltransferases(HCA-GTs) we assume for B. napus the presenceof HCA-GT homologues po tentially in volved insinapine biosynthesis.

By homology-based cloning techniques two full-length cDNAs with strong sequence identity toHCA-GTs could be isolated from B. napus.When

expressed in E. coli, one of these enzymes dis-plays HCA-GT activity. From seeds varioussecondary compounds were isolated and identi-fied by MS and NMR as sinapic acid esters of glu-cose, gentiobiose and kaempferol glycosides,besides sinapine, sinapoylmalate and an unusualcyclic spermidine amide.

We started a transgenic approach to suppresssinapine accumulation. A dsRNAi construct de -signed to silence the BnSGT1 gene was used totransform B. napus (cv. Drakkar).Transgenic plantshomozygous for the insertion of the BnSGT1suppression cassette showed a severe reductionof sinapine content to about 20 percent of theun transformed plants accompanied by a strongde crease in total sinapate ester content.

HPLC analysis of seed extracts of the trans -formed plants revealed that the concentration ofsi napoylglucose and most of the the minor sina-pate esters wasunder the detec-tion level. This in -dicates that 1-O-sinapoylglucose isa central precur-sor not on ly forsinapine biosyn-thesis but also fortransacy la t ionreac tions lea dingto all the other si -napate es ters inB. napus seeds.As the si napoyl -t r a n s f e r a s e s

The work of our group is focused on biochemistry, genetics and molecular regulation of sinapate esterbiosynthesis in oilseed rape (Brassica napus) and the model plant Ara bi dopsis with the aim to improverapeseed quality by metabolic engineering.The sinapate esters sinapoylcholine (sinapine) and sinapoyl-glucose (SinGlc) are abundant compounds, known to accumulate in seeds of Brassicaceae plants. Toproduce the highly diverse pattern of metabolites, plants have evolved a wide array of enzymes to fulfillthe need for driving plant secondary metabolism. SinGlc-dependent acyltransferases like SCT belongto this group of enzymes. They are homologous to hydrolases of the serine carboxypeptidase type.Our research interest is aimed at elucidating the molecular changes necessary for conversion of pep-tidases to acyltransferases by analysis of structure-function relationships. Sinapate esters are regardedas antinutritive factors. In B. napus they prevent the valuable seed protein from being used as humanfood supplement, a significant reduction of sinapine content is therefore a key requirement for esta-blishing rape as a protein crop.

Transcript levels of BnSGT1 and BnSCT genes,enzyme activities and metabolite accumulation indeveloping rape seeds. Maximal enzyme activity wasset to 100% (SGT=127 pkat/mg protein; SCT= 185pkat/mg Protein); metabolite concentration is givenin nmol/seed. A-G = stages of seed development

61

known so far, such as SCT and SMT, have beenshown to dis play high ac cep tor spe ci ficity we hypo-thesize that each of the sinapate es ters should bepro du ced by a distinguished acyltransferase of theSCPL-type protein giving rise to a lar ge proteinfamily of SCPL acyltransferases in B. napus as it hasbeen shown for Arabidopsis. To optimize suppres-sion of sina pine synthesis in rape, additional vec-tors have been constructed making use of theCaMV 35S promoter for constitutive silencing ofSCT sequences for BnSCT to be suppressed or ofa double in verted repeat including BnSGT1 andBnSCT se quences for simultaneous silencing ofboth genes.

Energy-rich b-acetal esters like 1-O-sinapoylglu -cose have been characterized as acyl donors intransacylation reactions of plant secondary meta-bolism. In Arabidopsis, four related genes (AtSGTand AtHCAGT1-3) were found to encode gluco-syltransferases catalyzing the formation of hydro-xycinnamate glucose esters in vitro.We have star -ted a sys tematic approach including modulationof gene ex pression, mutant analysis and crossingas well as promoter testing to elucidate the functi-on of these genes in planta. In wild type plants, itcould be shown by real-time RT-PCR that all fourgenes are expressed during seed development.However, AtSGT-mRNA is the most abundantone and ex pression of AtSGT1 follows roughlythat of BnSGT1 in B. napus with highest expressi-on in two-day-old seedlings. To modulate geneexpression in Ara bi dopsis, overexpression vectorsas well as specific dsRNAi constructs designed tosilence each of the four genes have been construc-ted and trans formed into wild type plants. SeveralArabidopsis in sertion lines are se gregating for iso-lation of ho mozygous mutants. For each of thegenes promoter-GUS fusions have been construc-ted and transformed. To elucidate potential linksto other metabolic pathways we have includedseveral mutants in our analyses.

Another main interest in our research is to eluci -date molecular factors that govern the functionaladaptation of hydrolases to perform transacyla -tion reactions. As model protein we investigatethe sinapoylglucose:L-malate sinapoyltransferase(SMT) from Arabidopsis. A key requirement forthis research aimed at investigation of structure-function relation ships is the heterologous over-expression of SMT-cDNA to produce appropri -

ate amounts of func tional en zymefor crystallization experiments.Since SMT is subject to several post-translational mo di fications like gly-cosylation or formation of disul fidebrid ges functional expression re -quires an eu ka ryotic system. By op -timiza tion of in duction conditionsand construct se quences we wereable to produce en zy matically ac tiveSMT in Saccharomy ces cerevisiae.However, further work is ne cessaryto in crease the yield. As the homo-logous serine carboxypeptidases be -long to the best characterized pro-teins, a structure modelling ap proachhas been started. Model structuresob tained give first evidence for ami -no acid residues that are critical fortransacylation function and sub strate recogni -tion. Site-directed mu tagenesis will be performedto prove the functional relevance of these aminoacid positions. Sequence analyses reveal thatSCPL acyltransferases form a distinct group with -in the large family of SCPL proteins. They are spe-cific for higher plants obviously reflecting the di -versity of secondary metabolism.

Collaborators

Clint ChapplePurdue University, West Lafayette, USA

Martin Frauen, Gunhild Leck -bandNorddeutsche Pflanzenzucht, Hans GeorgLembke KG, Hohenlieth, Germany

Christian MöllersUniversity of Göttingen, Germany

José OrsiniSaaten-Union Resistenzlabor GmbH,Leopoldshöhe, Germany

Jürgen Schmidt, Sabine RosahlLeibniz Institute of Plant Biochemistry,Halle, Germany

Milton T. StubbsUniversity of Halle, Germany

Victor WrayGerman Research Centre forBiotechnology, Braunschweig, Germany

Similarity tree of SCPL proteins from Arabidopsis and functionally proven enzymes, the carbox-ypeptidases CPDY from yeast and CPDWII from wheat as well as the acyltransferases fromArabidopsis (SCT, SMT), rape (BnSCT) and wild tomato (GAC-Lp).

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PublicationsCacace, S., Schröder, G., Wehinger, E.,Strack, D., Schmidt, J. & Schröder, J. A fla-vonol O-methyltransferase from Catha -ran thus roseus performing two sequen tialme thylations. Phytochemistry 62, 127-137.

Doll, J., Hause, B., Demchenko, K., Paw -lowski, K. & Krajinski, F. A member of thegermin-like protein family is a high ly con-served mycorrrhiza-specific induced gene.Plant Cell Physiol. 44, 1208-1214.

Eckermann, C., Schröder, G., Eckermann,S., Strack, D., Schmidt, J., Schneider, B. &Schröder, J. Stilbenecarboxylate biosynthe-sis: A new function in the family of chal -cone synthase-related proteins. Phy toche -mis try 62, 271-286.

Hause, B., Hause, G., Kutter, C., Miersch, O.& Wasternack, C. Enzymes of jasmonatebiosynthesis occur in tomato sieve ele-ments. Plant Cell Physiol. 44, 643-648.

Hause, B., Stenzel, I., Miersch, O. & Was -ternack, C. Occurrence of the allene ox idecyclase in different organs and tissues ofArabidopsis thaliana. Phytoche mistry 64,971-980.

Ibdah, M., Zhang, X.-H., Schmidt, J. & Vogt, T.A novel Mg++-dependent O-me thyl trans fe -rase in the phenylpropanoid metabolismof Mesembryanthemum crystallinum. J.Biol. Chem. 278, 43961-43972.

Liu, S., Chen, K., Schliemann, W. & Strack, D.Isolation and identification of two flavoneglycosides in burdock (Arctium lap pa L.)leaves by polyamide column chromatogra-phy and high performance liquid chroma-tography in combination with e lec trosprayionization-mass spectrometry. Chinese J.Anal. Chem. 31, 1023.

Monostori, T., Schulze, J., Sharma, V. K.,Maucher, H., Wasternack, C. & Hause, B.Novel plasmid vectors for homologous

transformation on barley (Hordeum vulga-re L.) with the JIP23 cDNA in sense andantisense orientation. Cereal Res.Commun. 31, 17-24.

Münzenberger, B., Hammer, E., Wray, V.,Schauer, F., Schmidt, J. & Strack, D. De -toxification of ferulic acid by ectomycor-rhizal fungi. Mycorrhiza 13, 117-121.

Opitz, S., Schnitzler, J.-P., Hause, B. &Schneider, B. Histochemical analysis ofphenylphenalenone-related compounds inXiphidium caeruleum (Haemodo ra ceae).Planta 216, 881-889.

Peng, Z. F., Strack, D., Baumert, A., Sub -ramaniam, R., Goh, N. K., Chia, T. F., Tan, S.N. & Chia, L. S. Antioxidant flavonoids fromleaves of Polygonum hydropiper L. Phyto -che mistry 62, 219-228.

Proels, R. K., Hause, B., Berger, S. & Roitsch, T.Novel mode of hormone in duction of tan-dem tomato invertase genes in floral tissues.Plant Mol. Biol. 52, 191-201.

Stenzel, I., Hause, B., Maucher, H., Pitzsch ke,A., Miersch, O., Ziegler, J., Ry an, C. A. & Was -ternack, C. Allene oxide cyclase de pendenceof the wound re sponse and vascular bundle-specific ge neration of jasmonates in tomato- amp lification in wound signalling. Plant J. 33,577-589.

Stenzel, I., Hause, B., Miersch, O., Kurz, T.,Maucher, H., Weichert, H., Ziegler, J., Feuss -ner, I. & Wasternack, C. Jasmonate biosynthe-sis and the allene oxide cy clase family of Ara -bidopsis thaliana. Plant Mol. Biol. 51, 895-911.

Strack, D., Fester, T., Hause, B., Schlie mann,W. & Walter, M. H. Arbuscular my corrhiza:Biological, chemical, and molecular as -pects. J. Chem. Ecol. 29, 1955-1979.

Strack, D., Vogt, T. & Schliemann, W. Re centadvances in betalain research. Phy toche -mis try 62, 247-269.

Varet, A., Hause, B., Hause, G., Scheel, D. &Lee, J. The Arabidopsis NHL3 gene en -codes a plasma membrane protein and itsoverexpression correlates with in creasedresis tance to Pseudomonas syringae pv.tomato DC3000. Plant Physiol. 132, 2023-2033.

Wang, W., Hause, B., Peumans, W. J.,Smagghe, G., Mackie, A., Fraser, R. & VanDamme, E. J. M. The Tn-antigen specific lec-tin from Glechoma hederacea is an in sec -ticidal protein with an unusual physiology.Plant Physiol. 132, 1322-1334.

Books and Book ChaptersFester, T. Plastiden bei der arbuskulären My -korrhizasymbiose. In: Wurzelindu zier te Bo -denvorgänge, 14. Borkheider Se mi nar zurÖkophysiologie des Wurzelraums(Merbach, W., Egle, K. & Augustin, J., eds.)B.G. Teubner, Stuttgart, pp. 39-42.

Stenzel, I., Hause, B., Feussner, I. & Was ter -nack, C. Transcriptional activation of jas-monate biosynthesis enzymes is not re -flected at protein level. In: Advanced Re -search on Plant Lipids (Murata, N., Ya mada,M., Nishida, I., Okuyama, H., Sekija, J. & Ha -jime, W., eds.) Kluwer Academic Publi -shers, Dordrecht, pp. 267-270.

Strack, D. & Milkowski, C. Recruitment ofse rine carboxypeptidase-related proteinsin to phenylpropanoid metabolism. In: Poly -phenols 2002: Recent Advances in Poly -phenols Research. Vol. 1 (El Hadrami, I. &Daayf, F., eds.) Elwatanya Faculté des Scien -ces, Sem lalia, Marrakech (Morocco), pp. 50-61.

Stumpe, M., Stenzel, I., Weichert, H., Hau se,B. & Feussner, I. The lipoxygen ase pathway inmycorrhizal roots of Me dicago trun catula. In:Advanced Research on Plant Lipids (Murata,N., Yamada, M., Nishida, I., Oku ya ma, H.,Sekiya, J. & Hajime, W., eds.) KluwerAcademic Publi shers, Dordrecht, pp. 287-290.

Thorson, J. S. & Vogt, T. Glycosylated na -


Department of

Secondary Metabolism

PublicationsBücking, H., Förster, H., Stenzel, I., Miersch,O. & Hause, B. Applied jasmonates accumu-late extracellularly in to ma to, but in tra -cellularly in barley. FEBS Lett. 562, 45-50.

Camacho-Cristóbal, J. J., Lunar, L., Lafont, F.,Baumert, A. & González-Fontes, A. Bo rondeficiency causes accumulation of chloro-genic acid and caffeoyl polyamine conju -gates in tobacco leaves. J. Plant Physiol.161, 879-881.

Hans, J., Brandt, W. & Vogt, T. Site-directedmutagenesis and protein 3D-homologymodelling suggest a catalytic me ch anismfor UDP-glucose dependent be ta nidin 5-O-glucosyltransferase from Doro -theanthus bellidiformis. Plant J. 39, 319-333.

Hans, J., Hause, B., Strack, D. & Walter, M. H.Cloning, characterization, and im muno lo ca -lization of a mycorrhiza-indu cible 1-deoxy-D-xylulose 5-phosphate re duc toisomerasein arbuscule-contain ing cells of maize. PlantPhysiol. 134, 614-624.

Isayenkov, S., Fester, T. & Hause, B. Rapiddetermination of fungal colonization andarbuscule formation in roots of Me di cagotruncatula using real-time (RT) PCR. J.Plant Physiol. 161, 1379-1384.

Lukacin, R., Matern, U., Specker, S. & Vogt,T. Cations modulate the substrate specifi-city of bifunctional class I O-me thyl trans -fe rase from Ammi majus. FEBS Lett. 577,3 6 7 -370.

Maucher, H., Stenzel, I., Miersch, O., Stein,N., Prasa, M., Zierold, U., Schwei zer, P., Do -rer, C., Hause B. & Waster nack, C. The al -lene oxide cyclase of barley (Hordeumvulgare L.) -Cloning and organ-specific ex -pression. Phytochemis try 65, 801-811.

Miersch, O.,Weichert, H., Stenzel, I., Hau -se, B., Maucher, H., Feussner, I. & Was ter -nack, C. Constitutive overexpression of al -lene oxide cyclase in tomato(Lycopersicon esculentum cv. Lukullus)elevates levels of some jasmonates andoctadecanoids in flower organs but not inleaves. Phyto che mistry 65, 847-856.

Milkowski, C., Baumert, A., Schmidt, D., Neh -lin, L. & Strack, D. Molecular regula tion ofsinapate ester metabolism in Bras si ca napus:Expression of genes, properties of the enco-ded proteins and correlation of enzymeactivities with metabolite accumulation.Plant J. 38, 80-92.

Milkowski, C. & Strack, D. Serine carbo xy -peptidase-like acyltransferases. Phyto che -mis try 65, 517-524.

Schaarschmidt, S., Qu, N., Strack, D., Son -newald, U. & Hause, B. Local induc tion ofthe alc gene switch in transgenic tobaccoplants by acetaldehyde. Plant Cell Physiol.45, 1566-1577.

Vogt, T. Regiospecificity and kinetic proper-ties of a plant natural product O-me thyl -transferase are determined by its N-termi-nal domain. FEBS Lett. 561, 159-162.

Publications in PressGünther, C., Hause, B., Heinz, D., Krau ze -wicz, N., Rudolph, R. & Lilie, H. Com bi na -tion of listeriolysin O and a tumor-specificantibody facilitate efficient cell-type speci-fic gene delivery of conjugated DNA. Can -cer Gene Ther.

Hause, B. & Fester, T. Molecular and cell bio-logy of arbuscular mycorrhizal symbiosis.Planta.

Doctoral ThesesHans, Joachim: Isoprenoidbiosynthese inmy korrhizierten Maiswurzeln: Klonie rung,Charakterisierung und Lokalisie rung einer1-Desoxy-D-xylulose-5-phosphat-Reduk -toisomerase (DXR). Uni versity of Halle-Wittenberg, Depart ment of Biology,24/02/2004.

Hans, Judith: Klonierung und Charakte ri -sierung von Flavonoidglucosyl transfe ra senaus Beta vulgaris L. University of Halle-Wit tenberg, Department of Bio chemis try/Biotechnology, 23/06/2004.

HabilitationHause, Bettina: Biosynthese und Funk tio -nen von Jasmonaten - eine zellbiologischeAnalyse. Univer sity of Halle-Wit tenberg,Faculty of Mathematics, Natural Sciencesand Techno logy 02/12/2004.

63


Professor Dieter Strack organized a DFG supported symposium in Japan entitled "Germany-Japan Seminar on Mole cular Regulation of Plant Secondary Metabolism".The Japanese organizer was Kazuki Saito of the Chiba University. The symposium took place from September 20-23, 2004 at the Kazusa Akademia Center, Kisarazu,Chiba, Japan and was meant to serve as a starting point for more cooperation and exchange between Japanese and German scientists in the area of plant naturalproduct research. Ten German and 19 Japanese groups presented their work and discussed future plants for possible collaborations.

64

Administration and Technical ServicesHead: Lothar FranzenSecretary: Heide Pietsch until April 2004, afterwards Cindy Maksimo

Administration and

Technical Services

65

During the past two years, two construction projects have dominatedthe activities of the IPB administration. The construction of a new

sha red-use building started in January 2003 and was completed in March2004. In June 2004, construction of additional fully climate-controlled green-houses was initiated.

With a total area of 500 square meters, the shared-use building providesspace for 15 researchers and technicians from the Department of Bio -organic Chemistry. In addition to the two wide-span laboratories, thereare many rooms for special uses such as fermentation, distillation, liquidchromatography and robotics for combinatorial syntheses. With an inertgas extinguishing system, experiments can also be carried out in a special"night lab" without constant supervision. Laboratories for gas chromato-graphy, mass spectrometry, and DNA sequencing are used by scientistsfrom all departments. The total cost of about three million Euros was pro-vided by the Federal Ministry of Education and Research and by the Mi -nistry of Education and Cultural Affairs of the State of Saxony Anhalt.Nearly half the building outlay is required for the technical facilities forventilation, special gases, heating and a closed-cycle cooling system.

The new greenhouses, with an additional 350 square meters of floor space,will meet the growing demand of the departmental research programs in -volving transgenic plants. The nine air-conditioned chambers will be ope-ned in the near future for cultivation of tomato, poppy, tobacco, rape andplants from the mycorrhization projects. In addition, there will be two tro-pical chambers and incubators for Arabidopsis. The technical equipmentfor the fine control of climatic conditions is state-of-the-art and costsabout 1.1 million Euros. The total construction cost of 2.5 million Euroswas also provided by the Federal Ministry of Education and Research andby the Ministry of Education and Cultural Affairs of the State of SaxonyAnhalt. These new plant growth facilities will be ready for occupancy inMay 2005.

The building that currently houses employees of facilities and construc-tion management, graphic arts and horticulture will be replaced by twonew buildings in the near future. Within this new complex, an additional200 square meters of laboratory space for biological and chemical re -search for guest profes-sors is planned as well asnew offices for the In -formation Technology spe-cialists from the De part -ment of Stress and Deve -lopmental Biology.

66

Administration Employees

Head of the AdministrationLothar Franzen

AccountingHead: Barbara WolfGudrun SchildbergBurgunde Seidl, until September 2004Kerstin Wittenberg

ChauffeurJürgen Gaul

Computer SupportHead: Hans-Günter KönigHolger BartzKevin Begrow, trainee since August 2003Ronald Scheller

Construction Management Head: Heike BöhmCatrin Timpel

Facilities ManagementHead: Michael KrägeDetlef Dieckmeyer, until February 2004Carsten KothMichael KrägeJörg LemnitzerKlaus-Peter SchneiderEberhard Warkus

General AdministrationHead: Rosemarie StraßnerChristel Düfer, until June 2004Heide Pietsch, since April 2004Elviera SchotteTrainees Maike Hildebrandt, since August 2003Antje Olschewski, until July 2003

Oliver Prudyus, since August 2003Mandy Schatkowski, since August 2003Clemens Schinke, until July 2003

Graphic ArtsHead: Christine KaufmannAnnett Kohlberg

HorticultureHead: Iris RudischMartina AllstädtAnnett Grün, Trainee since August 2004Nicole Mühlwald, Trainee until September2003Christian MüllerPhilipp Plato, Trainee since August 2004Kristina RejallSteffen RudischKatja Scheming, TraineeAndrea Voigt, Trainee

Human ResourcesHead: Kerstin BalkenhohlAlexandra BurwigCindy Maksimo, until April 2004Antje Olschewski, until September 2004Rita Stelzer, until August 2004Kathleen Weckerle

Library ServicesHead: Andrea PiskolJessica Ackermann, Trainee until February2003Anja Gärtner, Trainee since August 2004Antje Werner, Trainee until July 2004

Press and Public RelationsHead: Sylvia PieplowJana Krupik, until September 2004

67

Employee Statistics

2003 2004

Average number of employees 172 172

Full-time employees in % 73 67

Part-time employees in % 27 33

Permanent positions 92 92

Temporary positions 26 23

Employees externally funded 41 31

Employees remunerated by “University Science FundsProgram” (Hochschulwissenschaftsprogramm/HWP) 5 6

Proportion of female employees in % 61 59

Personnel turnover in % 20 16

Averade age of employees in years 39 37

Scholarship / fellowship holders 25 26

Vocational traininggeneral administrationhorticulturelibrary sciencescomputer supportresearch technical assistant

3211-

34112

Succesfully completed vocational training 3 1

Average number of apprentices 7 11

68

Involvement of the IPB in National and International Scientific Networks

CERC 3Chairmen of the European Research Coun cilsChemistry CommitteesGerman Research Foundation

COMBIOCAT

Evolutionary Discovery of Novel Drugs byOrchestration of Polymer-Supported Com bi -natorial Bio-/Chemistry Fifth Framwork Programme of the EuropeanCom munitiy

EPILAOpioid Treatment of Chronic Pain and In flam -mation of the Locomotor SystemFifth Framwork Programme of the EuropeanCom munitiy

EVOMET

Evolution of Metabolic Diversity Priority Program 1152 of the German Re -search Foundation

GABI

Genome Analysisof the Plant Biological Sys temGerman Federal Ministry for Education, Re -search and Technology (BMBF) and BusinessCompanies

GABI-NONHOST

A consortium-based functional geno mics ini -tiative on plant non-host disease resistanceGABI 1b

METABOLOMICS PLATFORM

Metabolite Profiling in Arabidopsis and Crop PlantsGABI 2

SARAFunctional genomics of local and systematicacquired resistance in ArabidopsisGABI - trilateral Cooperation, France, Ger -

many, SpainCOMPARATIVE GENOMICS

Comparative genomics between Arabi dopsisand Brassica for genes directing seed-specificflavonoid biosynthesisGABI -Génoplante - bilateral Cooperation,Ger many, France

HEATOS

A Vietnamese opiate detoxification symp tommedicationGerman Federal Ministry for Education, Re -search and Technology (BMBF)

MOLECULAR ANALYSIS

OF PHYTOHORMONE ACTION

Priority Program SPP 1067 of the German Re -search Foundation (DFG)

MOLECULAR

CELL BIOLOGY OF PLANT SYSTEMS

Collaborative Research Center SFB 363 of theGerman Research Foundation

MOLMYK

Molecular Basics of Mycorrhizal Sym bio sesPriority Program SPP 1084 of the German Re -search Foundation

NAPUS 2000 Rapeseed breeding for improved human nutritionGerman Federal Ministry for Education, Re -search and Technology (BMBF)

SELF-ORGANIZATION THROUGH COORDINA TION

AND NON- COVALENT INTERACTIONS

Graduate Program of the German Re searchFoundation

SELENOPROTEINS

Priority Programm SPP 1087 of the GermanResearch Foundation

69

Resources and Investments

in Mio. Euro in %

Basic financing funds

Personnel 9.7 37.9

Consumables 4.4 17.2

Grants/Subsidies 0.2 0.8

Investments 7.4 28.8

“University Science Funds Program”(HWP) 0.3 1.2

Subtotal 22.0 85.9

Funds from external sources

BMBF 0.8 3.1

MK-LSA 0.1 0.4

DFG 1.5 5.9

Industry 0.5 2.0

EU 0.5 2.0

other 0.2 0.7

Subtotal 3.6 14.1

Total 25.6 100

Investments in Million Euro

Equipment 2.3

Construction 5.1

Total 7.4

Research grants listed on this and thefollowing pages were given by

AFGN Arabidopsis FunctionalGenomics Network (DFG)

BPS BASF Plant Science

BMBF Bundesministerium fürBildung und Forschung -Federal Minsitry forEducation and Research

DAAD Deutscher AkademischerAus tauschdienst - GermanAca demic Exchange Service

D-BFoundation

Gottlieb Daimler and KarlBenz Foundation

DBU Deutsche BundesstiftungUmwelt - German Environ -mental Foundation

DFG Deutsche Forschungsge mein -schaft - German ResearchFoun dation

Elsevier Elsevier Science Publisher

EU European Union

Firmenich Firmenich Company

Hopsteiner Hopsteiner Company

HWP Hochschulsonderprogramm -University Science FundsProgram

IconGenetics

Icon Genetics AG

MK-LSA Kultusministerium des LandesSachsen-Anhalt - Ministry ofEducation and Cultural Affairsof the State of Saxony Anhalt

MLU Martin Luther UniversitätHalle-Wittenberg - Universityof Halle

PPP Hungary ProjektbezogenerPersonenaustausch (DAAD)-Project Based Personnel Ex -change Program

Probiodrug Probiodrug AG

VWFoundation

Volkswagen Foundation

Wella Wella AG

Investments28.8 %

Personnel37.9 %

HWP1.2 %

Consumables & Grants18.0 %

Funds from external sources

14.1 %

70

Funds from External Sources

Title & Head of Project Duration Financial SourceAmount in Euro(2003-2004)

Personnel PostsFunded

Department of Natural Product Biotechnology

Jasmonate biosynthesis regulationProf. C. Wasternack & O. Miersch

02/0404/05

DFGDFG

49,30016,000

11

12-Hydroxyjasmonic acidProf. C. Wasternack & O. Miersch

03/05 DFG 30,200 1

Glutamat cyclaseProf. C. Wasternack

01/05 Probiodrug 24,400 0

AllenoxidcyclaseProf. C. Wasternack

01/03 Firmenich 6,500 0

Analysis of genesProf. T. Kutchan

00/04 Icon Genetics 57,200 1

Molecular genetics of isoquinoline alk. biosynth.Prof. T. Kutchan

01/0404/05

DFGDFG

93,70041,000

22

Molecular genetics in Liana Triphyoph. PellatumProf. T. Kutchan

03/05 DFG 82,800 1

Cellular signalingProf. T. Kutchan

02/04 DFG/MLU 46,400 1

Transformation and regeneration of P. somniferumS. Frick

02/04 DFG 115,300 2

Metabolic engeneeringS. Frick

04/06 DFG 15,000 2

Modulation of jasmonates by transgenic plantsProf. C. Wasternack & O. Miersch

02/04 DFG/SFB 363 126,100 1

Salvia fragrancesProf. T. Kutchan

02/03 DBU 31,200 0

HUM-NEUJ. Page

03/05 Hopsteiner 54,200 1

Subtotal: 789,300 16

Department of Bioorganic Chemistry

HEATOSProf. L. Wessjohann

00/03 BMBF 90,300 2

COMBIOCATProf. L. Wessjohann

01/04 EU 185,600 2

EPILAW. Brandt

01/04 EU 3,700 2

71



MCR ligand synthesisProf. L. Wessjohann

02/04 DAAD/Probral 11,900 0

Chrom-(II)-mediated reactionsProf. L. Wessjohann

02/03DAAD/

PPP Hungary5,200 0

Daimler Benz fellowshipProf. L. Wessjohann

02/03 D-B Foundation -1,100 1

Reactivity of selenopeptidsProf. L. Wessjohann & W. Brandt

03/0504/06

DFGDFG

100,40015,800

11

CERC-3Prof. L. Wessjohann

04/06 DFG 23,000 0

MUSKARINW. Brandt

03/04 MLU 31,900 1

HUMULUSProf. L. Wessjohann

03/04 Hopsteiner 20,000 0

Fungi metabolitesN. Arnold & J. Schmidt

04/06 DFG 21,000 1

ScreeningProf. L. Wessjohann

04/06 Wella 42,000 1


Department of Stress and Developmental Biology

Heavy metal toleranceD. Neumann & S. Clemens

02/04 DFG/SFB 363 70,600 1

CRISPProf. D. Scheel

01/04 EU 39,000 1

Heavy metal tolerance and siliconU. zur Nieden & S. Clemens

00/04 MK-LSA 28,500 1

Resistence in potatoesProf. D. Scheel & S. Rosahl

04/05 DFG 34,500 1

The role of jasmonates in pathogene defenseProf. D. Scheel

01/03 DFG 22,900 1

Jasmonate-insensitive mutantProf. D. Scheel

99/03 MK-LSA 13,400 1

Arabidopsis halleriS. Clemens

00/03 DFG 7,600 1

MetallophytesS. Clemens

01/04 EU 101,800 1

72

Funds from External Sources



MetalhomeS. Clemens

03/06 EU 64,700 1

BiomineralisationD. Neumann

01/04 DFG 23,700 1

Cooperation with South AfricaProf. T. Nürnberger

01/03 VW Foundation 28,100 0

NODOS. Rosahl

02/04 EU 147,800 1

Receptor kinasesProf. T. Nürnberger

02/04 DFG/AFGN 95,000 2

Bioinformatics and mass spectronomyProf. D. Scheel

02/03 BMBF 152,100 6

GABI-NONHOSTProf. D. Scheel

02/06BPSBMBF

304,700 4

Metal homeostasisS. Clemens

04/06 DFG 20,200 1

GABI-GENOPLANTES. Clemens

04/06 BMBF 10,000 1

SARAProf. D. Scheel

04/07 BMBF 11,200 1

Subtotal: 1,175,800 26

Department of Secondary Metabolism

Betanidin-GlucosyltransferasesT. Vogt & Prof. D. Strack

01/03 DFG 22,800 2

Metabolism of isoprenoidsM.H. Walter & T. FesterM.H. Walter

00/0303/0404/06

DFGDFGDFG

6,80049,70020,000

111

NAPUS 2000Prof. D. Strack

99/04 BMBF 186,200 2

The role of jasmonatesduring the establishment of mycorrhizaB. Hause & Prof. D. Strack

00/0404/06

DFGDFG

62,40016,700

11

Carotenoid biosynthesisin arbuscular mycorrhizal rootsT. Fester

00/0404/06

DFGDFG

42,80024,000

11

Metabolite profilingW. Schliemann

02/0404/06

DFGDFG

115,20023,000

11

PhytochemistryProf. D. Strack

02/04 Elsevier 53,200 1

73



HCA-GlucosyltransferasesC. Milkowski & A. Baumert

03/05 DFG 58,200 1

SCPL-AyltransferasesC. Milkowski & Prof. D. Strack

03/05 DFG 48,500 1

PFOMTT. Vogt

04/05 DFG 3,200 0


Joint projects

Metabolite profilingin Arabidopsis and crop plants, GABIDepartment of Stress and Developmental BiologyDepartment of Bioorganic ChemistryS. Clemens

00/04 BMBF 228,300 4

Metabolite profilingin Arabidopsis and crop plants, GABI-2Department of Stress and Developmental BiologyDepartment of Bioorganic ChemistryProf. D. Scheel, S. Clemens, Prof. L. Wessjohann, J. Schmidt

04/07 BMBF 87,800 3

Subtotal: 316,100 7

Total projects granted: 3,563,600 78

General overview

DFG 1,497,400 38

BMBF 765,900 19

EU 542,600 8

Industry 534,400 8

Other sources 181,400 3

MK-LSA 41,900 2

Total: 3,563,600 78

HWP 350,900 4

Total: 3,914,500 82

74

Seminars, Workshops and Colloquia 2003

January 14Luc Varin, Professor, Concordia University Montreal, CanadaFunctional analysis of sulfotransferases in Ara -bi dopsis thaliana.

January 23Ekkehard Neuhaus, ProfessorUniversity of Kaiserslautern, GermanyPhysiology and evolution of nucleoside trans-porters in plastids and bacteria.

Yves Marco, Dr.Laboratoire de Biologie Moleculaire des Re -lations Plantes-Mi cro-Organism CNRS, FranceResistance and disease development in Ara bi -dop sis thaliana in response to the bacterialpathogen Ralstonia solanacearum.

January 30-31First colloquium of Priority Program 1152,Evolution of Metabolic Diversity

February 4Athanassios Giannis, ProfessorUniversity of Leipzig, GermanySynthese und biologische Untersuchungenvon In hibitoren der Angiogenese.

February 6Zsuzsanna Schwarz-Sommer, Dr.Max Planck Institute for Plant Breeding Research, Cologne, Germany Homeotic control of floral organ identity insnapdragon: The problem of restricting geneexpression domains.

February 18Guy R. Cornelis, ProfessorBiocenter of the University of Basel, Switzerland How to switch off the innate immunity: TheYer sinia lesson.

February 20Mugio Nishizawa, ProfessorBunri University, Tokushima, Japan Synthetic approach into sterol biosynthesis byde veloping mercuric triflate.

February 25Klaus Naumann, Dr.Bayer AG, GermanyChlor - ein wichtiger Substituent in biologischaktiven Substanzen aus Natur und Synthese.

March 3Yang Ye, ProfessorInstitute of Materia Medica, Shanghai Chemical studies on traditional chinese medi -cine.

March 5Wolfram Weckwerth, Dr.Max Planck Institute of

Molecular Plant Physiology, Golm, GermanyMultidimensionale LC-MS für die Pflanzen -pro te omanalyse.

March 12Christine Schimek, Dr.University of Jena, GermanyTrisporoids, apocarotenoid signal moleculesin zy gomycetes.

March 25Mike Toorneman, Dr.University of Amsterdam, The Netherlands A (-)- Myrtenol based trigger system for thecontrolled formation of enediynes.

April 3Jörg Hacker, ProfessorUniversity of Würzburg, GermanyEscherichia coli as a model to study microbialpa thogenicity.

April 10Patrick Covello, Dr.NRC Plant Biotechnology InstituteSaskatoon, Kanada Genomics approaches to bioactive lignan bio-synthesis in Podophyllum.

April 15Ralph Panstruga, Dr.Max Planck Institute for Plant Breeding Research,Cologne, GermanyCorruption of host seven-transmembraneproteins by pathogenic microbes: A commontheme in animals and plants?

April 30Bernhard Westermann, Dr. University of Paderborn, GermanySynthesen zu Peptid- und Glycopeptidmimetika.

May 20Helmut Duddeck, ProfessorUniversity of Hannover, GermanyChirale Erkennung von Phosphorderivaten mitei nem Dirhodiumkomplex als NMR-Auxiliar.

May 21Gernot Frenking, ProfessorUniversity of Marburg, GermanyDie Natur der chemischen Bindung - alteFragen, neue Antworten.

May 26Bernadette Lippok, Dr.University of Freiburg, GermanyPythium and Arabidopsis - an interaction thatis complex and full of surprises.

June 4Robert A. Dietrich, Dr.Syngenta Biotechnology, North Carolina, USAThe Arabidopsis Bos mutants affect toleranceto both biotic and abiotic stresses.

Franziska Krajinski, Dr.University of Hannover, GermanyTranscriptional changes in the model plantMe di cago truncatula in response to symbioticand pa thogenic microorganisms.

June 5Burkhard Schulz, Dr.University of Tübingen, GermanyThe Arabidopsis immunophilin AtFKBP42(TWIS TED DWARF) integrates hormonalsignaling pathways in plants.

June 6Elisabeth Fuss, Dr.University of Düsseldorf, GermanyBiosynthesis of lignans in Linum spp.

June 12Bruno Lemaitre, Dr.Centre de Genetique Moleculaire, CNRS, Gif-sur-Yvette, FranceGenomic and genetic analysis of theDrosophila immune response.

Károly Micskei, Dr.University of Debrecen, HungaryAssymmetric synthesis induced by naturalamino acids in aqueous medium.

Tamás Patonay, ProfessorUniversity of Debrecen, HungaryAlpha-azido ketones: Useful tools in the syn-thesis of polyfunctionalized synthons andheterocycles.

June 19Regine Kahmann, ProfessorMax Planck Institute for Terrestrial Micro biology, Marburg, GermanyThe Ustilago maydis /maize pathosystem:Mating and beyond.

June 24Naohiro Kato, ProfessorUniversity of New Brunswick,USALive from plant cells: Analysis of plant nuclearfunctions using fluorescent proteins.

June 25Peter Bäuerle, ProfessorUniversity of Ulm, GermanySelbstorganisierende Oligo-, Poly- undCyclothio phene.

June 26Peter Bäuerle, ProfessorUniversity of Ulm, GermanyCombinatorial Organic Materials Research(COMR) A faster route to organic materials.

Ralf Oelmüller, ProfessorUniversity of Jena, GermanyRegulatory proteins and novel processes in

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photosynthesis.

June 30Volkmar Vill, ProfessorUniversity of Hamburg, GermanyNaturstoff-Datenbank Phytobase -aktuellerStand der Arbeiten.

July 2R. Madsen, ProfessorTechnical University of Denmark, LyngbySynthesis of natural products from carbohy-drates by the use of ring-closing olefin me ta -thesis.

July 3Axel Mithöfer, Dr.University of Munich, Germany Aspects of friend and foe discrimination inlegume-microbe interactions.

July 8Thomas Weimar, Dr.University of Lübeck, GermanyUntersuchung biomolekularer Wechsel -wirkungen mit Hilfe der NMR-Spek tro skopieund der Ober flächen-Plasmonen-Resonanz.

Ahlert Schmidt, ProfessorUniversity of HannoverThioredoxine steuern die assimilatorischeSulfat reduktion und die assimilatorischeNitrat re duk tion.

July 15Franz Nussbaum, Dr.Bayer AG, GermanyWarum-und vor allem wie-sollte man bioakti-ve Naturstoffe derivatisieren? Anregungen ausder Chemie des -(-)Galiellalactons.

July 16Werner Engewald, ProfessorUniversity of Leipzig, GermanyDie Kopplung GC-MS/MS-wievielChromatogra phie brauchen wir noch?

August 2003 Wolfgang Knogge, Dr. University of Adelaide, AustraliaNIPping grass leaves - a favorite fungal pasti-me and its consequences.

October 2Ulrich Zähringer, ProfessorLeibniz Center for Medicine and Biosciences, Borstel, Germany Peptidoglycan - Toll or Nod? Pathogenic pat-tern recognition in innate immunity.

October 6David Mackey, Dr.The Ohio State University, Columbus, USAThe interface between bacterial pathogens

and plants: Virulence functions and resistanceresponses.

October 15Harald Schmidt, ProfessorUniversity of Linz, AustriaNeues vom Holz.

October 16Hugo Kubinyi, ProfessorBASF AG Ludwigshafen, GermanyWirkstoffe:Vom Zufall zum gezielten Entwurf.

Murray Grant, Dr. Imperial College London, UKLocal and systemic signalling networks me -diated by the plant disease resistance proteinRPMI.

October 21Reinhard Agerer, ProfessorUniversity of Munich, Germany Ökologische und systematische Aspekte derEkto mykkorrhiza.

October 22Teun Nunnik, Dr.University of Amsterdam, The NetherlandsPhosphatidic acid - an emerging plant lipid se -cond messenger.

October 23-24Second colloquium of Priority Program 1152,Evolution of Metabolic Diversity

October 23Birgit Piechulla, ProfessorUniversity of Rostock, GermanyControl of time-specific scent emission inplants.

October 24Shin-Ichi Aizawa, Dr.Core Research for Evolutional Scienceand Technology, Tokyo, JapanFlagella as the type III secretion system.

October 28Günter Gerlach, Dr.Botanical Garden Munich, GermanyBlütenöle als Belohnung - Über die Biologiedes Ölblumensyndroms.

November 6Thomas Martin, Dr.University of Cambridge, UKAre Arabidopsis T-DNA mutants suitable forthe analysis of 14-3-3 specificity?

November 13David Stern, Dr.Boyce Thompson Institutefor Plant Research, New York, USA

A medley of ribonucleases: How plastidsadjust mRNA levels during plant develop-ment.

November 19Lutz H. GadeUniversity of Strasbourg, FranceStrategien in der Homogenkatalyse: Von derKo ordinationssphäre in Einkernkomplexen zuma kromolekularen Dendrimerkatalysatoren.

November 20Georg Frank, ProfessorBayer Bitterfeld GmbH, GermanyPhasen der Arzneimittelforschung am BeispielAs pirin.

November 27Klaus Apel, ProfessorSwiss Federal Instituteof Technology, Zurich, SwitzerlandSinglet oxygen-mediated stress responses ofAra bidopsis.

December 1Karin Groten, Dr.Rothamsted Research, Harpenden, UKMechanisms contributing to the senescenceof pea nodules.

December 4Andreas Freialdenhoven, Dr.Max Planck Institute for Plant Breeding Research, Cologne, GermanyControl of race-specific resistance and cell-death in barley.

December 10Thomas Schrader, ProfessorUniversity of MarburgSelf -organization and molecular recognitionof biomolecules by artificial receptors.

December 11Guido van den Ackerveken, Dr. University of Utecht, The Netherlands Susceptibility to downy mildew inArabidopsis: Mo lecular aspects of hosting anuninvited pathogenic guest.

WorkshopsFebrurary 2003Collaborative Research Center MolecularCell Biology of Plant Systems (SFB 363) of theGer man Research Foundation

June 2003,GABI Nonhost

MeetingsJanuary 2003

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Seminars, Workshops and Colloquia 2004

January 8Andreas Schaller, ProfessorUniversity of Hohenheim, GermanyJasmonate signaling in plant defense and pol-len development.

January 12Ian W. M. Smith, ProfessorUniversity of Birmingham, UKChemistry amongst the stars: Reaction kine-tics at a new frontier.

January 14Joachim Stöckigt, Professor University of Mainz, GermanyMolekulare Analyse der Vinorin Synthase - einzentrales Enzym der Alkaloidbiosynthese inRau volfia.

Manfred Psiorz, ProfessorBoehringer Ingelheim Pharma GmbH & Co KG, GermanyTropan-Stukturen in Natur- und Wirkstoffen.

January 15Iwona Adamska, ProfessorUniversity of Konstanz, GermanyProtective mechanisms against light stress inthechloroplast of higher plants.

February 12Georg Coupland, ProfessorMax Planck Institute for Plant Breeding Research, Cologne, GermanyThe regulation of plant development by sea-sonal cues.

February 18A. Llobet, ProfessorUniversity of Girona, SpainAzomacrocyclic complexes and their applica-tion in bioorganic and coordination chemi-stry.

February 19Steffen Backert, Dr.University of Magdeburg, GermanyFunction of two type IV secretion systems inHe liobacter pylori: Protein translocation andconjugative chromosomal DNA transfer.

February 26Helle Ulrich, Dr.Max Planck Institute for Terrestrial Micro biology, Marburg, GermanyControl of genome stability by ubiquitin andSUMO.

March 18Imre Somssich, Dr.Max Planck Institute for Plant Breeding Research, Cologne, GermanySearch for in vivo target genes of WRKY

trans cription factors involved in plant defenseand leaf senescence.March 25Bonnie Carolyn McCaig, Dr.Michigan State University, USAThe role of jasmonate perception in tomatore production and development.

March 31Ute Wittstock, Dr.Max Planck Institute for Chemical Ecology, Jena, Germany Special weapons - exceptional countermeasu-res: How insects cope with an activated plantdefense system.

April 7Jürgen O. Metzger, ProfessorUniversity of Oldenburg , GermanyMassenspektrometrische Untersuchung vonRe aktionen in Lösung oder wie kann manCarbe niumionen bei SN1-Reaktionen undRadikale bei Radikalkettenreaktionen sehen.

Ruth Niemetz, Dr. University of Ulm, GermanyZur Biosynthese komplexer Gallotannine inRhus typhina und Ellagitannine in Tellima gran-diflora.

April 22Martin Parniske, Dr.John Innes Centre, Norwich, UKPlant genetics of root symbiosis with fungiand bacteria.

April 29Widmar Tanner, ProfessorUniversity of RegensburgPlant membrane transport: From physiologyto molecular biology and back.

May 3Laurent Zimmerli, Dr.University of Fribourg, Switzerland Early events in Arabidopsis nonhost resi-stance.

May 6Hanjo Hellmann, ProfessorFreie Universität Berlin, GermanyCullins as regulators in the ubiquitin protea-som pathway.

May 10György HorvathUniversity of Antwerp, The NetherlandsBiosynthesis of tocotrienols.

May 11Koop Lammertsma, ProfessorUniversity of Amsterdam, The NetherlandsGenerating and applying new organophorusrea gents.

Randolph J. Alonso-Herrera, Dr.Venezuelan Institute for Scientific Research Caracas, VenzuelaSynthetic derivatives of natural products withpo tential chemotherapeutic applicability.

May 12Burghard König, ProfessorUniversity of Regensburg, GermanyMolecular recognition with coordinationcompounds.

May 18Jens Rohloff, Dr.Norwegian University of Science and Technology, Trondheim, NorwayApproaches in Arabidopsis research: Spacebiology and differences of smell.

May 26Jakob Ley, Dr. Symrise GmbH & Co KG, Holzminden, GermanyGeschmack - Physiologie, Moleküle,Modifizierung.

June 3Herman Spaink, ProfessorUniversity of Leiden, The NetherlandsSimilarities of microbial recognition by plantsand animals.

June 9Birgit Kersten, Dr. Max Planck Institute for Molecular Genetics, Berlin, GermanyTowards plant proteomic studies using pro-tein mi croarrays.

June 10Markus Pauly, Dr. Max Planck Institute of Molecular Plant Physiology, Golm, Germany Life on the outside: Why do plant cell wallshave to be so complex?

June 18Cornelia Mrosk, Dr.University of Jena, GermanyDie ß-Amylase in Turionen von Spirodelapolyrhiza: Regulation durch Licht und Nitrat.

June 22Harro J. Bouwmeester, Dr.Plant Research International, Wageningen, The NetherlandsRole of terpenoids in signaling between plantsand other organisms.

June 24Olivier Voinnet, Dr.Institut de Biologie Moleculaire des PlantesCNRS, Strasbourg, France

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Mechanisms and roles of RNA silencing inplants and animals.

June 29Pierre Potier, ProfessorInstitut de Chimie des Substances Natu -relles, CNRS, Giv sur Yvette, FranceResearch and discoveries of new antitumordrugs: NAVELBINE ® & TAXOTERE ®.

July 5Vincenzo de Luca, ProfessorUniversity of Ontario, KanadaNew tools for understanding metabolic path-ways in single cells: The case for Catharanthusroseus indole alkaloid biosynthesis.

July 8Steffen Abel, ProfessorUniversity of California, Davis, USAGlucosinolates: From biosynthesis to pathwayre gulation.

July 15Jeff Dangl, ProfessorUniversity of North Carolina, Chapel Hill, USAP. syringae type III effector proteins manipula-te host cell biology, and plant disease resi-stance gene products stop them.

July 23Gynheung An, ProfessorPohang University of Science and Technology, South KoreaT-DNA insertional mutagenesis for reproduc-tive development in rice.

September 2Stefanie RanfUniversity of Souht Carolina, Columbia, USAVirus induced gene silencing of MAP kinasesin to mato.

Roger Wise, ProfessorIowa State University, USA

Interplay of gene-specific resistance to barley-powdery mildew and the suppression of host-re sponses.

September 8Virinder Parma, ProfessorUniversity of Massachusetts, Lowell, USABiocatalytic generation of novel materials ofim portance in drug and gene delivery.

October 4-5Third colloquium of Priority Program 1152,Evolution of Metabolic Diversity

October 10Ko Shimamoto, Dr.Nara Institute of Science and Technology, JapanRac GTPase is a key regulator of defensesignaling in rice.

October 18Bertil Helgee, ProfessorUniversity of Technology of Göteborg, SwedenN-Vinylpyrrolidone co-polymers, possiblesupport materials for chemical reactions.

October 21Joachim Uhrig, Dr.Max Planck Institutefor Plant Breeding Research, Köln, GermanyProtein interaction networks: Functional ana-lysis of plant protein families and plant-virusinterac tions.

October 27Gerd Jürgens, ProfessorUniversity of TübingenApical-basal pattern formation in Arabidopsisem bryogenesis.

November 4Raoul Bino, ProfessorUniversity of Wageningen, The NetherlandsPlant Metabolomics for Plant Breeding.

November 11

G. Ungar, ProfessorUniversity of Sheffield, UKPeriodic and quasiperiodic patterns in dendri-mer nanostructures.November 16Christa Kamperdick, Dr.Institute of Molecular Biotechnology Jena, GermanyDetection of protein-ligand interactiuons bysaturation transfer difference NMRSpectroscopy (STD).

November 18Beat Keller, ProfessorUniversity of Zurich, SwitzerlandDiversity and evolution of resistance genes incultivated and wild wheat: Exploring theresources of a crop plant.

November 24Klaus Müllen, ProfessorMax Planck Institute for Polymer ResearchGraphitmoleküle.

November 26Gopalan Selvaraj, Dr.National Research Council of CanadaMolecular aspects of reproductive develop-ment in cereals and oilseeds.

December 8Albrecht Berkessel, ProfessorUniversity of Cologne, Germany Biomimetik und Organokatalyse für dieSynthese enantiomerenreiner Epoxide, Aldoleund Amino säuren.

December 9Anne Osbourn, Dr.John Innes Centre Norwich, UKSecondary metabolism and plant defense.

Workshop

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Department of Bioorganic Chemistry

Department of Stress and Developmental Biology

Susanne Aust, Dr., Germany01.03. 2003 - 31.12. 2004

Lilechi Danstone Baraza, TanzaniaNAPRECA-DAAD Fellow01.06. 2004 - 30.11. 2004

Alessandra Basso, Dr., ItalyEU Fellow12.01. 2004 - 29.01. 2004

Claudia Bobach, Germany01.10. 2004 - 30.11. 2004

Christiano Rodrigo Bohn Rhoden,BrazilDAAD Fellow01.10. 2004 - 31.12. 2004

Antonio Luiz Braga, Prof., BrazilDAAD Fellow16. 09. 2004 - 31.10. 2004

Tran Van Chien, Vietnam07.10. 2002 - 30.09. 2003

Nguyen Manh Cuong, VietnamDAAD Fellow01.06. 2004 - 31.08. 2004

Katalin Czifrak, HungaryDAAD Fellow04.08. 2003 - 30.09. 2003

Victor

Dick, GermanyFellow Stifterverband für die DeutscheWissenschaft01.02. 2004 - 31.12. 2004

Simon Dörner, GermanyFellow Studienstiftung des DeutschenVolkes01.03. 2004 - 31.12. 2004

Kanchana Dumri, ThailandDAAD-Leibniz Fellow01.03. 2004 - 31.12. 2004

Othilie Vercillo Eichler, BrazilDAAD Fellow01.08. 2004 - 31.12. 2004

Gergely Gulyás, Hungary01.08. 2004 - 30.09. 2004

Zsuzsa Juhasz, HungaryDAAD Fellow01.10. 2003 - 31.10. 2003

Christine Kamperdick, Dr., GermanyHumboldt Fellow22.04. 2003 - 21.10. 2003

Myint Myint Khine, MyanmarDaimler Benz Fellow01.01. 2003 - 31.12. 2004

Christine Neuhaus, Germany01.03. 2004 - 30.09. 2004

Luay Rashan, Prof., JordanHumboldt Fellow12.07. 2004 - 30.07. 2004

Daniel Garcia Rivera, CubaGraduiertenkolleg01.10. 2003 - 31.12. 2003Juliana Schneider, Brazil

DAAD Fellow16.04. 2003 - 15.10. 2003

Paulo Henrique Schneider, BrazilDAAD Fellow16.04. 2003 - 15.10. 2003

Jasqer Alonso Sehnem, Brazil03.10. 2004 - 31.12. 2004

Jana Selent, Germany12.01. 2004 - 30.04. 2004

Katalin Sepreny, HungaryDAAD Fellow01.11. 2003 - 30.11. 2003

Tran Van Sung, Prof., VietnamFebruary, June and September 2004

Mieke Toorneman, The Netherlands01.01. 2003 - 30.10. 2003

Larissa Vasilets, Dr., Russia01.01. 2003 - 31.05.2004

Nguyen Hong Thi Van, Vietnam01.01. 2003 - 16.04. 2003

Heike Wilhelm, Dr., GermanyFellow BioService GmbH, EU and theState of Saxony Anhalt01.01. 2004 - 31.12. 2004

Hasliza Yusof, Malaysia06.10. - 20.10.2004

Reetta Ahlfors, FinlandDAAD Fellow08.07. 2002 - 30.04. 2003

Emiko Harada, Dr., JapanHumboldt Fellowsince 22.02. 2002

Ingo Hofmann, Germany01.01. 2003 - 31.12. 2003

Thorsten Nürnberger, Prof.,Germany01.08. 2003 - 31.12. 2003

Claudia Simm, GermanyGraduiertenkollegsince 01.10. 2003

Aleksandra Trampczynska, Dr.,Poland10.11. 2003 - 30.11. 2003

Guest Researchers and Fellows

Myint Myint Khine, Tran Thi Phuong Thaoand Kanchana Dumri (from left to right).

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Department of Natural Product Biotechnology

Andrea Andrade, ArgentinaDAAD Fellow05.08. 2003 - 30.09.2003

Andrea Borgogni, Italy09.04. 2004 - 16.06. 2004

Kum-Boo Choi, Dr., KoreaHumboldt Fellow07.10. 2002 - 30.09. 2003

Bonnie Carolyn McCaig, Dr., USA06.01. 2004 - 02.04.2004Maria Luisa Diaz Chavez, Mexiko

DAAD Fellow01.10. 2003 - 30.09. 2004

Aphacha Jindaprasert, ThailandDAAD Fellowsince 18.10. 2004

Kristin Krukenberg, USAFulbright Fellow23.09. 2002 - 15.07. 2003

Natsajee Nualkaew, ThailandDAAD Fellow01.11. 2004 - 31.12.2004

Alfonso Lara Quesada, Costa RicaDAAD Fellowsince 01.04. 2003

Luc Varin, Prof., Canada01.10. 2002 - 31.01. 2003

Department of Secondary Metabolism

Ma Hnin Hnin Aung, Singapore13.01. 2003 - 28.02. 2003

Ana Cenzano, Dr., Argentina01.09. - 30.11. 2004

Diana Schmidt, GermanyFellow, Bio Service GmbH, EU and the State of Saxony Anhalt01.08. 2001 - 31.07. 2004

Rostand Tonleu Tonfack

Administration and

Technical Services

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Press and Public RelationsHead: Sylvia PieplowAssistant: Jana Krupik

Press and

Public Relations

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National Year of Chemistry 2003The public relations activities in 2003 were effec-ted by the national Year of Chemistry. This the-matic year, initiated by the Donors Associationfor the Promotion of Sciences in Germany, hasbeen enriched by our chemists with many pro-jects and ideas.

Special LecturesIn the context of special lectures for the Year ofChemistry, Professor Ludger Wessjohann spokein March 2003 about flavors and pharmaceuticallyimportant ingredients from plants. The lecture,named Pizza Taste and Jungle Drugs - Che micalSubstances in Nature, was addressed to primaryschool students, high school students and inter-ested laymen.

Very interesting as well was the lecture about thebiology of oil flowers in October, given by the cu -rator of the botanical garden in Munich, GünterGerlach. The talk was organized by the IPB re -searchers who are investigating the biologicalrole of the poorly understood flower oils in co -operation with the botanical garden.

The Ship of Chemistry 2003In summer 2003 the, so-called Ship of Che mistrystarted as a floating exhibition its tour on theRhine. The exhibits about chemistry in every daylife could be visited by a large number of peopleduring the stops of the ship in every major har-bor. The IPB participated in this exhibition with amultimedia project about the scientific work onthe institute. The presentation contained a virtualtour through the IPB and a game, designed toinform people about bioorganic chemist’s work.The player has to isolate and analyze active sub-stances from medical plants. At the end of thegame, the player knows the structure of the com-pounds and is crowned with a doctoral hat. Thegame was planned mainly by our scientists LarsBräuer, Wolfgang Brandt, Andrea Porzel and Pro -fessor Ludger Wessjohann from the De partmentof Bio organic Chemistry.

The game and the virtual tour were burned onCD and distributed in August 2004 to many se -condary schools, universities, firms, publishing com-

panies and private ci tizens inGermany, Aus tria and Switzer -land. The response to the gamewas consistently positive and re -sulted both in numerous pressarticles and in an increase in ge -neral name recognition of the IPBin Ger many.

Public Events 2003 and 2004Germany’s Day of Re uni fica tionOn October 3, 2003, the IPB participated toge -ther with other scientific institutes of the state ofSa xony Anhalt in the celebration of Germany'sDay of Reunification in Mag de burg. Together withthe university of Halle, the IPB exhibited the dif-ferences between human and plant cell cultures.

Long Nightof Science 2003 and 2004The second and the third LongNight of Science were both greatsuccesses for our institute. On thisfirst Friday in July, the institute'sdoors were open from 7 pm tomidnight to welcome more then300 members of the public. Vi si -tors enthusiasti cally participatedin guided tours through the labsand greenhouses. In addition, ourscientists displayed experiments and ex hibits inthe foyer. Special lectures were fa vo red by the pu -blic. Stephan Cle mens spoke about the chancesfor and risks of green gene technology, whereasProfessor Ludger Wess jo hann gave an overviewabout plant natural products and their applica tionas pharmaceuticals.

Guided Tours for ScholarsAs in the previous years, the IPB organized manyguided tours through the institute for schoolclasses. Teachers and foreign students were alsovery interested in this possibility to learn moreabout the institute. In addition, all of the fourscientific departments sponsored several periodsof practical training, which allowed many highschool students to gain insight into lab work andto try out experiments on the bench.

The Phytolator crowned with a doctoral hat.

Carsten Milkowski showed electrophoreticexperiments in the Long Night of Science2004.

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Participation in Trade ShowsBiotechnika in 2003In October 2003, the IPB partici-pated in Biotechnika, Germany'smost important international bio-technology exhibition. Togetherwith universities and many scien-tific institutes and firms, our re -searchers presented their work atthe Saxony Anhalt booth in Han -nover. In this trade show, Mi chaelH. Walter from the Depart mentof Secondary Metabolism display-ed topics on mycorrhiza - a sym-biosis between plant roots andfungi.

ABIC in 2004 In a similar Saxony Anhalt booth the, IPB partici-pated in September 2004 in ABIC in Cologne.For this Agricultural Biotechnology InternationalConference, resear chers of the Department ofStress and Developmental Biology presented me -chanisms of metal hyperaccumulation in plants.

Projects in 2004In 2004, our institute was highly present in theregional and national press. Some of our scientificwork also resulted in interest by public broadca-sters. In addition, several high-ranking dignitariesvisited the IPB to get an impression about ourscientific projects and the conditions of work.With the organization of two art exhibitions, wecould revitalize an old custom of the past, whichgave many artists the possibility to show theirown perspectives about science and other im -portant aspects of life.

Visits of DignitariesMembers of the German Lower House ofParliament visit the IPBIn April 2004, Dr. Christoph Bergner, member ofthe German Lower House of Parliament, visitedthe IPB not only for nostalgic reasons, but also todiscuss further promotion of the Leibniz Societyinstitutes. One central point in discussions withthe former scientist of this institute was the re -solution of the new gene technology law in thenear future.

Also in April, Christel Rieman-Hahnewinckel, Ul -rich Kasparick, and Dr. Wolfgang Eichler visitedthe IPB to gain information on the major re -search projects on the institute. The members ofthe German Lower House of Parliament wereimpressed by the excellent research conditions atthe IPB. With this visit, the politicians were re -sponding to an initiative of the government toprovide more support to the best scientific insti-tutes in the eastern part of Germany. Togetherwith the directors, they discussed the necessarysteps of economic fortification by a better pre-sentation of the whole scientific landscape in thisregion. "In spite of the fact that many of East Ger -many's research institutes work on an excellentprofessional level, they have image problems inthe western part of the country", Ulrich Kas pa -rick said. These problems could be resolved byvisiting the corresponding institutes and by morepersonal contact between scientists and politi-cians.

President of the Leibniz Societytours our Labs and GreenhousesWith pleasure, members of the IPB welcomedthe president of the Leibniz Society Hans OlafHenkel in June 2004. Henkel, who is visiting at re -gular intervals all of the 84 Leibniz Institutes, wasimpressed by the scientific projects of the insti -tute. After the tour, he discussed with the direc-tors some general questions about the futurerole of the Leibniz Society within national and in -ternational science politics.

European Commission Of ficervisits important scientific insti-tutes of Saxony AnhaltIn context of the recently star -ted biotechnology offensive bythe State of Saxony Anhalt, inOc to ber 2004, Dr. ChristianPater mann from the Eu ropeanDi rectorate-General for Re -search viewed the IPB andother re gional scientific esta-blishments with similar orien-tation in plant biochemistry. Inaddition to green biotechnolo-gy, the im ple mentation strate-

In Biotechnika 2003 in Hannover, the IPB, re -presented by Dr. Michael H. Walter (left), exhibi-ted topics on mycorrhiza. Here Dr. Walter is dis-cussing with Dr. Horst Rehberger (right), Saxony-Anhalt's Minister for Work and Eco no my.

Hans Olaf Henkel (right), Presi -dent of the Leibniz So ciety, visitedthe IPB in June 2004. Here, hespeaks with Prof. Dierk Scheel, Ma -na ging Director of the institute.

gy of this of fensive aims to support the pharma-ceutical industry, neu rotechnology and red bio-technology as well. Af ter the circuit, Dr.Patermann informed interested persons aboutcurrent and planned scientific projects of the EU.

New information about Mycorrhiza Our multimedia presentation about mycorrhizawas in this year completely updated by its crea-tor Thomas Fester. The scientist of the De part -ment of Secondary Metabolism completed thepresentation with the newest scientific re sultsand with references from the literature. The firstversion of the interactive course about this fasci-nating biological interaction was produced in2001 for students and interested nonscientistsalike. In September 2004, the renewed programwas again burned on CD and sent to manyschools, firms, publishing companies and privatecitizens throughout the German-speaking regionin Europe.

Art Exhibitions 2004Monotypes from Hanno LehmanThe exhibition Farbansichten in July and August2004 showed pictures of the painter Hanno Leh -man. Hanno Lehmann studied chemistry and wor-

ked for many years as scientific co-wor-ker at the IPB. His paintings displayed awide range of themes from landscapesand scientific motifs to dreams andhuman relationships. All pictures weremade as monotype, a spe cial printingtechnology, which was modified andimproved by the painter.

Oil on Canvas from Giacomo PiccoliThe Italian painter and sculptor Gia co -mo Piccoli abducted us into the border-land between science and philosophy.His exhibition named Perpetuum Mobile- Magic of Biology in Imaginary Worldswas shown from October 2004 untilJanuary 2005 at the IPB. Inspired byplants, fruits and microscopic structu-res, Giacomo Piccoli displayed amazingspaces of surrealistic perspectives.Giacomo Piccoli, born in Catania(Sicilia) in 1949, he was elected in 1983National President of the ANTITESI cul-tural circuits in Rom. Currently he ischancellor of the Italian BibliographicalCenter.

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Our CD about mycorrhiza in a new layout.

TemptationMonotype from Hanno Lehmann.

Hair CellOil in canvas, Giacomo Piccoli

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Articles and Press ReleasesJANUARY 2003Stäudner, F. Pflanzen als Schadstoff schluk -ker. Leibniz, p. 5.

Pieplow,S. Pflanzliche "Staubsauger" ziehenSchwermetalle aus dem Boden. Wirt -schaftsspiegel, p. 40.

This article was also published by:- www.analytik-news.de- www.chemie.de- ww.e2nn.de- www.innovations-report.de- www.uni-protokolle.de- www.unitools.de- www.vln-saar.de- www.zalf.de

08.01.2003Jäckel, E. Land strebt in die Weltklasse -Netzwerk Innoplanta sorgt für Aufsehen.Mitteldeutsche Zeitung, p. 17.

28.01.2003Crodel, C. Junge Gärtner lösen manchknifflige Frage. Mitteldeutsche Zeitung.

15.02.2003Pieplow, S. Staubsauger und Schnell rei ni -ger. Laborjournal, p. 34-35.

21.03.2003PIEPLOW, S., PIZZAGESCHMACK UND UR -WALDDROGE, PRESS RELEASE.

24.03.2003PIEPLOW, S., SUBTILE STRATEGIEN DERPFLANZEN GEGEN KRANKHEITSERREGER, PRESSRELEASE.

Based on this press release articles were published by

- www.innovations-report.de- www.uni-protokolle.de- www.zalf.de

MARCH 2003Niemann,T. Nachgefragt Stephan Cle mens.Bild der Wissenschaft.

16.03.2003Krause, I. Biologe mit Schwäche für diehöchsten Gipfel der Welt. MitteldeutscheZeitung.

13.05.2003Pieplow, S. Wenn das Grünzeug einenSchnupfen hat. Mitteldeutsche Zeitung, p.17.

13.05.2003Krause, I. Entscheidung für Halle als positi-ves Signal. Mitteldeutsche Zeitung, p. 17.

01.08.2003PIEPLOW, S., EINLADUNG ZUM RICHT FEST, PRESSRELEASE

08.08.2003Krause, I. Richtfest auf dem Campus -Institut erhält neues Labor. MitteldeutscheZeitung.

11.08.2003Ronzheimer, M., Die WGL im DFG-Ran -

king, www.berlinews.de

25.09.2003Daur, V. Biotechnologie im Land gewinntProfil. Mitteldeutsche Zeitung.

25.09.2003Daur, V. Bund setzt Tradition in Qued lin -burg fort. Mitteldeutsche Zeitung.

08.10.2003WALTER, M. H., NATÜRLICHE HELFER FÜR DIEPFLAN ZENERNÄHRUNG: BIO DÜN GUNG DURCHSYMBIONTISCHE MY KOR RHIZA PIL ZE, PRESSRELEASE.

23.10. 2003PIEPLOW, S., BLÜTENÖLE UND DAS ÖL -BLUMENSYNDROM, PRESS PELEASE.

04.12. 2003MAX-PLANCK-GESELLSCHAFT,NEUE HOFFNUNG FÜR SCHWERMETALL-BELA-

STETE BÖDEN, PRESS PELEASE.

Book ChapterFebruary 2003Pieplow, S. Pflanzenforschung im interna-tionalen Rampenlicht. In Bauen für dieWissenschaft Sachsen-Anhalt,GEHRIGVerlagsgesellschaft mbH, p. 80-81.

Radio TransmissionJanuary 2003Schilling, J. Pflanzenstaubsauger. NDR4.

17.03.2003Smiljanic, M. Grüne Staubsauger - wiePflanzen vergiftete Böden sanieren.Leonardo - Wissenschaft und mehr,WDR5.

Publications and Press Releases 2003

85


Articles and Press ReleasesJanuary 2004Winter, S., Schlachten in der Furche, DerSPIEGEL 4/2004, p. 80.

28.01.2004Witthuhn, B., Die grünen Sanierungshelfer,Berliner Zeitung, p. 13.

19.03.04PIEPLOW, S., LEIBNIZ-INSTITUT ERÖFFNETWEITE-RES FORSCHUNGSGEBÄUDE AUF DEMWEINBERGCAMPUS, PRESS RE LEASE.

26.03.04Neubau für Biochemiker, MitteldeutscheZeitung, p. 9.

27.03.04Krause, I., Viel Licht und ein Nachtlabor,Mitteldeutsche Zeitung, p. 12.

27.03.04Richter, S., Erst Kellerkind - jetzt im Licht,Wochenspiegel.

29.03.04Neues Forschungsgebäude seiner Be stim -mung übergeben,www.weinbergcampus.de.

24.05.04WINGERT, N., HALLES BIOLOGEN ENTTARNEN DIE

GE HEIM NISSE VON PFLANZEN UND PILZEN, PRESSRE LEA SE.

07.06.2004PIEPLOW, S., HANS OLAF HENKEL BE SUCHTDAS LEIBNIZ-INSTITUT FÜRPFLANZENBIOCHEMIE, PRESS RELEASE.

09.06. 2004Krause, I., Henkel besucht Institut fürPflanzenbiochemie, MitteldeutscheZeitung, p.12.

09.06.04PIEPLOW, S., TRANSGENER RAPS PRODUZIERTSAMEN OHNE BITTERSTOFFE,PRESS RELEASE.

11.06.04Transgener Raps produziert Samen ohneBitterstoffe, www.innovations-report.de.

18.06.04Präsident der Leibniz-Gemeinschaft am 8.Juni zu Besuch in Halle, www.weinbwerg-campus.de.

20.06.2004Pieplow, S., Bitte weniger bitter, Labor -journal 06/2004, p. 34-35.

21.06.04PIEPLOW, S., AUSSTELLUNG FARBAN SICH TENVON HANNO LEH MANN AM LEIBNIZ-INSTITUTFÜR PFLAN ZEN BIO CHEMIE,PRESS RELEASE.

23.06.04Lehmann zeigt Farbansichten, Mitteldeut -sche Zeitung.

25.06.04Krause, I., Einblicke auf dem Campus, Mit -teldeutsche Zeitung, p.16.

28.06.04PIEPLOW, S., LANGE NACHT DER WIS -SENSCHAFTEN AM LEIBNIZ-INSTITUT FÜR

PFLANZENBIOCHEMIE, PRESS RELEASE.

29.06.04Grafiker zeigt Ansichten von der Farbe,Mitteldeutsche Zeitung, p.17.

01.07.04Gabriele Herrmann vom Institut für Pflan -zenbiochemie setzt frische Zellkul tu ren fürdie Lange Nacht der Wissen schaf ten an,Mitteldeutsche Zeitung, Photography, p. 15.

09.07.04Krause, I., Forscher entdecken Ursachenfür den bitteren Geschmack,Mitteldeutsche Zeitung, p.19.

31.07.04Lehmanns Farbansichten im Institut, Mit -tel deutsche Zei tung, Photography, p.12.

31.07.04Schmundt, H., Dschungel unter den Füßen,DER SPIEGEL 31/2004, p. 116-118.

24.08.04PIEPLOW, S., MIT DEM PHYTOLATOR IN DENDSCHUNGEL DER ERKENN TNIS, PRESS RELEASE.

Based on this press release articleswere published by:

- www.analytik.de- www.bildung-branden burg.de- www.bildungsklick.de- www.chemie.de- www.chemikalien.de- www.chemlin.de- www.lehrer-online.de- www.studieren-im-netz.de- www.uniprotokolle.de- www.wgl.de- www.wifoe.halle.de

01.09.04Rapssamen ohne Bitterstoffe, Leibniz 3, p.3.

03.09.04Gauselmann, K., Verunsicherung in "Ex zel -lenz-Zentren", Mitteldeutsche Zeitung, p.6.

04.09.04Gauselmann, K., Zusage an Institute,Mittel deutsche Zeitung, p.1.

10.9.04Chemiespiel kostenlos, MitteldeutscheZei tung, p. 13.

29.09.04Schwägerl, C., Eine Pflanze ändert ihr Ima -ge - der Raps als Edelgewächs, Frank furterAllgemeine Zeitung 227.

30.9.04 PIEPLOW, S., MYKORRHIZA IM NEUEN GE WAND,PRESS RELEASE.

This press release was also published by:- www.innovationsreport.de- www.interconnections.de- www.mygeo.de- www.vdbiol.de- www.wgl.de- www.wissensschule.de- www.zeus.zeit.de

30.09.04PIEPLOW, S., AUSSTELLUNG PERPETUUM MOBILEVON GIACOMO PICCOLI AM LEIBNIZ-INSTITUTFÜR PFLANZEN BIO CHEMIE, PRESS RELEASE

This press release was also published by:- www.interconnections.de- www.prowissenschaft.de- www.qinx.de- www.uniprotokolle.de- www.webinfo1.de

09.10.04Italiener zeigt die “Magie der Biologie”,Mitteldeutsche Zeitung, Photography.

11.10.04Ausstellung am IPB, MitteldeutscheZeitung, p.9.

12.10.04Hoffman, P., Das “Stille-Post-Spiel” um dieLeibniz-Gemeinschaft, Magdeburger Volks -stimme.

14.10.04KULTUSMINSITERIUM SACHSEN-ANHALT,HOHER EU-BEAMTER INFORMIERT SICH ÜBERBIOTECHNOLOGIE-OFFENSIVE DES LANDESSACHSEN-ANHALT, PRESS RE LEASE.

22.10.04Im Dschungel der Erkenntnis, Chemie inunserer Zeit 38 /2004, p. 367.

22.10.04Wein, M., Lange Nacht war viel zu kurz,Scienta hallensis 10/2004, p. 6-7.

86


Press and

Public Relations

25.10.04PIEPLOW, S., NEUE DIREKTORIN AM LEIB NIZ-INSTITUT FÜR PFLANZENBIO CHEMIE, PRESSRELEASE.

This press release was also published by:- www.chemlin.de- www.innovationsreport.de- www.interconnections.de- www.uniprotokolle.de- www.weinbergcampus.de- Laborpraxis 12/2004, p. 14.- BIOforum 12/2004, p. 8

26.10.04Bank, M., Neue CD über Symbiosen,Mitteldeutsche Zeitung, p.20.

30.10.04Bank, M., Frau aus Amerika an der Spitze,Mitteldeutsche Zeitung, p. 20.

01.11.04PIEPLOW, S., NEUE WIRKSTOFFE AUS HEIMI-SCHENWÄLDERN, PRESS RELEASE

This press release are also published by:- www.chemie.de- www.chemlin.de- www.forstverein.de- www.geoscience-online.de- www.journalmed.de- www.medicineworldwide.de- www.physik-forum.de- www.shortnews.stern.de- www.tk-online.de- www.vdbiol.de

04.11.04Schilling, T., Mit dem Trick-”Phytolator”eine neue Medizin entdecken, Mittel deut -sche Zeitung, p. 22.

26.11.2004Pieplow, S., Pilze kontra Eiter und Bak -terien, Mitteldeutsche Zeitung, p. 22.

10.12.04Spilok, K., Neue Bioprodukte haben Zu -satznutzen, VDI Nachrichten.

20.12.2004Stäudner, F., Gehaltvolle Pilze, Leibniz 4, p.4.

20.12.2004Bank, M. Morphium in Zellen produziert,Mitteldeutsche Zeitung, p. 24.

Radio Transmission05.06.04Jacobs, D., Schnecklinge als Antibiotika,Mitteldeutscher Rundfunk, RadioSachsen-Anhalt

03.11.04Kienzlen, G., Medizin aus dem Wald,Deutschlandfunk, Forschung aktuell

Television Transmission26.03.04Neues Gebäude für Leibniz-Institut,Mitteldeutscher Rundfunk, Sachsen-Anhalt heute

16.06.04

87

Map and Impressum

Publisher: SCIENTIFIC REPORT 2003 - 2004Leibniz Institute of Plant Biochemistry May 2005

Leibniz Institute of Plant BiochemistryWeinberg 306120 Halle (Saale)Germany

www.ipb-halle.de

Editor: Sylvia PieplowPress and Public RelationsPhone: +49 (0) 3 45 - 55 82 11 10Fax: +49 (0) 3 45 - 55 82 11 19email: [email protected]

Layout & Design: Sylvia Pieplow

Graphics & Pictures: Leibniz Institute of Plant BiochenistryChristine KaufmannAnnett KohlbergBettina Hause and others

Copyright © 2005, all rights reserved Leibniz Institute of Plant Bio chemistry (IPB), Halle, Germany. No parts ofthis publication may be reproduced by any mechanical, photographic, electronic process, or in form of a photo-graphic recording, nor may be stored in a retrieval system, transmitted or otherwise copied for public or privateuse, without written permission from the publisher.

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