international research training group · 8 characterization of the putative vacuolar sugar...
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INTERNATIONAL RESEARCH TRAINING GROUP
8th Joint Symposium
Weiskirchen
September 2-4, 2019
2
Compendium
Monday, 09/02/19 Tuesday, 09/03/19 Wednesday, 09/04/19
Arrival
Registration Welcome
8:00 Breakfast 8:00 Breakfast
9:00 G. Khandpur 9:00 M. Schöppe
10:00 A. Khan 9:20 J. Oestreicher 9:20 M. Saurette
10:15 J. Bak 9:40 E. Zöller 9:40 C. Martins Rodrigues
10:30 A. Russo 10:00 J. Laborenz 10:00 K. Badior
10:45 Coffee break 10:20 Coffee break 10:20 Coffee break
11:00 M. Sicking 10:45 T. Bentrcia 10:45 K. Ravichandran
11:15 R. Brassard 11:00 M. Beggs 11:05 P. Schepsky
11:30 N. Yadao 11:15 X. Liu 11:25 Concluding remarks
11:50 F. Wollweber 11:35 Poster flash talks 11:35 PI & Trainee Meetings
12:10 Lunch 12:30 Lunch 12:00 Lunch
13:15 -
22:00
Visit of the “Saarschleife”
&
Saarburg “Saarweinfest”
13:30 Hike
End of Meeting
&
Departure
16:00 Poster Session
18:00 Guidance Committee
Meetings
19:00 Barbecue
10 min talk & 5 min discussion
15 min talk & 5 min discussion
Poster flash talks max. 3 min
3
Day One (Monday, September 2, 2019)
Arrival and Registration until 9.45 am 09:55-10:00 Ekkehard Neuhaus Welcome
Session 1 (Chair: Gurleen Khandpur)
10:00-10:15 Azkia Khan Characterization of the putative vacuolar sugar transporter AtERDL4
10:15-10:30 Jessi Bak Characterizing the “regulin” family of SERCA-regulatory peptides
10:30-10:45 Antonietta Russo Translocon-associated protein (TRAP) complex and co-translational protein transport
10:45 – 11:00 Coffee break
Session 2 (Chair: Duc Phuong Vu)
11:00-11:15 Mark Sicking Malfunctions of kidney disease associated Sec61α mutations
11:15-11:30 Raelynn Brassard Understanding PARL-dependent cleavage of PINK1 in mitochondrial health and Parkinson's disease
11:30-11:50 Nilam Yadao Differential sorting of mitochondrial preproteins via the TIM23 machinery
11:50-12:10 Florian Wollweber Regulation of MICOS activity during mitochondrial cristae remodelling
12:10 – 13:00 Lunch
13:15 – 22:00 Visit of the Saarschleife & Saarburg (“Saarweinfest”)
Day Two (Tuesday, September 3, 2019)
08:00 – 09:00 Breakfast
Session 3 (Chair: Jonas Höring)
09:00-09:20 Gurleen Kaur Khandpur Changes in amino acid availability severely impacts yeast cell growth and redox homeostasis
09:20-09:40 Julian Oestreicher Identification of Opt3 as a putative endoplasmic reticulum-localized glutathione disulphide exporter
09:40-10:00 Eva Zöller Mix23 – a novel yeast protein in the intermembrane space of mitochondria
10:00-10:20 Janina Laborenz The ER-protein EMA19: a role in mitochondrial import?
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10:20 – 10:45 Coffee break
Session 4 (Chair: Leo Bellin)
10:45-11:00 Teqiyya Bentrcia A novel modulator of TRPC6 channel function
11:00-11:15 Megan Beggs Increased calcium permeability across small intestine in early life conferred by claudin-2
11:15-11:30 Xiong Liu Molecular mechanism of TRPP3 regulation by calmodulin
11:30 – 11:35 Short break Poster flash talks (Chair: Cristina Martins Rodrigues)
11:35-11:38 Leo Bellin Structural and functional insights into the ATCase of Arabidopsis thaliana
11:38-11:41 Wassilina Bugaeva Physiological analysis of the plastid fatty acid export proteins in Arabidopsis thaliana
11:41-11:44 Damayante Das The novel role of TMEM33 as a regulator of voltage gated Potassium channels
11:44-11:47 Daniel Fajonyomi Unique gating properties of Kv1.2 glycosylation deficient mutants
11:47-11:50 Jonas Höring Micellization thermodynamics of fluorinated and hydrogenated surfactants
11:50-11:53 Annalisa John Identification of chloroplast envelope proteins with critical importance for cold acclimation of Arabidopsis
11:53-11:56 Xiaobing Li Analysis of intracellular trafficking & localization of the human kidney anion exchanger 1 (kAE1) in yeast
11:56-11:59 Hasib Sarder Dissecting intracellular trafficking and mis-trafficking of human kidney AE1 in yeast and mammalian cells
11:59-12:02 Andrea Tirincsi Transmembrane protein 109 (TMEM109), a putative hsnd3 protein
12:02-12:05 Pratiwi Prananingrum The warm and high light stress; insight on a plastidic sugar transporter
12:05-12:08 Duc Phuong Vu Vacuolar sucrose mobilization is critical for the development of Arabidopsis thaliana
12:08-12:11 Janet Zhou The influence of selenium on arsenic hepatobiliary transport
12:30 – 13:30 Lunch
13:30 – 16:00 Hike / Time at free disposal (e.g. swimming, sports)
5
16:00 – 18:00 Poster Session (including coffee break)
1 Leo Bellin Structural and functional insights into the ATCase of Arabidopsis thaliana
2 Wassilina Bugaeva Physiological analysis of the plastid fatty acid export proteins in Arabidopsis thaliana
3 Damayantee Das The novel role of TMEM33 as a regulator of voltage-gated potassium channels
4 Daniel Fajonyomi Unique gating properties of glycosylation-deficient Kv1.2 channels
5 Jonas Höring Micellization thermodynamics of fluorinated and hydrogenated surfactants
6 Annalisa John Identification of chloroplast envelope proteins with critical importance for cold acclimation of Arabidopsis
7 Hasib Sarder Dissecting intracellular trafficking and mis-trafficking of human kidney AE1 in yeast and mammalian cells
8 Xiaobing Li Analysis of intracellular trafficking & localization of the human kidney anion exchanger 1 (kAE1) in yeast
9 Andrea Tirincsi Transmembrane protein 109 (TMEM109), a putative hsnd3 protein
10 Pratiwi Prananingrum The warm and high light stress; insight on a plastidic sugar transporter
11 Duc Phuong Vu Vacuolar sucrose mobilization is critical for the development of Arabidopsis thaliana
12 Janet Zhou The influence of selenium on arsenic hepatobiliary transport
In order to allow guidance committee meetings at the posters and that the presenting trainees can visit the posters of their colleagues, the posters remain hanging the whole evening.
18:00 – 19:30 Guidance Committee Meetings Free choice of meeting places, e.g. hotel lobby, terrace, conference room, at the posters.
18:00–18:20
H. Sarder M. Schmitt E. Neuhaus E. Cordat
J. Laborenz J. Herrmann S. Lang N. Touret
F. Wollweber M. van der Laan M. Hoth J. Casey
T. Bentrcia V. Flockerzi T. Alexander
New trainees/ trainees without guidance committees L. Bellin (T. Möhlmann) J. Höring (S. Keller) A. John (E. Neuhaus) J. Oestreicher (B. Morgan) A. Tirincsi (S. Lang) M. Sicking (S. Lang)
18:20–18:40
A. Russo R. Zimmermann M. Hoth X-Z. Chen
G. Khandpur B. Morgan M. van der Laan N. Touret
W. Bugaeva E. Neuhaus K. Philippar J. Casey
K. Ravichandran J. Rettig V. Flockerzi ?
18:40 –19:00
C. Martins Rodrigues E. Neuhaus K. Philippar X.-Z. Chen
P. Schepsky J. Engel V. Flockerzi (H. Kurata)
N. Yadao M. van der Laan J. Herrmann J. Casey
M. Schöppe B. Niemeyer S. Lang T. Alexander
19:00-19:20
E. Zöller J. Herrmann M. van der Laan T. Alexander
D. Vu E. Neuhaus T. Möhlmann J. Lemieux
X. Li M. Schmitt B. Morgan E. Cordat
19:00 Barbecue at the “Eventhütte”
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Day Three (Wednesday, September 4, 2019)
08:00 – 09:00 Breakfast
Session 5 (Chair: Janina Laborenz)
09:00-09:20 Mona Schöppe Characterization of a novel splice variant of the stromal interaction molecule1 (STIM1)
09:20-09:40 Matthew Saurette Phosphorus source and intestinal absorption: inorganic phosphate is absorbed by the paracellular pathway
09:40-10:00 Cristina Martins Rodrigues To BEet or not to BEet. A short story about sugar beet transporters and their impact on cold acclimation
10:00-10:20 Katie Badior Investigation of RBC aging
10:20 – 10:45 Coffee break
Session 6 (Chair: Xiaobing Li)
10:45-11:05 Keerthana Ravichandran Characterization of flower containing vesicles in mouse cytotoxic T lymphocytes
11:05-11:25 Pauline Schepsky Slack (Slo2.2) K+ channels and the tight junction protein Claudin-12 in the cochlea
11:25-11:35 Barbara Niemeyer &
Joe Casey
Concluding remarks
11:35 – 12:00 Trainee & PI Meeting
12:00 - 13:00 Lunch / End of Meeting
7
Abstracts in chronological order
8
Characterization of the putative vacuolar sugar transporter, a homolog of
AtERDL6
Azkia Khan, Patrick Klemens & H. Ekkehard Neuhaus Department of Plant Physiology, University of Kaiserslautern, Germany
In plants, the central vacuole is the largest organelle and essential for plant growth. Within a
cell, the vacuole can serve as temporary storage for many metabolites and signaling
compounds. Both the availability of sugars and the accumulation of macro- and
micronutrients ensure proper plant growth. Sugars in plants provide energy for metabolic
processes as well as act as precursors in the synthesis of starch and amino acids. The
movement of sugar to and from the vacuole rely on numerous vacuolar transporters.
Vacuolar sugar transport is mediated by sucrose transporter family, the monosaccharide
transporters (MST) and members of a family called SWEET. Early response to dehydration
(ERD) 6–like1 (ESL1), is a member of the ERD-6-like clade, is a vacuolar protein and a
member of MST family. Other members of this clade are also targeted to the tonoplast. The
current study is on a homolog of AtERDL6, which we have identified as a vacuolar located
sugar carrier, induced by cold, drought and salt stress. GUS promoter analysis revealed its
expression mainly in the roots and pollens. Overexpressor and knockout plants also exhibit
variation in sugar accumulation under different stress conditions.
Characterizing the “regulin” family of SERCA-regulatory peptides
Jessi Bak & Howard Young Department of Biochemistry, University of Alberta, Edmonton, Canada
Calcium signalling is important for a multitude of physiological processes like skeletal and
cardiac muscle function, neurotransmitter release, and the cell cycle. Because of this central
role, it is tightly regulated at the intracellular level through the sarco(endo)plasmic calcium
ATPase, or SERCA. SERCA is a P-type ATPase that utilizes ATP to pump two calcium ions
across the membrane of the sarcoplasmic or endoplasmic reticulum membrane into the
lumen of the organelle for storage. This action is further regulated by single-pass
transmembrane peptides that interact with SERCA. Two regulators, phospholamban and
sarcolipin, have been investigated for a number of years but advances in bioinformatic
screening have led to the expansion of SERCA-regulatory peptides - a group that we now
refer to as the “regulins.” The regulins include myoregulin, another-regulin, endoregulin, and
DWORF, and these regulators are found in both muscle and non-muscle tissue. Our lab
studies the affect of these regulators on SERCA by co-reconstituting these components into
proteoliposomes and then measuring SERCA activity through a coupled-enzyme assay.
Here, our recent studies to understand these regulators will be presented.
Translocon-associated protein (TRAP) complex and co-translational protein
transport
Antonietta Russo Department of Medical Biochemistry and Molecular Biology, Saarland University, Homburg
A milestone in the understanding of protein translocation is the “Signal Hypothesis” proposed
by G. Blobel in 1971, which explains how the proteins are translocated from the cytosol into
the endoplasmic reticulum (ER). The signal peptide (SP) typically has the same structure but
it is very heterogeneous in protein sequence composition, this implies a fast evolution
probably connected with the mature protein.
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Nowadays, we know that the complexity of the SP meant to control many biological
processes: signal recognition particle (SRP) binding, interaction with the translocon Sec61
(gating), early folding prevention, signal peptidase (SPase) interaction and cleavage, and
even post-cleavage functions such as antigen presentation.
In the last years, much progress has been made to address the structure of the translocation
machinery, thanks also to the improved microscopic techniques such as Cryo-EM and Cryo-
ET; most of the structures are known, but the role of some components is still unclear.
TRAP is a heterotetrameric complex, α-β-γ-δ subunits, is located close to Sec61 complex,
where also TRAM, OST, SPase, Calnexin are present. It is a Ribosome Associated Protein
(RAP) that is involved in co-translation translocation; however, its role needs to be
determined. I will present computational and experimental results that can contribute to
shedding more light on the function(s) of TRAP at the molecular level and the entire
biological process.
Malfunctions of kidney disease associated Sec61α mutations
Mark Sicking1, Martina Zivna
2, Adolfo Cavalié
3, Richard Zimmermann
1 & Sven Lang
1
1Medical Biochemistry and Molecular Biology, Saarland University, Homburg;
2 Institute of Inherited Metabolic Disorders,
Charles University, Prague; 3 Experimental and Clinical Pharmacology and Toxicology, Saarland University, Homburg
The endoplasmic reticulum (ER) is a crucial organelle concerning the calcium homeostasis
and transport of nascent polypeptides in human cells. The heterotrimeric Sec61 complex is
an essential protein in the ER membrane and ensures a flawless functionality of the
mentioned processes. This complex forms an aqueous pore in the membrane, and the
entrance for multiple nascent proteins during their transport into or across the ER membrane.
During or shortly after the polypeptide transport, calcium leaks out through the complex from
the ER lumen into the cytosol in a regulated manner. Mutations in the main component of the
complex, the Sec61α protein, show multiple disease phenotypes in humans.
The two here investigated mutations of Sec61α, SEC61A1-V67G and SEC61A1-T185A, are
located at central positions of the protein and both variants cause forms of the Autosomal-
dominant tubulointerstitial kidney disease (ADTKD). However, the underlying pathogenic
mechanisms causing the Sec61α-related forms of ADTKD are unknown. In our cell culture
approaches we are using transgenic kidney cells, HEK293 cells, which express an
endogenous wild type and a mutated form of the Seec61α protein. This heterozygosity
mimics the patient families situation. Both mutations show a reduced protein transport
efficiency of specific substrates, both kidney related and unrelated polypeptides. The
reduced transport of affected substrates was traced back to the composition of their N-
terminal signal sequences. Furthermore, both mutations show multi-level irregularities of
calcium homeostasis. This includes, (i) the altered abundance of key players of calcium
homeostasis like SERCA2 or SOCE associated proteins, (ii) the suppressed sensitivity of the
mutants to the calcium leak modulating drug Eeyarestatin1, (iii) the slower Sec61-mediated
calcium leakage in case of the T185A variant, and (iv) the reduced calcium storage capacity
of the ER. Yet the small differences between the two Sec61α mutations, regarding protein
transport and calcium homeostasis could explain the phenotypic differences of the patients
and open up an interesting avenue of investigation. With the SEC61A1 gene being
expressed in all nucleated cells, there is still no explanation for the organ specificity of
SEC61A1 mutations associated diseases. Hence, we are currently studying other ER related
10
processes like its ATP supply to address the issue of cell type specific defects and the
aforementioned phenotypic originalities.
Understanding PARL-dependent cleavage of PINK1 in mitochondrial health and
Parkinson's disease
Raelynn Brassard1, Elena Arutyunova
1, Laine Lysyk
1, Emmanuelle Tayki
2, Hélène Lemieux, Nicolas Touret
1 & Joanne
Lemieux1
Department of Biochemistry1 and Neuroscience and Mental Health Institiute
2, University of Alberta, Edmonton, AB, T6G 2R3.
Parkinson’s disease (PD) is a devastating neurodegenerative disease that is characterized
by a loss of dopaminergic neurons located in the substantia nigra. Mitochondrial dys-
regulation has been observed in numerous neurodegenerative diseases, including PD, due
to the high energy requirements of neuronal tissues.
Genetic mutations in the gene encoding for PINK1, have been associated with cases of early
onset PD. In a neuroprotective role, PINK1 accumulates on damaged mitochondria, flagging
them for destruction; while in healthy cells PINK1 is turned over by the PARL protease. PD
associated mutations are found near the PARL cleavage site and may lead to a loss of
PINK1 cleavage.
To reveal the etiology of PINK1 PD variants, in vivo analysis was conducted. Cleavage and
localization of the variants were assessed using confocal imaging and western blot analysis
of HeLa cells transfected with PD linked PINK1 variants. Like wild-type PINK1, all variants,
except the R98W mutant, displayed a diffuse pattern of PINK1 throughout the cell. The
PINK1-R98W PD variant displays high retention of the protein in the mitochondria.
These results were compared to in vitro cleavage analysis with recombinant PARL protease
and substrate, which shows no cleavage defect for the R98W variant, suggesting this variant
has defects in mitochondrial trafficking and not PARL processing. Thus, in these cells,
healthy mitochondria may be targeted for destruction with the PINK1-R98W mutation. To
evaluate this, assessment of mitochondrial respiratory function will be conducted. This work
provides insight into mutations causing early-onset inherited forms of PD.
Differenial sorting of mitochondrial prproteins via the TIM23 machinery
Nilam Yadao & Martin van der Laan Department of Medical Biochemistry and Molecular Biology, Saarland University, Homburg/Saar
The majority of mitochondrial proteins is encoded by nuclear genes and synthesized as
precursors in the cytosol. Mitochondrial preproteins contain a variety of import and sorting
signals that guide them to their destined locations within the organelles. The focus of my
project is the translocation and membrane-insertion of preproteins with amino-terminal
targeting signals. These proteins pass the outer membrane via the TOM complex and are
then taken over by the presequence translocase of the inner mitochondrial membrane, the
TIM23 complex. Depending on the physicochemical properties and sorting information of the
preproteins, they are either translocated completely into the matrix or integrated into the
inner membrane. Matrix import requires the interaction and close cooperation of TIM23 with
the presequence translocase-associated import motor. Membrane insertion of hydrophobic
preprotein segments via a stop-transfer mechanism is supported by the direct physical
coupling of proton-pumping respiratory chain complexes. It is unknown how transmembrane
segments are recognized by the TIM23 machinery and how they laterally escape from the
translocon into the lipid bilayer. Recent studies from my laboratory have shown that the small
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membrane-integral TIM23 subunit Mgr2 controls the lateral release of preproteins, likely via
interactions with charged amino acid residues flanking transmembrane segments. Site-
specific photo-crosslinking data suggest that Tim17 differentially interacts with hydrophilic
and hydrophobic preprotein segments within the protein-conducting pore of the TIM23
complex. In my studies, I use purified mitochondria from knock-out and conditional tim
mutants of the baker's yeast (Saccharomyces cerevisiae) to unravel how Tim17 and Mgr2
cooperate in the decoding of inner membrane sorting signals and the release of preprotein
segments into the phospholipid bilayer, a process referred to as "lateral gating". Supported
by IRTG1830
Regulation of MICOS activity during mitochondrial cristae remodelling
Florian Wollweber & Martin van der Laan
Department of Medical Biochemistry and Molecular Biology, Saarland University, Homburg/Saar
Mitochondria show a remarkable structural complexity as a result of their endosymbiotic
origin. Their double-membrane architecture is crucial for many essential functions in
metabolism and cell-fate decisions. The mitochondrial outer membrane connects
mitochondria to the cytosol and other organelles while the inner membrane harbours the
oxidative phosphorylation machinery for ATP synthesis and consists of a boundary region as
well as tubular or disc-shaped protrusions, termed cristae. One of the main organisers of this
intricate inner membrane ultrastructure is the mitochondrial contact site and cristae
organising system (MICOS), which is a large and highly conserved protein machinery that
can be found in all cristae-containing eukaryotes.
MICOS is essential for maintaining crista junction structure and creates a central hub for
mitochondrial biogenesis. Recent studies found that the two core subunits Mic10 and Mic60
form genetically and biochemically separable modules with distinct functions in membrane-
shaping and membrane-bridging to stabilise the membrane curvature at crista junctions and
connect mitochondrial protein machineries across multiple sub-organellar compartments. As
cristae are highly dynamic structures that have to adapt to e.g. altered metabolic demands of
the cell, the regulation of MICOS activity plays a pivotal role. We are currently investigating
how accessory MICOS subunits, such as the lipid-binding components Mic26/Mic27 and the
redox-regulated subunit Mic19 modulate the assembly of MICOS core subunits to coordinate
MICOS activities for cristae maintenance and remodelling.
Changes in amino acid availability severely impacts yeast cell growth and
redox homeostasis
Gurleen Kaur Khandpur1,4
, Martin Van der Laan2, Nicolas Touret
3 & Bruce Morgan
1
1Department of Biochemistry, University of Saarland, Germany
2Faculty of Medicine Medical Biochemistry and Molecular
Biology, University of Saarland, Germany, 3Department of Biochemistry, University of Alberta, Canada,
4Department of Biology,
Technical University of Kaiserslautern, Germany
Changes in amino acid handling have been observed in a wide-range of human pathologies
including diabetes and cancer. We used Saccharomyces cerevisiae as a model to
investigate how changes in amino acid availability influences cell growth and fitness.
Intriguingly, we observe that increasing the general availability of amino acids relative to the
availability of leucine leads to striking growth defects on glucose containing media. We also
observed that increasing the concentration of Ehrlich amino acids (amino acids which are
degraded to fusel acids/alcoholic compounds) in the media, severely impacts the growth of
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the cells which can be rescued by providing more Leucine. Furthermore, the deletion of Ilv2,
Ilv5 or Ilv3 (enzymes involved in leucine bio-synthetic pathway) makes the cell growth better
than wild type yeast cell in presence of increased amino acids availability. This growth
phenotype suggests that Ilv3 is the switching point and links the cellular growth and redox
homeostasis. Surprisingly, the amino acid-dependent growth phenotypes are completely
absent when cells grown in media containing non-fermentable carbon sources. We found
that deletion of the mitochondrial external NADH dehydrogenase-1 (Nde1) in combination
with Cox6 (an essential component of complex IV) partially rescued amino acid-dependent
growth phenotypes. However, deletion of the Nde1 homolog, Nde2, in combination with Cox6
had the opposite effect, further decreasing growth rate. We speculate that specific respiratory
chain components, but not the respiratory chain function per se, can play an important role in
‘buffering’ cells against these changes, although the mechanism remains to be determined.
Identification of a putative endoplasmic reticulum glutathione disulphide
exporter
Julian Oestreicher & Bruce Morgan Department of Biochemistry, Saarland University, Germany
In the endoplasmic reticulum the glutathione redox couple is generally believed to be more
oxidized than in the cytosol. However, it is unclear how glutathione redox homeostasis is
maintained with lacking glutathione reductase. In this study, we investigated the possibility
that the glutathione disulphide (GSSG) is exported from the secretory pathway in the cytosol
for reduction.
Specifically, we investigated the role of an uncharacterized, putative oligopeptide transporter
homolog, Opt3. We observed a synthetic lethality when OPT3 was deleted in combination
with either of the two glutathione biosynthetic enzymes, GSH1 or GSH2. We found that
manipulation of OPT3 expression levels affected whole cell GSSG concentrations, with
overexpression leading to a strong decrease in cellular GSSG content. These findings would
be consistent with a transport of GSSG to the cytosol where it is reduced. Upon targeting of
the glutathione synthetase, Gsh2, specifically to the ER, the impact of Opt3 expression level
on total cellular GSSG content was amplified. Finally, we demonstrate that the impact of
Opt3 on cellular GSSG levels is independent of the vacuolar-localized GGSG transporter,
Ycf1, therefore speaking against a role of Opt3 in regulating vacuolar GSSG storage. In
summary, we hypothesize that Opt3 mediates the export of GSSG from the endoplasmic
reticulum to the cytosol.
Mix23 – a novel yeast protein in the intermembrane space of mitochondria
Eva Zöller & Johannes Herrmann Department of Cell Biology, University of Kaiserslautern, Germany
Yeast cells synthesize the majority of their mitochondrial proteins in the cytosol which then
get imported into the mitochondria through their outer (OMM) and inner (IMM) mitochondrial
membranes. A variety of proteins targeted to the intermembrane space (IMS) relies on the
specific MIA pathway. Here, proteins enter the IMS in a reduced state and bind to Mia40, an
oxidoreductase that oxidizes the target proteins, which traps them in the IMS. In a proteome
screen, the cysteine containing protein Mix23 with unknown function was found to be
localized in the IMS. We could verify the IMS localization, the interaction with Mia40, and the
MIA dependent import of Mix23. In a RNA sequencing approach, we found Mix23 to be
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upregulated during import stress. Surprisingly, Mix23 is the only MIA substrate to be
upregulated under those circumstances. Furthermore, Mix23 is co-regulated with Mia40 and
the proteasome by the transcription factor Rpn4. This upregulation is specific to import stress
since it is not triggered by cytosolic aggregates. The import efficiency of MIA substrates was
not effected in a MIX23 deletion strain. Overexpression of Mix23 leads to a strong growth
phenotype. It is known that IMS proteins can be retro-translocated back into the cytosol for
their degradation. Due to the direct interaction of Mix23 with Mia40 and its cysteine rich
sequence, we hypothesize Mix23 to be one player in the so far uncharacterized retro-
translocation pathway. A number of experiments is planned to further investigate into this
direction.
The ER-protein EMA19: a role in mitochondrial import?
Janina Laborenz & Johannes Herrmann Department of Cell Biology, University of Kaiserslautern, Germany
Most mitochondrial proteins are initially synthesized in the cytosol as precursor proteins and
imported into mitochondria. While the import of soluble mitochondrial proteins was well
studied in the past, only little is known how cells manage to translocate the many
hydrophobic membrane proteins of the inner membrane. We developed a genetic screen in
yeast cells to identify genes that are critical for the efficient translocation of the hydrophobic
inner membrane protein Oxa1 into mitochondria. Surprisingly, in this screen we identified
several so far uncharacterized, though conserved ER proteins that are crucial for
mitochondrial targeting of Oxa1. By combining biochemical and genetic analysis we
characterized the function of the protein Djp1 in this process, which belongs to the family of
J-domain cochaperones of the Hsp70 system. We found that a large fraction of the newly
synthesized Oxa1 precursor associates with the ER surface from where it is recognized by
Djp1 to be directed to the mitochondrial outer membrane translocase. We propose that the
ER surface can serve as a collection system that facilities intracellular protein transport to
mitochondria. We called this import route the ER-SURF pathway. In addition to Djp1, we
identified an uncharacterized ER membrane protein, Ema19. First results about the function
of this protein in the context of mitochondrial preprotein sorting will be presented.
A novel modulator of TRPC6 function
Teqiyya Bentrcia & Veit Flockerzi Department of Pharmacology & Toxicology, Saarland University, Homburg, Germany
The canonical transient receptor potential channel 6 (TRPC6), a Ca2+ permeable cation
channel comprises four TRPC6 α subunits and may contribute to platelets function and
haemostasis. To identify the TRPC6 protein and potential β subunits of the channel in human
platelets, we generated antibodies for TRPC6, which enable protein detection in tissue
homogenates by Western blots. First, we established an antibody-based affinity purification
procedure to enrich solubilized TRPC6 protein from platelets. The bound TRPC6 protein was
eluted under non denaturing condition, run on blue native gels and analysed by mass
spectrometry. By the latter approach, the G-protein-coupled receptor kinase interacting
protein-1 (GIT1) and proteins of the phospholipase Cγ, ARHGEFs, ERK1, PAK2 and IP3R
pathway were found to be associated with the TRPC6 protein. In contrast, none of these
proteins were retained by a non-specific antibody used as a control. The physical interaction
between TRPC6 and GIT1 was confirmed by coimmunoprecipitation and in vitro pull down
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assays. We could show that the GIT1 protein binds to the TRPC6 protein via its ankyrin
repeat domain. Similarly, coimmunoprecipitation showed that the N-terminus of TRPC6 is
essential for TRPC6-GIT1 interaction. By calcium imaging as well as by patch clamp
recordings, calcium entry and TRPC6 currents are activated by diacylglycerol or its derivative
OAG. The OAG-induced calcium entry and current in HEK293 cells stably expressing Trpc6
cDNA was reduced in the presence of the GIT1 protein indicating a potential inhibitory effect
of GIT1 on TRPC6 mediated calcium entry and current. Calcium entry is required for cell
migration. In an in vitro migration scratch assay, we could show that the presence of GIT1,
results in slower migration. Furthermore, depletion of endogenous GIT1 protein by
overexpression of the TRPC6 N-terminus abolishes the inhibitory effect of GIT1 on TRPC6
function and results in larger currents and faster cell migration. This study identifies a novel
TRPC6 channel regulatory subunit (GIT1) which has a modulatory impact on TRPC6 channel
function. To elucidate the roles of TRPC6 and GIT1 in the phospholipase Cγ, ARHGEFs,
ERK1, PAK2 and IP3R containing signalom is the aim of our ongoing study.
Increased calcium permeability across small intestine in early life conferred by
claudin-2
Megan Beggs1, Allen Plain
1, Justin J. Lee
1 & R. Todd Alexander
1,2
1Physiology,
2Pediatrics, University of Alberta, Canada
Infants and children must maintain a net positive calcium (Ca2+) balance in order to achieve
an optimal peak bone mineral density by early adulthood. Across the small intestine, Ca2+ is
absorbed via transcellular and paracellular pathways although the molecular details of these
pathways in early life are not well delineated. Claudins are tight junction proteins that confer
selective permeability to epithelia. Claudins-2, -12 and -15 are expressed and have been
implicated in paracellular Ca2+ permeability across intestinal epithelia. To date, the functional
contribution of these claudins to the Ca2+ permeability (PCa2+) of small intestine segments has
not been defined. The objective of this study was therefore to 1) determine if PCa2+ across
small intestine segments changed with postnatal development 2) to asses whether claudin-2
or -12 contribute PCa2+
to the small intestine. To this end, basolateral to apical 45Ca2+ fluxes
on ex vivo intestinal segments in Ussing chambers were employed to indirectly assess
permeability to Ca2+. We observed greater 45Ca2+ flux in P14 vs 2-month mice across the
duodenum (31.8 ± 2.2 vs 9.5 ± 2.0 nmol/h/cm2, P<0.0001) and jejunum (42.3 ± 3.9 vs 21.8 ±
3.5 nmol/h/cm2, P<0.01) but not ileum (40.5 ± 3.2 vs 30.1 ± 5.7 nmol/h/cm2, P=0.13). Next,
we measured PCa2+ directly in Ussing chambers using diffusion potentials. PCa
2+ was
significantly greater in younger mice across the duodenum (1.43 ± 0.04 vs 1.13 ± 0.08 x10-
4cm/s, P<0.01), jejunum (1.64 ± 0.09 vs 0.84 ± 0.04 x10-4cm/s, P<0.0001) and ileum (1.72 ±
0.08 vs 0.81 ± 0.05 x10-4cm/s, P<0.0001). To determine if claudin-2 or -12 facilitates greater
PCa2+ at P14, diffusion potential experiments were repeated on Cldn2 or Cldn12 knockout
(KO) mice. PCa2+ was decreased in Cldn2 KO mice at P14 across the jejunum (1.85 ± 0.09 vs
1.24 ± 0.07 x10-4cm/s, P<0.0001) and ileum (1.92 ± 0.07 vs 1.09 ± 0.06 x10-4cm/s,
P<0.0001) to levels not different than 2-month WT and KO mice. No differences were
observed between Cldn12 knockout mice compared to wildtype littermates at either age. We
conclude that the younger mice have enhanced Ca2+ permeability along the small intestine
relative to older animals. Further, the increased jejunal and ileal Ca2+ permeability is
mediated by claudin-2 which likely contributes to a positive calcium balance for normal
growth.
15
Characterizing the role of CAM in TRPP3 channel function
Xiong Liu & Xing-Zhen Chen Department of Physiology, Membrane Protein Diseases Research Group, University of Alberta, Edmonton, AB, Canada, T6G 2H7
Transient receptor potential (TRP) polycystin-3 (TRPP3) is a non-selective cation channel
activated by calcium ions and protons and is involved in regulating ciliary calcium
concentration, hedgehog signaling and sour tasting. Previous reports have shown that
TRPP3 activation is followed by desensitization but the molecular basis underlying the
channel activation and the ensuing desensitization is not well understood. Calmidazolium
(CMZ), which is a commonly used calmodulin (CaM) antagonist, has been reported to
activate TRPP3 channel. Since CaM has been involved in the Ca2+ dependent regulation of
many TRP proteins, whether and how CaM regulates TRPP3 channel function has never
been explored. One possible mechanism is that CaM directly binding to TRPP3 channel
induces inhibitory effect and therefore the CaM antagonist, CMZ, will consequently activate
the channel. Another explanation is that CaM activating kinase, Ca2+/CaM dependent protein
kinase II (CaMKII) is inhibited by CMZ, leading to a functional TRPP3 form. In this study, we
observed that CaMKII and CaM decrease TRPP3 channel function. TRPP3 was
phosphorylated by CAMKII with ongoing experiments trying to identify the phosphorylation
site(s). We are also testing possible direct binding of CaM to TRPP3 and the role of Ca2+ in
the binding. These studies would allow to uncover how CaM blocks TRPP3 function. We
propose to verify that CaM promotes the inhibition of TRPP3 channel function through direct
interaction and CaMKII’s phosphorylation on TRPP3.
Support: Department of Physiology stipend, 75th Anniversary Graduate Student Award (to
XL), NSERC Discovery Grant, and Kidney Foundation of Canada Biomedical Grant (to XZC).
(Poster flash talk 1)
Structural and functional insights into the ATCase of Arabidopsis thaliana
Leo Bellin & Torsten Möhlmann
Department of Plant Physiology, University of Kaiserslautern
Pyrimidine nucleotides are some of the most crucial cellular metabolites for plant
development and growth. In plants, aspartate transcarbamoylase (ATCase), the enzyme
catalyzing the condensation of aspartate (Asp) and carbamoyl phosphate (CP) to carbamoyl
aspartate (CASP), is the control point of the de novo biosynthesis of pyrimidines.
This essential metabolic enzyme is unique in plants for being directly inhibited by uridine
mono-phosphate (UMP), the final product of the biosynthetic pathway. Despite numerous
biochemical and structural studies on ATCases from bacteria, fungi and animals, little is
known about ATCases in plants and no structural information has been reported so far.
In the course of this project we determined 6 different high resolution crystal structures of the
ATCase from Arabidopsis thaliana, providing first atomic detail about the catalytic and
regulatory mechanisms of this essential enzyme in plants. The structural data combined with
mutagenic, ITC and in vivo assays reveal the mechanism of inhibition by UMP.
In addition, we found out that the plant ATCase is a trimeric protein where only one subunit
can catalyze the reaction at a time. These results confirm that despite sharing an overall
structural similarity with the enzyme from other kingdoms, the ATCase in plants functions in a
different manner that could be targeted for the design of new herbicide strategies. Based on
the acquired structural and functional knowledge about Arabidopsis ATCase we propose
16
different approaches that could guide us in the development of plant specific ATCase
inhibitors.
Further and in addition to first insights into the structure- experiments for the relevance of the
ATCase in living plants have been performed. Thereby a reduction of the transcript by
artifical micro RNA leads to a drastical loss of biomass and to deficits in the development.
(Poster flash talk 2)
Physiological analysis of the plastid fatty acid export proteins in Arabidopsis
thaliana
Wassilina Bugaeva, Anne Könnel, Janick Peter & Katrin Philippar
Molecular Plant Biology, Center for Human- and Molecular Biology, Saarland University Saarbrücken, Germany
In plants, fatty acids (FAs) are synthesized in the plastid stroma and become available for
lipid assembly mainly in the form of long-chain FAs (C16–18). Some of these FAs are
integrated into lipids inside plastids (prokaryotic pathway), but the majority is exported to the
ER for further elongation, acyl editing, and lipid assembly (eukaryotic pathway). The
identification of FAX, a novel fatty acid export protein in the inner envelope membrane of
chloroplasts significantly contributes to the understanding of the FA transport mechanism
and the importance of FAX1 in plant development, biomass formation and fertility. In
Arabidopsis thaliana, seven proteins belong to the FAX family and besides FAX1, FAX2 and
FAX3 are also integrated into the inner envelope of chloroplasts. FAX2 and FAX3 are
assumed to partly complement FAX1 function due to increased transcript levels in leaves of
fax1 knockouts. Segregation analysis of fax1 single mutants and fax1/fax3 double mutants
on agar plates with and without sucrose shows a reduced amount of homozygous fax1/3
double mutants due to a missing complementation of FAX1 by FAX3. On soil, homozygous
fax1/3 double mutants are not viable, consequently the loss of both genes leads to a lethal
effect without additional carbon source supply.
Reference: Li et al. (2015) PLoS Biology. 13(2): e1002053
(Poster flash talk 3)
The novel role of TMEM33 as a regulator of voltage-gated potassium channels
Damayantee Das & Harley Kurata Department of Pharmacology, Faculty of Medicine & Dentistry, University of Alberta, Edmonton, Canada
The voltage gated potassium (Kv) channel Kv1.2 is a prominent potassium channel in the
CNS, playing an important role in regulating neuronal excitability and plasticity. Kv1.2
knockout mice have elevated seizure susceptibility, leading to 100% mortality by the third
post-natal week. Several channelopathies, including cerebellar ataxia, epileptic
encephalopathy, and hereditary spastic paraplegia are linked to Kv1.2 mutations. Kv1.2
channel structural and functional properties have been well studied, but its interactions with
regulatory proteins are still incompletely understood. Therefore, we aim to identify Kv1.2
channel regulatory proteins, in order to understand mechanisms that may impact channel
gating or proteostasis. Using mass spectrometry analysis of immunoprecipitated Kv1.2
complexes, we identified and validated several unknown regulatory proteins. One interacting
partner that stood out was TMEM33, an ER-resident molecule with effects on Kv1.2 channel
expression. Although TMEM33 is a conserved protein, little is known about its function, and it
is had not been previously recognized as a regulator of voltage-gated ion channels. This
17
study investigates mechanisms underlying TMEM33 regulation of Kv1.2, and how this may
lead to altered neuronal excitability.
EGFP-tagged TMEM33 prevents Kv1.2 trafficking to the cell surface and hinders maturation
into a fully glycosylated form. A strong suppression of Kv1.2 currents by EGFP-TMEM33 is
also observed. EGFP-TMEM33 generates a strong signal with Kv1.2 luminescence donors in
a BRET assay, suggesting their close proximity. TMEM33 is therefore a strong regulator of
Kv1.2 expression and function. Unlike Kv1.2, EGFP-TMEM33 did not affect Kv1.5 current
level and maturation. Current density measurements of different Kv1.2-Kv1.5 chimeras
suggested that substitution of the Kv1.2 pore abolished the effects of EGFP-TMEM33.
Untagged TMEM33, however, promotes total expression of Kv1.2 and rescues channel
surface expression in a TMEM33-knockout cell line. The distinct effects of N-terminally
tagged vs. untagged TMEM33 suggest that N-terminus of TMEM33 might have an important
functional role. Using a chimeric approach combining segments of Kv1.2 (TMEM33-
sensitive) and Kv1.5 (TMEM33-insensitive), we identify the pore and C-terminus of Kv1.2 as
critical determinants of TMEM33 sensitivity.
Channelopathies linked to Kv1.2 are associated with several diseases related to cell
excitability and epilepsy. Developing new knowledge into the regulatory mechanisms of
channel expression will contribute to a better understanding of these severe diseases. This
research will map out detailed biological functions of TMEM33, a poorly understood ion
channel regulator.
(Poster flash talk 4)
Unique gating properties of glycosylation-deficient Kv1.2 channels
Daniel Fajonyomi & Harley Kurata Department of Pharmacology, Faculty of Medicine & Dentistry, University of Alberta, Edmonton, Canada
Rationale: Kv1.2 channels are neuronal voltage-gated potassium ion channels belonging to
the Shaker (Kv1) family. Kv1.2 channels exhibit a unique behaviour termed use-dependent
activation (UDA), in which repetitive depolarizing stimuli produce progressive and
considerable increases in channel activity. However, the underlying molecular mechanism of
this behaviour is unknown. Previous work has demonstrated that mutation of a threonine at
position 252 of Kv1.2 to an arginine (Kv1.2 [T252R]) considerably diminishes use
dependence. Several lines of evidence have suggested that use-dependent activation of
Kv1.2 channels is due to an extrinsic regulatory molecule. To investigate the role of
glycosylation in the interaction of Kv1.2 with a candidate regulatory protein, the Kv1.2 N-
linked glycosylation site at residue N207 was mutated to an alanine (Kv1.2 [N207A]) or
glutamine (Kv1.2 [N207Q]), and effects on gating and maturation were tested.
Results: Kv1.2 [N207A] and Kv1.2 [N207Q] mutant channels exhibit dramatically slower
activation kinetics, slower deactivation kinetics, and a positive shift in voltage-dependence of
activation compared to WT Kv1.2. A signature feature of use-dependent activation is that
repetitive activation of Kv1.2 channels leads to accelerated activation. Similarly, the
activation kinetics of Kv1.2 [N207A] are markedly accelerated with prior stimuli. Furthermore,
the T252R mutation in Kv1.2 strongly attenuates sensitivity to use-dependent activation.
Engineering this mutation into glycosylation deficient [N207A] channels significantly reduces
the effects of the N207 mutation on the kinetics and voltage-dependence of channel
activation. Lastly, co-expression of glycosylation deficient channels with Slc7a5, a novel
regulator of Kv1.2, appears to occlude the use-dependent activation mechanism of Kv1.2.
This regulatory effect of Slc7a5 also affects the slow gating properties of Kv1.2[N207A]
18
channels, leading to significant acceleration of activation, along with a negative shift in
voltage-dependence of activation.
Conclusions: Slow activation kinetics and altered voltage-dependence of glycosylation
mutants are caused by increased sensitivity to use-dependent activation. These findings
suggest a previously unrecognized role for glycosylation as a regulator of Kv1.2 sensitivity to
extrinsic regulatory processes.
(Poster flash talk 5)
Temperature dependence of fluorinated-surfactant micellization
Jonas Höring1, Grégory Durand
2 & Sandro Keller
1
1Molecular Biophysics, University of Kaiserslautern, Kaiserslautern, Germany; 2Institut des Biomolécules Max Mousseron, University of Avignon, Avignon, France
Membrane proteins (MPs) are crucial for plentiful physiological processes of the cell. This
makes the comprehension of their structure paramount for, among others, the
pharmaceutical industry, as MPs represent more than 60 % of drug targets. However,
compared with soluble proteins, only a small number of high-resolution structures of MPs are
available. This is rooted in their embedment in the lipid bilayer, which hampers the
accessibility of MPs to biophysical investigation [1]. Therefore, it is essential to extract MPs
from their native membrane. While canonical detergents, due to their solubilizing nature, are
well-suited for the extraction of MPs, they are often invasive towards the structure of the MP.
Therefore, new approaches have emerged to combine detergent capability for membrane
extraction with a milder effect on MPs to form membrane-mimetic systems for the study of
MPs [2]. Fluorinated surfactants are one of the compounds which were designed for this
task. The bulky, fluorinated carbon tail of these molecules renders them both hydrophobic
and lipophobic, resulting in poor miscibility with hydrocarbon chains. Therefore, fluorinated
surfactants are less invasive towards extracted MPs compared with hydrocarbon-based
surfactants, rendering their micelles promising membrane-mimetic systems [3]. To gain
detailed insight into the molecular mechanisms underlying the mildness of fluorinated
surfactants toward MPs, we characterized their micellization behavior by several calorimetric
techniques. In particular, we studied the temperature-dependent properties of micellization of
a homologous series of fluorinated surfactants and their hydrogenated analogs. Isothermal
titration calorimetry (ITC) was utilized to investigate the temperature dependence of their
critical micellar concentration, as well as to allow a full thermodynamic characterization of the
micellization. Differential scanning calorimetry (DSC) was employed to further explore
micellization of the surfactants and the associated heat-capacity changes over a wide
temperature range. Data from both methods were compared to show differences between
corresponding critical micelle concentrations and critical micelle temperatures of the
surfactants.
References
[1] Y. Arinaminpathy et al., Drug Discovery Today 14, 23/24, 1130–1135 (2009)
[2] J.-L. Popot, Annual Review of Biochemistry 79, 737–775 (2010)
[3] G. Durand et al., in Membrane Proteins Production for Structural Analysis chapter 8, Ed.
I. Mus-Veteau, Springer, New York, USA, 205–251 (2014)
19
(Poster flash talk 6)
Identification of chloroplast envelope proteins with critical importance for cold
acclimation of Arabidopsis
Annalisa John , Oliver Trentmann & Ekkehard Neuhaus Department of Plant Physiology, University of Kaiserslautern, Kaiserslautern, Germany
Higher plants are organisms performing a sessile lifestyle and thereby are exposed to many
different weather and temperature conditions. Especially coldness shows a great effect on
plant vitality, mainly because of a downregulation of enzymatic reactions, a decreased fluidity
of membranes and the damage by ice crystals. This is why plants like Arabidopsis thaliana
developed different strategies to cope with low temperatures and become cold-hardy. A
common adaption while acclimation is the storage of different solutes like sugars in the
vacuole to function as osmotica and prevent dehydration. Next to the involvement of the
vacuole, cold acclimation is a more complex process including also other organelles like the
chloroplast. Not only photosynthesis takes place in here, the chloroplast functions as a
central coordinator of metabolic adjustment and acclimation processes. Proteome analysis of
chloroplast envelopes of cold treated Arabidopsis plants identified proteins with critical
importance for cold acclimation. Some of them are already known and analyzed, like the
plastidic ATP/ADP antiporter NTT2, the maltose translocator MEX1 or the fatty acid exporter
FAX1. But in this connection also new proteins were identified and thereby the most
interesting candidates like e. g. a small RAB-B-class GTPase, a putative steroid-5-alpha
reductase and a putative subfamily F ABC protein were selected because they showed the
highest alterations in relation to the protein content compared to the others. To gain a first
insight in the impact of these proteins for adaption to lower temperatures the phenotype of
corresponding Arabidopsis knock-out lines should be investigated, especially while cold
treatment. In future work the creation of overexpressing lines is planned as well as
subcellular localization studies.
(Poster flash talk 7)
Dissecting intracellular trafficking and mis-trafficking of human kidney AE1 in
yeast and mammalian cells
Hasib A. M. Sarder, Björn Becker & Manfred J. Schmitt Molecular & Cell Biology, Department of Biosciences, Center of Human and Molecular Biology (ZHMB), Saarland University, D-66123 Saarbrücken, Germany
Kidney anion exchanger 1 (kAE1) is a membrane protein located in the basolateral
membrane of α-intercalated cells (A-IC) of the human kidney which is responsible for the
reabsorption of bicarbonate ions (HCO3-) by exchanging with chloride ions (Cl-), thereby,
ensuring acid excretion in the urine [1]. Various genetically inherited mutations in the kAE1
encoding gene have been reported to negatively affect HCO3-/Cl- exchange and ultimately
result in clinical disorders known as distal renal tubular acidosis (dRTA). Until now,
autosomal dominant (AD) and recessive (AR) mutations have been identified [2], but the
underlying molecular mechanisms for the disease are still poorly understood. In this study,
we are using baker’s yeast (a simple eukaryotic model organism) and a mouse kidney cell
line (mIMCD3) to dissect the pathological reasons of dRTA. So far, a yeast codon-optimized
kAE1 variant could be successfully expressed as full-length protein in yeast. Proper kAE1
localization at the cell periphery was confirmed via indirect immunofluorescence microscopy,
cell surface biotinylation experiments as well as colocalization with the yeast plasma
membrane marker Pma1p. In vivo functionality of kAE1 was analyzed by using a pH-
20
sensitive ratiometric dye. Furthermore, during my research stay at the Cordat Lab in Canada,
I was focused on the characterization of novel dominant and recessive dRTA mutants
(R296H, Y413H and S525F) by using mIMCD3 cell line, the closest cellular model for A-IC at
the moment. All the mutants showed less stability in degradation assays as well as the p62
levels, a reporter protein for active autophagy, was likewise upregulated compared to the
wild-type situation. However, a cytotoxic effect after expression of these kAE1 mutants was
not observed under hyperosmotic condition.
(Poster flash talk 8)
Analysis of intracellular trafficking and localization of human kidney anion
exchanger 1 (kAE1) in yeast
Xiaobing Li, Björn Becker & Manfred J. Schmitt Molecular and Cell Biology, Department of Biosciences and Center of Human and Molecular Biology (ZHMB), Saarland University, D-66123 Saarbrücken, Germany
Human kidney anion exchanger 1 (kAE1) represents a bicarbonate transporter in the
basolateral membrane of renal epithelial cells that participates in the fine-tuning of acid-base
homeostasis by mediating electroneutral Cl-/HCO3- exchange. Several autosomal mutations
in the kAE1 encoding gene (SLC4A1) can cause clinical disorders known as distal renal
tubular acidosis (dRTA) which are linked to kAE1 misfolding, ER/Golgi retention, and/or
premature degradation. Despite that some proteins involved in kAE1 trafficking had been
identified, the precise mechanism(s) resulting in dRTA still remain unclear. In this study, we
are using yeast as experimental model system to identify proteins which affect intracellular
kAE1 trafficking to the plasma membrane and/or its turnover, both of which is vital for proper
kidney function. Our current data indicate that kAE1 can be successfully expressed in yeast
and partially colocalizes at the plasma membrane. However, kAE1 overexpression leads in
part to the intracellular kAE1 aggregation which subsequently induces a strong UPR
activation in yeast cells. In further experiments, we aimed to address the underlying
principles resulting in kAE1 aggregation and to identify tools to finally prevent or at least
minimize intracellular kAE1 protein accumulation in yeast. Furthermore, we will initially
establish a fluorescence-based screening approach in S. cerevisiae to test to which extent
the overexpression of single yeast genes can modulate both cellular expression and plasma
membrane localization of kAE1.
This study is kindly supported by the Deutsche Forschungsgemeinschaft (IRTG 1830).
(Poster flash talk 9)
Transmembrane protein 109 (TMEM109), a putative hsnd3 protein
Andrea Tirincsi, Sarah Haßdenteufel, Monika Lerner, Mark Sicking, Sven Lang & Richard Zimmermann Medical Biochemistry & Molecular Biology, Saarland University, Homburg
Protein transport to the endoplasmatic reticulum (ER) mostly relies on the signal recognition
particle (SRP) pathway. However, tail-anchored proteins can be transported into the ER in an
SRP independent (SND) manner. Previous work identified the SND pathway in S. cerevisiae
and we aim to identify it in human cells. The SND pathway has three components (hsnd1, 2,
3) so far no human hsnd3 protein has been identified. Here we report transmembrane protein
109 (TMEM109) as a putative hsnd3 protein, which might be involved in the SND targeting
machinery. TMEM109 also known as Mitsugumin23 (MG23) is a transmembrane protein
21
within the ER and in the nuclear membrane and has been described as a calcium channel
within the ER membrane.
Co-immunoprecipitation experiments showed an interaction between hsnd2 and TMEM109.
Further mass spectrometry analysis revealed the simultaneous reduction of TMEM109 with
hsnd2 depletion; this might suggest that they are associated within the snd pathway. Here
we report our initial results about TMEM109 as a putative hsnd3 protein and its role in
calcium homeostasis.
(Poster flash talk 10)
The warm and high light stress; insight on a plastidic sugar transporter
Pratiwi Prananingrum, Kathrin Patzke, Patrick A.W. Klemens, Oliver Trentmann, Ilka Haferkamp & H. Ekkehard Neuhaus Department of Plant Physiology, Faculty of Biology, University of Kaiserslautern, Germany
The major carbohydrates in plants are glucose, fructose, sucrose, cellulose and starch. The
transport of sugars across membrane barriers is essential for higher plants, since sugar
represents transport and storage units of cellular energy generation and thus play a
fundamental role during developmental processes and stress responses. In addition to
transport across the plasma membrane, carrier-mediated sugar transport has also been
demonstrated across organellar membranes, such as the inner plastid envelope or the
vacuolar membrane, named tonoplast. The monosaccharide transporter family is diverse. In
this study, we will focus on VGT-like protein family of monosaccharide transporter in which
comprised of three genes Arabidopsis thaliana Vacuolar Glucose Transporter 1 (AtVGT1),
AtVGT2, and AtVGT3. Recently, two genes from this family, AtVGT1 and AtVGT2 have been
shown to localize to the vacuolar membrane of Arabidopsis thaliana (A. thaliana) (Aluri and
Büttner, 2006). It has been shown that VGT1 transports glucose and a proton-coupled
antiport. In contrast, here we characterize that the protein encoded by AtVGT3 (we further
address it as plastidic sugar transporter /pSuT), locates to the chloroplast membrane.
Transport analyses with yeast cells expressing a truncated, vacuole-targeted version of pSuT
indicate that both glucose and Suc act as substrates, and nonaqueous fractionation supports
a role for pSuT in Suc export from the chloroplast. The latter process is required for a correct
transition from vegetative to reproductive growth and influences inflorescence architecture.
Moreover, pSuT play a role in the adaptation to the warm, cold, and high light stress. These
data further underline the central function of the chloroplast for plant development and the
modulation of stress tolerance.
(Poster flash talk 11)
Vacuolar sucrose mobilization is critical for the development of Arabidopsis
thaliana
Duc Phuong Vu & Ekkehard Neuhaus Department of Plant Physiology, University of Kaiserslautern, Germany
Sugars are involved as signal molecules in plant development. To achieve these functions,
cellular sugar homeostasis must be tightly regulated, via control of membrane transporters
and enzyme activities. Among these enzymes are the vacuolar invertases, which cleave
sucrose into glucose and fructose.
To study the impact of an altered sugar homeostasis on Arabidopsis thaliana development
by increasing the luminal sucrose concentration, we generated amiRNA lines targeting both
22
vacuolar invertases. The mutants showed increased sucrose concentrations and decreased
monosaccharides levels. The early plant development is delayed, most probably because of
a blocked sugar mobilization. In addition, plant seed yield is decreased and impaired sugar
export from source to sink tissues might contribute to this.
Hence, vacuolar sucrose mobilization is of superior importance for modulation of plant
development and seed weight.
(Poster flash talk 12)
Studying the influence of selenium on arsenic hepatobiliary transport
Janet R. Zhou1,2
, Gurnit Kaur1,2
, Denis Arutyunov2,3
& Elaine M. Leslie1,2,3
1. Division of Analytical and Environmental Toxicology, Department of Laboratory Medicine and Pathology, University of Alberta, Canada; 2. Membrane Protein Disease Research Group, University of Alberta, Canada; 3. Department of Physiology, University of Alberta, Canada
Background: Worldwide, at least 200 million people are exposed to the proven human
carcinogen arsenic at levels exceeding the World Health Organization guideline (10 µg/L). In
animal models arsenic and selenium are mutually protective via the formation and biliary
excretion of the seleno-bis (S-glutathionyl) arsinium ion [SeAs(GS)2]-. Despite ongoing
human selenium supplementation trials in arsenic endemic regions, the influence of selenium
on human hepatic handling of arsenic is not adequately understood. We hypothesized that
selenium would increase the biliary excretion of arsenite (AsIII) from human HepaRG cells, an
immortalized cell lined used as a surrogate for primary human hepatocytes.
Objective: To study the influence of selenite (SeIV), selenide (SeII), methylselenocysteine
(MSC) and selenomethionine (SM) on arsenic efflux from HepaRG cells.
Methods: Fluorescence microscopy after treatment with 5(6)-carboxy,2’,7’-
dichlorofluorescein (CDF) diacetate was performed to visualize the canalicular networks and
assess function of multidrug resistance protein 2 (MRP2/ABCC2), which transports
conjugated and unconjugated organic anions, including [SeAs(GS)2]- into bile. The
expression of genes involved in hepatic arsenic metabolism (As3MT) and efflux transport
(ABCC2 and ABCC4) were assessed with an agarose gel. Crude membrane preparations
subjected to SDS-PAGE and immunoblots were used to determine if MRP2 and the related
arsenic sinusoidal transporter multidrug resistance protein 4 (MRP4) were present at the
protein level. HepaRG cells were treated with 73AsIII ± SeIV, SeII, MSC or SM (1 µM) and
efflux was measured across sinusoidal and canalicular membranes. Biliary excretion indices
(BEIs) were calculated to quantify biliary excretion.
Results: CDF accumulated in canalicular networks, suggesting the presence of functional
MRP2. ABCC2, ABCC4, and As3MT are expressed in the cell line, and MRP2 and MRP4
proteins were detected by immunoblot. SeII increased biliary excretion of 73AsIII, with a BEI of
24%, but other forms of selenium did not.
Conclusion: This work will lead to a better understanding of the influence of selenium on
arsenic handling by human liver and provide valuable information for ongoing selenium-
supplementation trials.
23
Characterization of a novel splice variant of the stromal interaction molecule 1
(STIM 1)
Mona Schöppe1, Kathrin Förderer
1, Yvonne Schwarz
3, Annette Lis
2 & Barbara Niemeyer
1
1Molecular Biophysics, Saarland University, 66421 Homburg, Germany/
2Biophysics, Saarland University, 66421 Homburg,
Germany/ 3Molecular Neurophysiology , Saarland University, 66421 Homburg, Germany
Changes in intracellular free calcium concentration [Ca2+] probably represent the most
widespread and important signaling event in cellular physiology, since transient elevations of
Ca2+ directly or indirectly control and regulate a plethora of cellular responses. Therefore
cells must be able to react to minor changes in [Ca2+]i and changes must be tightly regulated.
The major Ca2+ pathway in electrically nonexcitable cells is the store operated calcium entry
(SOCE) via calcium release activated calcium channels. The Ca2+ selective channel is
located in the plasma membrane and formed by Orai-family proteins. Stromal interaction
molecule (STIM1 and STIM2) proteins activate SOCE by sensing changes in the luminal
Ca2+ concentration in the endoplasmic reticulum via their N-terminal EF hand motif. Upon
store depletion, STIM molecules change conformation, multimerize and trigger SOCE by
directly gating Orai channels within ER-PM junctional regions.
Here, we report the identification and characterization of a novel STIM1 splice variant,
STIM1A, which retains an additional 31 amino acid long exon within its C-terminal cytosolic
region. The so called exon A is spliced into the mRNA downstream of the channel activating
region and also downstream of a region encoding an acidic inhibitory domain (ID) that
mediates fast Ca2+ inactivation of Orai1. On mRNA level the variant is ubiquitously
expressed, but its abundance relative to the more common STIM1 variant varies upon cell
type. In contrast to the RNA analysis, STIM1A could be detected only in murine testis on
Western blots. Transient overexpression of the splice variant leads to an overall reduced
SOCE and ICRAC when compared to STIM1, whereas a knockdown leads to an increased
SOCE. The new splice variant colocalizes with the wildtype STIM1 upon co-overexpression.
Furthermore, the interaction with Orai1 is not impaired. Using mutation assays, two specific
amino acids within the exon A could be identified to be responsible for the Ca2+ phenotype
of the splice variant. Future experiments aim to understand the physiological role of STIM1A
in Testis and to identify splice-specific interaction partners.
Supported by IRTG 1830 and SFB 894
Phosphorus source and intestinal absorption: inorganic phosphate is ab-
sorbed by the paracellular pathway
Matthew Saurette1,2
, Tate MacDonald1,2
& R. Todd Alexander1,2,3
1. Department of Physiology, Membrane Protein Disease Research Group, University of Alberta, Edmonton, Alberta, T6G 2H7, Canada. 2. The Women’s and Children’s Health Research Institute, Edmonton, Alberta, T6G 1C9, Canada. 3. Department of Pediatrics, University of Alberta, Edmonton, Alberta, T6G 2R7.
Hyperphosphatemia, elevated serum phosphate (Pi), is an electrolyte imbalance frequently
occurring with chronic kidney disease (CKD) and end stage renal disease (ESRD).
Hyperphosphatemia is associated with a number of negative sequelae including decreased
renal function, cardiovascular disease and death. Consequently, lowering serum is a clinical
priority for patients with CKD and ESRD. One of the main ways to reduce serum Pi is
reducing the amount of Pi ingested. There are two main forms of Pi in our diet: organic and
inorganic. Protein is one of the main sources of organic Pi and, therefore, reducing dietary Pi
often requires reducing protein consumption which can lead to protein malnutrition. Inorganic
Pi is a common food additive and preservative, and are ubiquitous in the “Western” diet.
Compared to organic Pi, inorganic Pi is more bioavailable, however, it is unclear why. We
24
hypothesized that inorganic Pi is more bioavailable than organic Pi because it does not
require digestion therefore is readily absorbed down the intestinal electrochemical gradient
(N.B. the intestinal lumen is – 5 mV relative to blood).
To test our hypothesis, wild-type and claudin-12 null mice were consecutively placed on an
organic then predominantly inorganic Pi diet in metabolic cages, permitting urine and feces
collection. The total amount of Pi in each diet was identical (0.7%). Claudin-12 is a tight
junction protein that we propose blocks paracellular Pi flux. Urinary and fecal Pi excretion,
and Pi bioavailability were determined for mice fed each diet to assess differences between
organic and inorganic Pi absorption. Both wild-type and claudin-12 null mice had increased
oral Pi bioavailability and urine Pi excretion on the inorganic Pi diet, consistent with
enhanced intestinal absorption. In contrast, potassium, another ion absorbed by the
paracellular route in the gut, did not display differences in oral bioavailability or urinary
excretion between the different diets. Further, claudin-12 null mice had significantly greater
oral Pi bioavailability, serum Pi and urine Pi excretion relative to wild-type mice on the
inorganic but not organic Pi diet. Our data confirms that inorganic Pi is more bioavailable
than organic Pi and is consistent with i) claudin-12 forming a paracellular Pi barrier that
inhibits inorganic but not organic Pi absorption and ii) organic Pi being absorbed through the
paracellular pathway. Ultimately this works provides biological rationale for CKD/ESRD
patients to consume diets low in inorganic phosphate and provides a molecular target to help
block intestinal Pi absorption.
To BEet or not to BEet. A short story about sugar beet transporters and their
impact on cold acclimation
Cristina Martins Rodrigues & Ekkehard Neuhaus Department of Plant Physiology, University of Kaiserslautern, Germany
Sugar beet (Beta vulgaris) is the exclusive sugar source for the food industry in temperate
climate zones (Europe and North America). Sugar beet taproots accumulate sucrose to as
much as 20% of their fresh weight at maturity. Although a biennial plant species, sugar beet
is grown annually due to its prominent sensitivity to freezing. A protective mechanism of
higher plants against low temperatures is the accumulation of soluble solutes, especially
sugars. The accurate compartmentalization of sugars into the vacuole, the cellular sugar
storage organelle, is guided by several sugar carriers. To understand how an altered
compartmentalization of the distinct sugar species might influence the cold response of
plants, we investigated the behavior of different Arabidopsis thaliana lines each
overexpressing one of the mentioned sugar carriers deriving from sugar beet. Therefore, we
analyzed the metabolic status and expression pattern during the onset of cold.
Investigation of RBC aging
Katie Badior Department of Physiology, University of Alberta, Edmonton, Canada
During their circulating lifetime of 120 days, RBCs are exposed to extreme physical and
chemical stresses. As damages accumulate, aged RBC transport function becomes less
efficient, and they must be removed from circulation. Aged RBCs are removed by white
blood cells of the immune system.
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White blood cells recognize aged RBCs through antibody mediators, which I turn recognize
the major membrane protein component on RBCs, Band 3. Previous studies indicate that
there are two regions of Band 3 (epitopes) in particular that are recognized by these
antibodies, comprising amino acid residues 538-553, and 813-830. Recently, the 813-830
region was shown to be inside the red cell, where it would be inaccessible to antibodies in
blood sera. We propose that the Band 3 epitope marking RBCs as ‘aged’ to antibodies is
dynamic, and can access both the intracellular and extracellular sides of RBC membranes.
The ability of these Band 3 senescence epitopes to access the extracellular environment was
assessed using substituted cysteine scanning mutagenesis and antibody binding assays,
using an HEK293 cell model.
In our model of RBC aging red cell viability is damaged over time following antibody binding
to Band 3. This has implications for developing strategies improving RBC storage and shelf
life, as well as increasing transfusion efficiency.
Characterization of flower containing vesicles in mouse cytotoxic T lympho-
cytes
Keerthana Ravichandran, Claudia Schirra & Jens Rettig. Center for Integrative Physiology and Molecular Medicine (CIPMM), Saarland University, Homburg/Saar, Germany
A transmembrane protein, flower, was shown to be associated in synaptic vesicle
endocytosis in neurons (Yao et al., 2009) and our group recently have shown that flower
consists of four transmembrane domains with its N- and C-terminus facing the cytoplasm.
We further demonstrated that it plays an important role in regulating cytotoxic granule (CG)
endocytosis in primary cytotoxic T lymphocytes (CTLs), which was delayed in cells from
flower KO mice. Despite these findings, it is still not clear how and from where flower exerts
the action of controlling endocytosis. Endocytosis as such in CTLs is important for function in
serial killing of target cells. The endogenous expression of flower by immunocytochemistry
was found to be very low but found mostly in vesicles and partially at the plasma membrane.
Real time killing assay with different effector: target cell ratios show a clear defect in multiple
killing efficacy of flower KO CTLs. To further understand the endogenous localization of the
protein, flower-Halo-HA knock-in mouse was generated using CRISPR-CAS9 gene editing
technology and the fusion protein was characterized using specific antibodies. The major aim
of this study is to find the localization of the protein flower and characterize the proteome of
flower containing vesicles and subsequently decipher a possible mechanism of how flower
aids the CGs in endocytic process.
Slack K+ channels and the tight junction protein Claudin-12 in the cochlea
Pauline Schepsky & Jutta Engel Biophysics, Saarland University, Homburg, Germany
Slack (Slo2.2, gene Kcnt1) is a Na+- and voltage-activated potassium channel that reduces
neuronal excitability in response to neuronal activation and Na+ influx. Slack mRNA and
Slack currents have been described in spiral ganglion neurons (SGN; Reijntjes et al.,
Scientific Reports 2019). The role of Slack in hearing however is unknown. I have
characterized hearing of 8 week-old Slack-/- mice by recording auditory brainstem responses
(ABR). Slack-/- mice had normal hearing thresholds. We hypothesized that Slack counter-acts
neuronal excitation and may reduce noise-induced cochlear damage. Therefore Slack-/- and
wild type mice were subjected to a mild noise trauma (100 dB SPL, 2 hrs, 8 – 16 kHz band
26
noise). The shift in hearing thresholds was assessed directly after trauma and between one
and 28 days after trauma. Thereafter, cochleae were analyzed for numbers and localization
of presynaptic ribbons and postsynaptic protein clusters.
Tight junctions are essential for cellular compartmentalization transforming tissues to highly
complex functional systems. In the cochlea, various claudins that are important components
of tight junctions form the paracellular barrier between the endolymph and perilymph
compartments and thereby maintain the large concentration differences of K+, Na+ and Ca2+
as well as the endocochlear potential needed for normal cochlear function.
Preliminary results point to mRNA expression of claudin-12 (Cldn12) in the cochlea but the
cellular expression pattern had not been identified. During my stay in Dr. Alexander’s
laboratory in Edmonton I analyzed the cellular expression of claudin-12 in the cochlea of
adult mice. By using claudin-12 knockout mice carrying a LacZ-reporter construct I found
strong promoter activity of claudin-12 exclusively in inner hair cells.
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List of Participants
28
Alexander, Todd Department of Pediatrics Division of Nephrology & Physiology 2B2.42 Walter Mackenzie Centre University of Alberta Edmonton, Alberta, Canada, T6G 2R7 [email protected]
Amoroso, Gabriele Department of Plant Physiology Faculty of Biology University of Kaiserslautern Erwin-Schrödinger-Straße 67663 Kaiserslautern Germany [email protected]
Badior, Katie Department of Biochemistry Faculty of Medicine & Dentistry University of Alberta 4020E Katz Group Rexall Building Edmonton, Alberta, Canada, T6G 2E1 [email protected]
Bak, Jessi Department of Biochemistry Faculty of Medicine & Dentistry University of Alberta 327 Medical Sciences Building Edmonton, Alberta, CanadaT6G 2H7 [email protected]
Beggs, Megan Department of Pediatrics Division of Nephrology & Physiology 2B2.42 Walter Mackenzie Centre University of Alberta Edmonton, Alberta, Canada, T6G 2R7 [email protected]
Bellin, Leo Department of Plant Physiology Faculty of Biology University of Kaiserslautern Erwin-Schrödinger-Straße 22 67663 Kaiserslautern Germany [email protected]
Bentrcia, Teqiyya Department of Experimental & Clinical Pharmacology & Toxicology Faculty of Medicine Saarland University University Hospital Building 46 66421 Homburg Germany [email protected]
Brassard, Raelynn Department of Biochemistry School of Translational Medicine Faculty of Medicine & Dentistry University of Alberta 451 Medical Sciences Building Edmonton, Alberta, Canada T6G 2H7 [email protected]
Bugaeva, Wassilina Department of Plant Biology Faculty of Natural Sciences & Technology Saarland University University Hospital, Building A 2.4 66421 Homburg /Saar Germany [email protected]
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Casey, Joe Department of Biochemistry Faculty of Medicine & Dentistry University of Alberta 4020 E Katz Group Rexall Building Edmonton, Alberta, Canada, T6G 2E1 [email protected]
Chen, Xing-Zhen Department of Physiology Faculty of Medicine & Dentistry University of Alberta 7-29A Medical Sciences Building Edmonton, Alberta, Canada, T6G 2H7 [email protected]
Cordat, Emmanuelle Department of Physiology School of Moleclar & Systems Medicine University of Alberta Medical Sciences Building/Room 7-34 Edmonton, Alberta, Canada, T6G 2H7 [email protected]
Das, Damayantee Department of Pharmacology Faculty of Medicine & Dentistry University of Alberta 6-040 Li Ka Shing Centre Edmonton, Alberta, Canada [email protected]
Deitmer, Joachim Department of General Zoology Faculty of Biology University of Kaiserslautern Erwin-Schrödinger-Straße 13 67663 Kaiserslautern Germany [email protected]
Engel, Jutta Department of Biophysics Faculty of Medicine Saarland University CIPMM , Building 48 66421 Homburg/Saar Germany [email protected]
Fajonyomi, Daniel Department of Pharmacology Faculty of Medicine & Dentistry University of Alberta 6-040 Li Ka Shing Centre Edmonton, Alberta, Canada [email protected]
Flockerzi, Veit Department of Experimental & Clinical Pharmacology & Toxicology Faculty of Medicine Saarland University University Hospital, Building 46 66424 Homburg/Saar Germany [email protected]
Herrmann, Johannes Department of Cellular Biology Faculty of Biology University of Kaiserslautern Erwin-Schrödinger-Straße 13 67663 Kaiserslautern Germany [email protected]
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Höring, Jonas Department of Molecular Biophysics Faculty of Biology University of Kaiserslautern Erwin-Schrödinger-Straße 13 67663 Kaiserslautern Germany [email protected]
John, Annalisa Department of Plant Physiology Faculty of Biology University of Kaiserslautern Erwin-Schrödinger-Straße 22 67663 Kaiserslautern Germany [email protected]
Keller, Sandro Department of Molecular Biophysics Faculty of Biology University of Kaiserslautern Erwin-Schrödinger-Straße 13 67663 Kaiserslautern Germany [email protected]
Khan, Azkia Department of Plant Physiology Faculty of Biology University of Kaiserslautern Erwin-Schrödinger-Straße 22 67663 Kaiserslautern Germany [email protected]
Khandpur, Gurleen Kaur Department of Biochemistry Faculty of Medicine Saarland University Campus, Building B2.2 66123 Saarbrücken Germany [email protected]
Laborenz, Janina Department of Cellular Biology Faculty of Biology University of Kaiserslautern Erwin-Schrödinger-Straße 13 67663 Kaiserslautern Germany [email protected]
Lang, Sven Department of Medical Biochemistry & Molecular Biology Faculty of Medicine Saarland University University Hospital, Building 44 66424 Homburg Germany [email protected]
Lemieux, Joanne Department of Biochemistry School of Translational Medicine Faculty of Medicine & Dentistry University of Alberta 451 Medical Sciences Building Edmonton, Alberta, Canada T6G 2H7 [email protected]
31
Li, Xiaobing Department of Molecular & Cell Biology Faculty of Natural Sciences & Technology III Saarland University Campus Saarbrücken, Building A 1.5 P.O. Box 151150 66041 Saarbrücken Germany [email protected]
Liu, Xiong Department of Physiology Faculty of Medicine & Dentistry University of Alberta 7-29A Medical Sciences Building Edmonton, Alberta Canada T6G 2H7 [email protected]
Martins Rodrigues, Cristina Department of Plant Physiology Faculty of Biology University of Kaiserslautern Erwin-Schrödinger-Straße 22 67663 Kaiserslautern Germany [email protected]
Möhlmann, Torsten Department of Plant Physiology Faculty of Biology University of Kaiserslautern Erwin-Schrödinger-Straße 22 67663 Kaiserslautern Germany [email protected]
Morgan, Bruce Department of Biochemistry Faculty of Medicine Saarland University Campus, Building B2.2 66123 Saarbrücken Germany [email protected]
Neuhaus, Ekkehard Department of Plant Physiology Faculty of Biology University of Kaiserslautern Erwin-Schrödinger-Straße 22 67663 Kaiserslautern Germany [email protected]
Niemeyer, Barbara Department of Biophysics Faculty of Medicine Saarland University University Hospital, Building 58 66424 Homburg Germany [email protected]
Oestreicher, Julian Department of Biochemistry Faculty of Medicine Saarland University Campus, Building B2.2 66123 Saarbrücken Germany [email protected]
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Philippar, Katrin Department of Plant Biology Faculty of Natural Sciences & Technology Saarland University University Hospital, Building A 2.4 66421 Homburg /Saar Germany [email protected]
Prananingrum, Pratiwi Department of Plant Physiology Faculty of Biology University of Kaiserslautern Erwin-Schrödinger-Straße 22 67663 Kaiserslautern Germany [email protected]
Ravichandran, Keerthana Department of Physiology Faculty of Medicine Saarland University CIPMM, Building 48 D-66421 Homburg Germany [email protected]
Rettig, Jens Department of Physiology Faculty of Medicine Saarland University CIPMM, Building 48 D-66421 Homburg Germany [email protected]
Russo, Antonietta Department of Medical Biochemistry & Molecular Biology Faculty of Medicine Saarland University University Hospital, Building 44 66424 Homburg Germany [email protected]
Sarder, Hasib Department of Molecular & Cell Biology Faculty of Natural Sciences & Technology III Saarland University Campus Saarbrücken, Building A 1.5 P.O. Box 151150 66041 Saarbrücken Germany [email protected]
Saurette, Matthew Department of Pediatrics Division of Nephrology & Physiology University of Alberta 2B2.42 Walter Mackenzie Centre Edmonton, Alberta, Canada, T6G 2R7 [email protected]
Schepsky, Pauline Department of Biophysics Faculty of Medicine Saarland University CIPMM , Building 48 66421 Homburg/Saar Germany [email protected]
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Schmitt, Manfred Department of Molecular & Cell Biology Faculty of Natural Sciences & Technology III Saarland University Campus Saarbrücken, Building A 1.5 P.O. Box 151150 66041 Saarbrücken Germany [email protected]
Schöppe, Mona Department of Biophysics Faculty of Medicine Saarland University University Hospital, Building 58 66424 Homburg Germany [email protected]
Sicking, Mark Department of Medical Biochemistry & Molecular Biology Faculty of Medicine Saarland University Building 45.2 D-66421 Homburg/Saar Germany [email protected]
Tirincsi, Andrea Department of Medical Biochemistry & Molecular Biology Faculty of Medicine Saarland University University Hospital, Building 44 66424 Homburg Germany [email protected]
Touret, Nicolas Department of Biochemistry School of Translational Medicine Faculty of Medicine & Dentistry University of Alberta 4-020H Katz Group-Rexall Centre for Pharmacy & Health Research Edmonton, Alberta, Canada T6G 2E1 [email protected]
Van der Laan, Martin Department of Medical Biochemistry & Molecular Biology Faculty of Medicine Saarland University Building 45.2 D-66421 Homburg/Saar Germany [email protected]
Vu, Duc Phuong Department of Plant Physiology Faculty of Biology University of Kaiserslautern Erwin-Schrödinger-Straße 22 67663 Kaiserslautern Germany [email protected]
Wollweber, Florian
Department of Medical Biochemistry & Molecular Biology Faculty of Medicine Saarland University Building 45.2 D-66421 Homburg/Saar Germany [email protected]
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Yadao, Nilam Department of Medical Biochemistry & Molecular Biology Faculty of Medicine Saarland University Building 45.2 D-66421 Homburg/Saar Germany [email protected]
Zhou, Janet Department of Physiology School of Translational Medicine University of Alberta 7-10A Medical Sciences Building Edmonton, Alberta Canada T6G 2S2 [email protected]
Zimmermann, Richard Department of Medical Biochemistry & Molecular Biology Faculty of Medicine Saarland University University Hospital, Building 44 66424 Homburg Germany [email protected]
Zöller, Eva Department of Cellular Biology Faculty of Biology University of Kaiserslautern Erwin-Schrödinger-Straße 13 67663 Kaiserslautern Germany [email protected]
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Hints
Location
Parkhotel Weiskirchen
Kurparkstraße 4
66709 Weiskirchen
Phone: +49 (0)6876-919-0
Fax: +49 (0)6876-919-519
E-Mail: [email protected]
WWW: http://www.parkhotel-weiskirchen.de/
Conference phone
0151 18234824 (Gabi)
Taxi-Service
1. Taxi Martin (Wadern), 06871-2284
2. Taxi Martin (Weiskirchen), 06876-700750 ([email protected])
3. Taxi Fries, 06874-1392
4. Taxi Göbel, 06876-500 Airport-Shuttle Fahrservice Rosenberger (Lauterecken), 0170-8934702 Airport-Runner (Höringen), 0173-9112423
Venue
1. By train
From Homburg, Kaiserslautern or Saarbrücken by train to Merzig and then by bus
(Regionalbus R1, direction: Wadern) from Merzig to Weiskirchen (Kirche); total travel time
1.40 h to 2.10 h; for suitable train connections, please use the homepage of the “Deutsche
Bahn” (https://www.bahn.de/p/view/index.shtml)
2. By car
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