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Essentials of Glycobiology
Lecture 29
May 18, 2004
Plant Glycans
Marilynn EtzlerSection of Molecular and Cellular BiologyUniversity of CaliforniaDavis, CA 95616e-mail: [email protected]
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LECTURE OUTLINE
• Introduction
• Classes of glycans in plants
• Structure and biosynthesis of plant N-linked glycans
• Other types of plant glycans
• Plant cell wall
• Plant glycosylation mutants
• Molecular “farming”
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PLANTS FUNGI ANIMALS
Urochordates
Yeasts
Red algae Green algae
Brown algae
Vertebrates
Chordates
MossesLiverworts
Ferns
GymnospermsAngiosperms
PR
OT
OZ
OA
EU
KA
RY
OT
ES
Mu
ltic
ellu
lar
Un
icel
lula
r
Ancestral Prokaryotes
Arthropods
Insects
Slime moldsSponges
Coelenterates
MollusksNematodes
Echinoderms
Adapted from Figure 1-38, Molecular Biology of the Cell, 3rd ed., Garland Publishing, Inc.
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Comparison of Classes of Plant and Animal Glycans
From Lecture 2 by Dr. Varki
Cell wall glycans
Glycolipids
MajorGlycan
Classes inPlantCells
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Major Classes of N-Glycans Found in Plants
High Mannose
3
2
4
Complex
Pauci-mannoseHybrid= Sialic acid
= Man
= Gal
= GlcNAc
= Fuc
= Xylose
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Sialic Acid Has Recently Been Found in Plants
Evidence:
Found in glycoproteins obtained from suspension-cultured cells from Arabidopsis thaliana, Nicotiana tabacum and Medicago sativa
Bound to Sambucus nigra and Maackia amurensis lectinsDid not bind to these lectins if pretreated with 2-3,6 sialidase
Sialic acids were released chemically, derivatized with DMB and analyzed by reverse phase chromatography, yielding a prominent peak of Neu5Gc and a smaller peak of Neu5Ac. Similar results were obtained with sialic acids released enzymatically.
Analyses of DMB-SA derivatives were confirmed by MALDI-TOF
Reference: Shah, M.M., K. Fujiyama, C.R. Flynn, and L. Joshi (2003) Nature Biotechnology 21: 1470 – 1471.
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Recognition and Processing of N-Glycans in the Plant Secretory Pathway
Endoplasmic reticulum:
Glucosidases I and IICalreticulinER mannosidase
-mannosidase I
Golgi:
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Processing in Golgi (continued)
GNT I-Man IIGNT II
2-XylT3-FucT
3 2
3-GalT4-FucT 4
4
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J. Cell Science (2002) 115: 2423
Most of the volume of a typical plant cell is occupied by the vacuole(s)
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Processing in Vacuole or Enroute to Vacuole:
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Other Types of Plant Glycans
Plant Glycolipids:Galactolipids – in chloroplast membranes
Monogalactosyldiacylglycerol
Digalactosyldiacylglycerol
O-Diacylglycerol
O-Diacylglycerol6
Sphingolipids – in plasma membraneGlucosylceramide ceramide
Glycosylphosphatidylinositol anchors:
462
NH2
Protein - Ethanol amine - PO4
Phosphatidylinositol
4 462
NH2
Protein - Ethanol amine - PO4
Phosphatidylinositol
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Types of O-Linked Glycans Found in Plants
GlcNAc
Ser/Thr
GalNAc Gal GlcNAc Gal (Ara)1-4
Hydroxyproline
(Glycosylation of hydroxyprolineis unique to plants and Chlorophyceanalgae)
Arabinogalactan proteins (carbohydrate usually > 90% by weight)
3 33 333)(
(many variations)
- Hyp
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Cell Wall Glycans
Cellulose [ Glc 4 Glc]n
Pectins:
Homogalacturanan [GalU 4 GalU 4]n
Rhamnogalacturonan I [GalU 2 L-Rha 4]n
Rhamnogalacturonan II
Figure 3 from Phytochem. 57: 929 (2001)
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Xyloglucan:
4 44 4 44
2
6 6 6 6 6 6
2
nn
Hemicelluloses:
Cell Wall Glycans (continued)
Galactomannan
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Plant Cell Wall
Alberts, et al., Molecular Biology of the Cell, Fig. 19-75
Constitutes the extracellular matrix
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cellulose
galactans
homogalacturonan
rhamnogalacturonan I
calcium
rhamnogalacturonan II
xyloglucan
arabinans
Plasma membrane
Adapted from Figure 2, Trends in Plant Science 9: 203 (2004)
Model of Plant CellWall
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Plant Glycosyltransferases and Glycosidases
Almost 800 glycosyltransferase and glycosidase-related genes have been found in the Arabidopsis genome. Comprises > 3.3% of its genes.
By contrast, human genome has about 350 glycosyltransferase and glyco- sidase-related genes.
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Arabidopsis thaliana as a Model Plant System
SOME ADVANTAGES:
Complete genome sequencedDiploidEasily transformedRelatively rapid life cycleMany mutants availablePlants small and thus take up little space
Arabidopsis seed
Corn kernel
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ARABIDOPSIS cgl MUTANT
Identified by screening leaf extracts of EMS mutants with antiserum against complex glycans.
DEFECT: Missing GNT I
PHENOTYPE: No apparent effect on development and morphology of plants. No complex glycans. Accumulates Man5GlcNAc2
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ARABIDOPSIS mur 1 MUTANT
DEFECT: Deficient in an isoform of GDP-D-mannose-4,6-dehydratase.
PHENOTYPE: Plants are dwarfed and have fragile cell walls. Deficient in fucose.
Identified by making acid hydrolysates of cell walls of EMS mutants and screening the alditol acetate derivatives by GLC.
3 2
L-Gal
4 44 4 44
2
6 6 6 6 6 6
2
nn
L-Gal
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ARABIDOPSIS SPY MUTANTS
DEFECT: Deficient in O-linked GlcNAc transferase activity.
PHENOTYPE: A variety of alterations in growth and development.
Originally identified in genetic screen for mutants with increased response to gibberellins from T- DNA mutants. Also from EMS mutants.
Proposed to be involved in various aspects of regulation of plant development.