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ESSENTIALS OF GLYCOBIOLOGY
Lecture 30
May 18, 2004
Marilynn Etzler
Section of Molecular and Cellular Biology
University of California
Davis, CA 95616
([email protected])
FREE GLYCANS AND THEIR ROLES AS
SIGNALING MOLECULES
LECTURE OUTLINE
• Background
• Oligosaccharide signals trigger the initiation of
the plant defense response
• Nod factor signals initiate the nitrogen-fixing
Rhizobium-legume symbiosis
• Chitin oligosaccharide signals in plant defense
and early animal development
• Other oligosaccharide signals in early plant and animal
development
• Pattern recognition receptors and innate immunity
•Background:
Potential of oligosaccharides as signals:
1) Variety of linkages between monomers enables a large number
of conformational variations.
Portion of Table shown in Lecture 1:
Macromolecule
Building block
Protein
Nucleic Acid
Carbohydrate
Amino acids
Nucleotides
Hexoses
Possible variations
in a trimer
6
6
1,056 to 27,648
2) Many hydroxyls available for modification.
Basic elements of signaling system:
Signal
Receptor
Transduction
mechanism
Response
• Oligosaccharide Signals Trigger the Initiation of the Plant
Defense Response
Plant Defense Responses:
Pathogen
Pathogen
Elicitor
Elicitor
Cell wall
Plasma membrane
Early Plant Responses
Changes in ion fluxes
Oxidative burst  H2O2 and O2
Activation of early defense-related genes
Phytoalexin production
Structural changes in cell walls
The first oligosaccharide signal was identified in the fungus,
Phytophthora megasperma, a fungal pathogen of soybean.
Isolated from cell walls
Elicits phytoalexin production in soybean seedlings
Structure confirmed by chemical synthesis
3
3
Hepta--glucoside
6
6
6
6
Relative Activities of Oligo--Glucosides
3
6
Relative
Elicitor Activity
3
6
6
Relative
Binding Activity
1000
1000
270
93
6
3
3
6
3
6
3
6
6
3
1.2
6
6
6
6
= reduced glucose
1.3
• Nod factor signals initiate the nitrogen-fixing
Rhizobium-legume symbiosis
Rhizobia
CH2O R 5
O
R O
R O
R4'
NR
1
HO
R2
O
NH
CO
CH3
R5'
R7
R5
3
O
O
4
R3'
OR 6
CH2
O
CH2OH
O
HO
n
NH
CO
O R 7
CH3
NOD Factor
R3
O
Flavonoids
Root hair
deformation
Infection thread
formation
GENERIC STRUCTURE OF NOD FACTORS
R5
R6
O
O
CH 2
R4 O
R3
CH 2OH
O
O
R2
O
HO
N
R1
CH 2
O
O
HO
NH
C=O
CH 3
n
R1 = H, Methyl
R5 = H, Ac
R2 = C16:2, C16:3
C18:1, C18:3, C18:4
C20:3, C20:4
R6 = H, Ac,
SO4
Fuc
AcFuc
MeFuc
R3 = H, Cb
R4 = H, Cb
O
O
R7
NH
C=O
CH 3
R7 = H
Glycerol
n=1-4
• Chitin oligosaccharide signals in plant defense
and early animal development
Elicit alkalinization of medium of tomato cell cultures
Biological activity
(nanomolar)
0.1
4
4
4
4
0.1
4
4
4
50
4
4
4
100,000
>1,000,000
Evidence for chitin oligosaccharide signaling in early animal
development:
In Xenopus laevis the developmentally regulated protein, DG42,
is homologous to Nod C, the enzyme that synthesizes the
chitin backbone of the Nod factors in rhizobia.
DG42 is only expressed between the gastrula and neurulaation
stages.
DG42 can direct the synthesis of chitin oligosaccharides in vitro.
Chitin oligosaccharides can be synthesized by extracts of
gastrulation stage embryos of cyprinid fishes (zebra fish and
carp).
Microinjection of fertilized eggs with antibodies against DG42
leads to severe defects in trunk and tail development.
• Other oligosaccharide signals in early plant and animal
development
Oligogalacturonides - isolated from plant cell walls
4
4
4
4
4
4
4
4
4
Serve as signals in both defense and in plant development
DEFENSE: Usually need DP of 10-14 to elicit phytoalexin accumulation.
DEVELOPMENT: Different DP elicit different responses.
= Galacturonic acid
DP = degree of polymerization
Xyloglucan nonasaccharide:
2
2
6
4
6
6
4
4
Concentrations of 10-8 M inhibit auxin-induced elongation of
stem fragments.
Hyaluronan fragments:
3
4
3
4
n
3
Activate antigen presenting cells and other proinflammatory
responses, signal cell motility and adhesion
• Innate immunity
Dependent on proteins and phagocytic cells that recognize conserved features of
pathogens that are absent in the host.
Found in vertebrates, invertebrates and plants.
Pattern recognition receptors
Recognize pathogen-associated immunostimulants
Examples of repeating patterns that often occur on pathogen surfaces
Chitin, glucan and other polysaccharides in cell walls of fungi
Peptidoglycan cell wall and flagella of bacteria
Lipopolysaccharide on Gram-negative bacteria
Teichoic acids on Gram-positive bacteria
Such repeating patterns called PAMPS (Pathogen-Associated
Molecular Patterns)
Pathogen
associated
immunostimulants
Pathogen
Pattern recognition
receptor
Macrophage
plasma membrane
When signal binds to the receptor,
it causes the rearrangement of actin
filaments as well as the transcription
of new genes.
The pathogen is endocytosed
Actin
rearrangement
Actin
Acid hydrolases
Lysozyme
Phagosome
Transcription of
target genes
Lysosome
Pattern recognition receptors
Distinguish self from conserved microbial structures.
Drosophila – identified Toll receptors
Mammals – identified Toll-like receptors (TLRs)
Leucine rich repeat (LRR) domain
Indirectly associated with binding PAMPs
May be part of receptor complex
Membrane
TIR domain
Interacts with adaptor protein
PLANTS
Angiosperms
FUNGI
ANIMALS
Vertebrates
Gymnosperms
Urochordates
Insects
Chordates
Arthropods
Mollusks
Nematodes
Brown algae
Coelenterates
Red algae
Green algae
Sponges
Slime molds
Unicellular
PROTOZOA
Yeasts
EUKARYOTES
Mosses
Liverworts
Multicellular
Echinoderms
Ferns
Ancestral Prokaryotes
Adapted from Figure 1-38, Molecular Biology of the Cell, 3rd ed., Garland Publishing, Inc.
Pathogen Perception and Defense Systems
Plants
Legumes
Animals
Mammals
Insects
Last common ancestor may have used a related TLR for pathogen recognition