The molecular basis of plant resistance and defense responses to pathogens: Current status

Sruthi.N

Introduction

Many plant-associated microbes are pathogens that impair plant growth and reproduction

Pathogens may proliferate in intercellular spaces (the apoplast) after entering through stomata or hydathodes (bacteria), enter plant epidermal cells, or extend hyphae onto the plant cells (fungi)

Innate immune receptors in plants detect the presence of microbial pathogens and trigger defense responses to terminate or restrict pathogen growth

Elicitors of defence responses

Any substance that has the capability of activating defense responses in plants

Include components of the cell surface as well as excreted metabolites

Elicitors

General Race specific

a) Oligosaccharide elicitors a)avr gene products

b) Protein/peptide elicitors(Ebel et al.,1998)

Perception of elicitor signals

Binding proteins:

Oligosaccharide-binding sites: -Specific glucan-binding sites on soybean root plasma

membranes -high-affinity binding sites for chitin fragments in tomato, rice

Glycopeptide- and peptide-binding sites: -Binding sites for peptidoglycans have been identified in wheat

plasma membranes

Plant defense to pathogens

• Plants respond to infection using a two-branched innate immune system

-Recognition and response to molecules common to many classes of microbes (basal disease resistance)

-Response to pathogen virulence factors (Liu et al.,2008)

Basal defense

Triggered by trans-membrane receptors that recognize conserved molecules released by a variety of (unrelated) microbes

Include cell wall fragments, chitin or peptide motifs in bacterial flagella - PAMPs or MAMPs

PAMP- triggered immunity (PTI)

(Liu et al.,2008)

Secondary defense response

Against virulence effector proteins produced by pathogens

Effector –triggered immunity (ETI)

Mediated by resistance (R) proteins

(Liu et al., 2008)

Phases of pathogen-triggered immunity

(Chisholm et al.,2006)

Concepts regarding pathogen recognition and defense

Gene-for-gene resistance (Flor , 1947)

Guard hypothesis (Beizen et al.,1998)

Gene-for-gene resistance

For resistance to occur, complementary pairs of dominant genes, one in the host and the other in the pathogen, are required (incompatibility)

A loss or alteration to either the plant resistance (R) gene or the pathogen avirulence (Avr) gene leads to disease (compatibility)

(Hammond-Kosack et al., 1997)

Types of genetic interactions between plants and pathogenic microbes

(Hammond-Kosack et al., 1997)

Plant disease resistance genes

Encode proteins that recognize Avr-gene-dependent ligands

Activate signaling cascade(s) that coordinate the initial plant defense responses to impair pathogen ingress

Capacity for rapid evolution of specificity

Common feature of resistance proteins is a leucine-rich repeat

(Hammond-Kosack et al., 1997)

Classes of resistance proteins

(Chisholm et al.,2006)

Extracellular LRR class of R genes

Have classic receptor-kinase formats - an extracelluar LRR, a membrane spanning region and an intracellular protein kinase domain

Against pathogens that have an extracellular lifestyle

Examples: rice Xa21 against Xanthomonas, cf genes of tomato against Cladosporium fulvum.

NB-LRR R proteins

Consists of four domains connected by linkers

A leucine-rich repeat domain (LRR) fused to a nucleotide binding (NB) domain

NB-LRR core equipped with variable amino- and carboxy-terminal domains

(Tameling et al., 2008)

NB-LRR subfamilies

TIR NB-LRRs (Toll/interleukin-1 receptor like NB-LRRs)

CC-NB-LRRs (Coiled Coil NB-LRRs) Solanaceous domain (SD) BED zinc finger DNA-binding domain

(Tameling et al.,2008)

Domains found in plant LRR R proteins

(Tameling et al.,2008)

Deviations from gene-for-gene concept

One R gene may confer specificity to more than one ligand- RPM1 in Arabidopsis confers resistance against P.syringae expressing

either avrRpm1 and avrB

More than one R gene may exist for a given Avr gene- Pto and Prf genes encode biochemically distinct components of the

same pathway- Two genes at the Cf-2 locus furnish identical functions (Bent, 1996)

Guard hypothesis

Key points

a) An effector acting as a virulence factor has a target(s) in the host

b) By manipulating or altering this target(s) the effector contributes to pathogen success in susceptible host genotypes

c) Effector perturbation of a host target generates a “pathogen induced modified self” molecular pattern, which activates the corresponding NB-LRR protein, leading to ETI

(Jones et al.,2006)

a) 9

Guard hypothesis

(Jones et al.,2006)

Plant defense responses

Hypersensitive response

Production of reactive oxygen species

Cell wall fortification

Production of antimicrobial metabolites (phytoalexins)

Defense signal transduction

Synthesis of enzymes harmful to pathogen (eg. chitinases, glucanases)

(Nurnberger et al.,2006)

Types of activated defense responses

(A) Hypersensitive response in single lettuce mesophyll cells penetrated by haustoria of an incompatible isolate of the biotrophic fungus Bremia lactucae B) H2O2 generation in lettuce cell walls, in the vicinity of the incompatible bacterium P. syringae pv phaseolicola(C) Papillae formation. Papillae develop beneath the penetration peg (PP) and germinating spore (S) of an avirulent isolate of the biotrophic fungusErysiphe graminis f sp hordei on barley leaves expressing the Mlg gene.

(Hammond-Kosack et al., 1997)

Defense signaling pathways

SA-dependant signaling

Effective against biotrophic pathogens

Activated by the initiation of HR in plants

Rise in SA levels dissociation of NPR1 oligomers to monomers interaction with TGA-type transcription factors

activation of PR gene expression

TGAs 2,5, and 6 and WRKY70 required for full expression of PR-1 (Glazebrook, 2005)

Defense signaling pathways

JA- and ET- dependent signaling

Effective against necrotrophic pathogens

Increase in JA levels and induction of effector genes (PDF1.2, VSP1)

Induction of transcription factors ERF1, RAP2.6 and JIN1 which activates many defense related genes

Some JA regulated genes also regulated by ET (PDF1.2 , ERF1) (Glazebrook, 2005)

Defense signaling pathways

JA- and ET- dependent signaling

JA levels regulated by cellulose synthases

JAR1 involved in conversion of JA to active form by conjugation with amino acids like isoleucine

In Arabidopsis, all activities of JA requires the function of CO11.

Some responses to JA require the function of an MAP kinase encoded by MPK4

(Glazebrook, 2005)

Defense signaling pathways

Cross-talk between SA and JA/ET signaling

Helps the plant to minimize energy costs and create a flexible signaling network that allows the plant to finely tune its defense response to the invaders encountered

Most reports indicate a mutually antagonistic interaction between SA- and JA dependent signaling.

(Koornneef et al., 2008)

Molecular players in SA/JA crosstalk

NPR1, required for transduction of SA signaling is a suppressor of JA response

Expression of GRX480 is SA inducible and dependent on NPR1

Overexpression of GRX480 completely abolished MeJA-induced PDF1.2 expression, but does not affect the induction of the JA-responsive genes LOX2 and VSP2

(Koornneef et al., 2008)

Molecular players in SA/JA crosstalk

Overexpression of WRKY70 caused enhanced expression of SA-responsive PR genes and suppressed methyl jasmonate (MeJA)-induced expression of PDF1.2

MPK4 is a negative regulator of SA signaling and a positive regulator of JA signaling in Arabidopsis (Koornneef et al., 2008)

PR proteins

Coded by host plants as a response to pathological or related situations

Accumulate not only locally in the place of infection, formed systemically following infection by pathogens.

Wide array of functions: hydrolases, transcription factors, protease inhibitors etc.

(Scherer et al.,2005)

Classes of PR proteins

(Scherer et al.,2005)

Non-Host Resistance

Two mechanisms In case of a potentially new host, pathogen’s effectors could be ineffective, resulting in

little or no supression of PTI, and failure of pathogen growth

One or more of the effector complement of the would-be pathogen could be recognized by the NB-LRR proteins of plants other than it’s coadapted host , resulting in ETI

Arabidopsis resistance to non-adapted powdery mildew Blumeria graminis f. sp. hordei (Jones et al., 2006)

Arabidopsis Nonhost Resistanceto Bgh

(Ellis, 2006)

How R genes initiate defense signaling pathways R proteins recognize pathogen effectors in the cytoplasm

Effector perception may result in altered intra and inter-molecular R protein interactions, including oligomerization.

Activated R proteins cycle into the nucleus and directly bind transcriptional repressors of innate immunity, resulting in transcriptional reprogramming of the plant cell

Studies conducted for N, MLA,RPS4, and RX (Liu et al., 2008)

Nuclear activity of MLA immune receptors

(Liu et al., 2008)

Objective of the study

Study of the interaction of tomato plants with tomato powdery mildew fungus, Oidium neolycopersici

The monogenic genes Ol-1, ol-2 and Ol-4 confer resistance to tomato powdery mildew Oidium neolycopersici via different mechanisms

Study of the molecular and biochemical mechanisms involved

Main interaction stages in compatible and incompatible interactions

H2O2 and callose accumulation in compatible and incompatible interactions

Different expression classes of the DE-TDF in cDNA AFLP analysis

BLAST Results

RT-PCR

Conclusion of the study

ROS, callose accumulation, and upregulation of DE-TDF associated with resistances conferred by dominant and recessive Ol genes

cDNA – AFLP profiling clarified that 81% of upregulated DE-TDF are common for both compatible and incompatible interactions

Class III DE-TDF were up regulated only in incompatible interactions and are specific for specific resistance genes

DE-TDF profiles of NIL-OL-1 and NIL-OL-4 deviated much but similarities were observed between NIL-OL-1 and F3-ol-2

Conclusion

An evolutionary arms race..

Discussion

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