Download Lectures 15-16 Molecular mechanisms of plant

Lectures 15-16
Molecular mechanisms of plant-pathogen interactions
During the contact between plant and pathogen, a
particular chain of events is produced in the plant
organism.
Two ways of possible interaction between plant and pathogen
• The plant is provided by a receptor that interacts with bacterial protein. As a result,
quick protective reaction is being developed. In such a situation, the bacteria is called
avirulent for a given plant genotype (Piffanelli P. et al., 1999, Martin G.B., 1999).
•• The proteins of the pathogenic organism are virulent for the given plant genotype. The
plant is affected by the pathogen, whereas protective mechanisms are being activated
more slowly (Maleck K and Lawton K., 1998). In both cases, with the start of
pathogenesis gene transcription, the cell walls strengthen. Then in the place of pathogen
penetration, the active forms of oxygen are formed, causing the death of infected cells.
Molecular mechanisms of plant resistance to disease Plant
disease caused by viruses, bacteria and fungi are controlled naturally by plant host
resistance (R) genes and avirulence (avr) genes of the pathogen.
This has been termed gene-for-gene resistance.
A simple model explains these gene-for-gene interactions: Avirulence gene products
generate signals (ligands) and resistance genes encode cognate receptors.
Disease resistance is usually mediated by dominant genes, but some recessive resistance
genes also exist.
In early 1900s it was recognized that resistance to plant pathogens was often inherited as
a single dominant or semidominant trait.
1940s – Harold Flor studied flax and flax rust pathogen.
Gene-for-gene interaction.
Model: Plant resistance will only occur when a plant possesses a dominant resistance
gene (R ) and a pathogen expresses complementary dominant avirulence gene (Avr).
Holds true for most plant-pathogen interactions.Flor’s model: For
resistance(incompatibility) to occur, complementary pairs of dominant genes, one in the
host one in the pathogen, are required. An alteration or loss of the plant resistance gene
(R changing to r) or of the pathogen avirulence gene (Avr changing to avr) leads to
disease (compatible).Different model – for pathogens that deploy host-selective toxins
for successful pathogenesis.
Pathogen virulence must be dominant because a functional toxin or enzyme (or both)
must be produced to cause disease.
Plant resistance is predominantly inherited as a dominant trait and is achieved through
enzymatic detoxification or through loss or alteration of the toxin in the pathogen.
Interactions involved in toxin-dependent compatibility.
The wild-type pathogen gene Tox is required for synthesis of a toxin that is crucial for
pathogenesis; tox is the corresponding recessive, nonfunctional allele. The dominant
allele of the host R gene is required for detoxification-based resistance. Resistance can
only occur when the host expresses a toxin-insensitive for of the toxin target. Disease
results only when the plant can not detoxify the toxin produced by the pathogen.The term
disease tolerance describes a neutral outcome observed in plant-pathogen associations.
The interaction is genetically compatible, but the plant somehow restricts the biochemical
processes required for symptom development.
As a result tissue damage is kept down even if the plants are heavily infected.
Disease-tolerant plants act as important reservoirs of pathogen inocula, which may go
and infect susceptible species.Properties of some avirulence genesAvr
genes are recognized as the genetic determinants of incompatibility toward particular
plant genotypes.
Functions of Avr genes of phytopathogenic bacteria or fungi are not completely
understood.
Plant viruses – conclusive exceptions.
In different incompatible interactions the viral CP, replicase and MP are recognized as
avirulence factors.
Changes in amino acids that do not substantially compromise the primary function of
protein in pathogenesis can still alter their avr specificity.
A mutation from avirulence (Avr) to virulence (avr) is often associated with the loss of
fitness for growth on plants that lack the R gene.
Thus, Avr gene products have important roles in microbial pathogenicity at different
stages: growth and reproduction in plants, symptom development, transfer to other
plants.1984 – the first avirulence gene was isolated from Pseudomonas bacteria infecting
soybean.
Later 30 avr genes were isolated from Pseudomonas and Xanthomonas species.
Bacterial avr genes code for soluble, hydrophylic proteins.
Avr proteins share little homology with other known proteins, but there is substantial
homology between some Avr proteins.
Two types of distinct Avr-generated signals are recognized. Exported syringolides (Cglucosides) are produced by enzymes encoded by the avrD locus of P. syringae pv.
glycinea, and these substances trigger a resistance response in soybean harboring
corresponding R gene, Rpg4.
For other bacterial species, the Avr protein itself is now thought to be a signal.The
Xanthomonas avrBs3 family of avr genes is distinct from the Pseudomonas genes.
avrBs3 family proteins contain a reiterated internal motif 34 aa lonf.
Eg: AvrBs3 gene product has 17.5 nearly identical repeats of this motif.
By deleting some of these repeats the specificity of this Avr gene can be altered, so that
it is no longer recognized by the corresponding Bs3 resistance gene in pepper plants.
Other internal deletions in the AvrBs3 lead to the gain-of-function phenotype, resulting in
resistance response in plant carrying recessive bs3 allele.Delivery of AvrBs3 in to the
plant cell cytoplasm via the type III secretion system appears to be necessary for the
function of the Xanthomonas Avr gene products.
Mutational studies of the two functional nuclear localization signals in the C-terminus of
the AvrBs3 protein reveal that the Avr protein has to be targeted to the plant cell nucleus
for Bs3-mediated resistance to operate.
In fungi only few Avr genes are known.
For fungal species that colonize only the intercellular airspaces of plants, a biochemical
approach can isolate small secreted peptides capable of eliciting R-gene dependent plant
defense in the absence of pathogens. They appear to be direct elicitors, but their role is
still elusive.Resistance (R) genesThere are many resistance (R) genes in plant
hosts, each conferring a unique specificity to various pathogen isolates. These R genes
often are clustered as complex gene-families in plant genomes.
In general, R genes function to recognize, directly or indirectly, "elicitor" molecules
produced by the invading pathogen.
This recognition results in a rapid signal cascade, leading to an active defense response. R
genes (and their associated responses) are exploited by plant breeders to offset yield loss
due to pathogen infection
R genes and R gene-mediated disease resistanceIsolation
strategies:
Locating R gene on the chromosome by using plant populations that segregate for
resistant and susceptible individuals.
Identifying the correct sequence by inserting either transposon to destroy biological
activity, or by using binary cosmid complementation to confer resistant phenotype on a
susceptible plant.
R genes were isolated from 3 monocotyledonous plants and from 5 dicotyledonous
plants.
Provide resistance to a range of taxonomically unrelated pathogens.
4of 6 classes of predicted R proteins which mediate dominant or semidominant
resistance, have leucine-rich repeats (LRRs) – structural motifs seen in proteins that
function in signal transduction pathways.
LRR motifs have been shown to mediate protein-protein or receptor-ligand interactions
in many different kinds of proteins.
Genetic evidence suggests that the beta-strand/beta-turn of the LRR is a key region in the
R protein and appears to determine its specificity.Many proteins also have a central
nucleotide-binding site (NBS) that contains several conserved domains, the function of
which is still unknown.
Although they do not have intrinsic kinase activity, they could bind ATP or GTP and
then activate the defense response.
Mutations in key residues in the NBS destroy the R protein function.
Some current models view NBS as adaptor region, linking the C-terminal LRR
recognition domain to various N-terminal effectors.
Some R protein possess a putative leucine zipper (LZ) or coiled-coil sequence between
the N terminus and the NBS domains.
Lzs are known for their roles in homo- and hetero-dimerization of eukaryotic
transcription factors as well as facilitating interactions between proteins with other
functions.
Other NBS-LRR R proteins contain a large N-terminal domain called Toll/interleukin1/resistance (TIR) domain, that is similar to the cytoplasmatic signaling domain in
Drosophila
How R and Avr gene products activate plant defense
responses is not understoodEach R gene product is thought to possess
two functions:
•Recognition of corresponding Avr-derived signal
•And activation of down-stream signaling pathways to trigger complex defense
responses.
The various predicted protein structures provide the clues of the possible
mechanisms.Example: Pto kinase – confers resistance to strains of Pseudomonas that
express avrPto, a protein that bacterium delivers into the cell by type II secretion system.
Yeast two-hybrid technology provided clues that it may interact directly with the avrPto
signal..
Several plant protein that interact with the Pto kinase were identified, including proteins
that have homology with transcription factors and another protein kinase, Pti1. Three of
the possible transcription factors – Pti4, Pti5 and Pti6 possess highly conserved DNAbinding domain that recognizes a core hexanucleotide sequence – a sequence
foundPlant-pathogen interactionThe basic principle behind these are that there is a
gene product that inhibits an effect or product of the viral infection. One gene for
infection, one gene to resist. A few examples are the L6 gene for the Flax Rust virus, the
N gene in tobacco for TMV resistance, as well as the RPS2 gene in Arabidopsis for
bacterial resistance.
These gene-for-gene resistance's operate on the basis of recognition of the infecting
particle by an interaction between the host resistance system, and the avirulence genes of
the infecting virus. This results in the activation of anti-pathogen molecules, or
hypersensitive cell death pathways, in the tissues around the point of infection.
The RPS2, and N gene products have domains for nucleotide binding, and repeats of
leucine-rich area. The postulated mechanism involves a signal transduction that is
dependent on A\GTP, responding to pathogen avirulence genes.
Pathogen – any organism which is able to cause
diseaseDisease would result in appearance of specific symptoms due to the change
of host metabolism required by pathogen or as a specific host response
Important definitions•Targeting of
R-gene mediated resistance leads to the
expression of salicylic acid (SA)
••SA expression results in hypersensitive response (HR) and systemic acquired resistance
(SAR)
••HR – necrotic lesions around the infection sites due to profound burst of reactive
oxygen species
••SAR – an inducible plant defence state, the activation of which depends mostly on the
accumulation of SA At the present moment, the best studied are the molecular
mechanisms providing hypersensitive response (HR) (Halterman D.A. and Martin G.B.,
1997).
In this case, the plant receptor interacts with the pathogen molecule. In order such
interaction could occur, the plant and bacteria of a certain genotype should meet, i.e., a
bacteria carrying the avirulence gene (avr) interacts with a plant, which has the
corresponding R-gene.
Such process is called an incompatible combination and leads to quick progressing of
events, or to hypersensitive response.
Receptors activate the passes of signal transduction and launch several protective
systems. In the place of pathogen penetration, the strong oxidants are being synthesized
such as H2O2, O2· - , OH· .
Then the oxidative burst is being developed, followed by the death of infected cells
according to the mechanism similar to apoptosis known in vertebrates (Lam E. et al.,
1999).
Hypersensitive response is being developed in the place of pathogen penetration into the
neighboring cells. Rapidly developing local process produces signal molecules spreading
along vascular system of a plant.
In dependence of local events, the set of signal molecules is being organized, which in
turn, forms this or that generalized response.
In the whole plant organism, the pathogenesis-related genes (PR-genes) are activated,
the cell walls strengthen, and the plant accumulates some amount of protective
substances, which are more effective in the struggle with this definite pathogenic form. In
the plant cells, the salicylic acid (SA) is produced in considerable amounts and causes
activation of SA-induced genes. The integrity of these events is named as systemic
acquired resistance (SAR) (Mittler R. et al., 1999).
The other systemic, or referring to the whole organism, response to pathogen infection is
an induced systemic resistance (ISR). Its differs from SAR by activation of some
differing set of pathogenesis genes and by other ways of signal transduction (without
participance of salicylic acid) (Pieterse C.M.J. et al., 1996).
The plant is not always supplied with receptors to the proteins of “attacking” bacteria or
fungi. In this situation, the pathogen is called viral for given plant genotype; and the pair
plant-pathogen is compatible. In this case, pathogen molecules are non-specific elicitors,
which are non-specific substances causing pathogenesis.
The ways of obtaining the signal from non-specific elicitors are still unknown. Various
external stimuli (wound, non-specific elicitors) activate protein kinases and genes of
signal molecules biosynthesis. In the course of signal transduction, the synthesis of JA,
NO, H2O2, SA, and ethylene is produced.
The processes are being activated that are well-known in animals: proteinkinase cascade,
polyubiquitine-dependent protein degradation, etc. The signal transduction paths
frequently intercross.
For example, the gene of one of the key enzymes, PAL (phenylalanine ammonia-lyase),
is activated not only in the course of hypersensitive response, but also in response to
various external stimuli (Dixon R.A. and Paiva N.L. 1995, Mauch-Mani B. et al., 1996).
PAL takes part in the synthesis of SA, phytoalexins, and lignin monomers. The latter,
both in hypersensitive response and non-specific pathogenesis induction, activate
pathogenesis-related (PR) genes What is the role of SA?
•SA plays an important role in the support of HR (transgenic plants which express the
enzyme degrading SA are not resistant to TMV infection)
•SA was shown to move from the infected leaf into the other parts of the plant (up to 70%
of labelled SA was found in the upper leaves).
ROS are often produced during the early stages of a plant resistance
responseVarious
PR proteins
From B.Buchanan, W. Gruissen, R. Jones
Part of the material adapted from
B.Buchanan, W. Gruissen, R. Jones, Biochemistry and molecular biology of plants, 2000,
ASPP Publisher.
http://taipan.nmsu.edu/EPWS310Dr. Roger Wise, USDA-ARS-Corn Insects and Crop
Genetics Research
Unithttp://www.public.iastate.edu/~imagefpc/Subpages/research.html Dr. Barbara Baker
http://plantbio.berkeley.edu/faculty/faculty_pages/Baker.htmlhttp://wwwmgs.bionet.nsc.r
u/mgs/papers/goryachkovsky/plant-trrd/pathogen1.htm