Dissecting the Arms Race between Pathogenic Bacteria and the Plant Immune System

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Pathogens and their hosts engage in a co-evolutionary arms race for survival, which is mediated by pathogen virulence factors and the host immune system. The first level of cellular interaction is the recognition of conserved microbe-associated molecular patterns (MAMPs) by host pattern recognition receptors (PRRs). Some pathogens are able to suppress this immune response via type III secreted effectors (T3SEs). Some plant hosts can in turn respond to T3SEs via nucleotide binding leucine rich repeat resistance proteins (NLRs), which trigger a second layer of immunity upon recognition of T3SE activity. This proposal has two major goals: Goal 1. Dissect the arms race between the HopZ family of T3SEs and their host targets. HopZ alleles are carried by the agronomically-important phytopathogenic bacterium Pseudomonas syringae. This diverse family of T3SEs has four major allele sub-families that can act as either virulence factors or immune elicitors depending on the presence of the ZAR1 NLR in Arabidopsis. We have recently shown that one HopZ allele acetylates a variety of host proteins including the pseudokinase ZED1 and a family of kinases closely related to ZED1 called ZRKs. ZED1 is monitored / guarded by ZAR1. - Our hypotheses are: HopZ T3SEs target ATP-binding domain proteins; different HopZ alleles target different members of the ZED1/ZRK family; HopZ acetylation of ZRKs interferes with their kinase activity; and inhibition of ZRK activity suppresses immune signaling. - Our specific aims for this goal are to: 1a) Determine which HopZ alleles are acetyltransferases. 1b) Assess physical and biochemical interactions between the HopZ and ZED1/ZRK families. 1c) Identify residues putatively important for interaction and post-translational modification in the HopZ and ZRK families. 1d) Determine if specific ZRK family members are important for bacterial virulence or host immunity. And 1e) Determine if post-translation modification of HopZ or ZRK proteins modulates virulence or immune activation. Goal 2. Dissect the arms race between bacterial MAMPs and plant PRRs. We have developed novel screens to identify new MAMPs and their cognate PRRs, and want to determine what role these interactions play during the very early stages of host adaptation. - Our hypotheses are: there are many more MAMPs in any given pathosystem than have been previously characterized; MAMP-PRR interactions are highly specific, but most interactions signal through a fairly small number of regulatory PRRs; MAMP variation can result in modulation of host specificity; and the first step in a host switch involves evolution of MAMPs away from PRR recognition. - Our specific aims for this goal are to: 2a) Identify the PRRs that recognize newly identified MAMPs. 2b) Validate the direct interaction of PRRs and MAMPs. 2c) Identify the first changes associated with host range expansion via experimental evolution. And 2d) Determine how MAMPs evolve over recent evolutionary time and the consequence of this for immune elicitation. These studies continue our comparative and functional dissection of one of the best established pathosystems for host-specificity research. Through these aims we will decode the range of functional interactions between a diverse family of bacterial T3SEs and the even more diverse family of host proteins with which they interact. We will then mechanistically characterize the interactions that modulate bacterial virulence and host immunity. We will also examine the diversity of MAMPs and their cognate PRRs. We will determine how this variation influences host specificity, and produce fundamental research that will be the foundation for future applied work focusing on the use of MAMPs as alternative antimicrobials.