Structural analyses of AvrRpm1 and HopBA1: Two TTSS effectors from the plant phytopathogen Pseudomonas syringae.

摘要

Plants recognize microbes via specific pattern recognition receptors that are activated by microbe-associated molecular patterns (MAMPs), resulting in MAMP-triggered immunity (MTI). Successful pathogens bypass MTI in genetically diverse hosts via deployment of effectors (virulence factors) that inhibit MTI responses, leading to pathogen proliferation. Plant pathogenic bacteria like Pseudomonas syringae utilize a type III secretion system to deliver effectors into cells. These effectors can contribute to pathogen virulence or elicit disease resistance, depending upon the host plant genotype. In disease resistant genotypes, intracellular immune receptors, typically belonging to the nucleotide binding leucine-rich repeat family of proteins, perceive bacterial effector(s) and initiate downstream defense responses (effector triggered immunity) that include the hypersensitive response, and transcriptional re-programming leading to various cellular outputs that collectively halt pathogen growth. Nucleotide binding leucine-rich repeat sensors can be indirectly activated via perturbation of a host protein acting as an effector target. AvrRpm1 and HopBa1 are two P. syringae type III effectors. Upon secretion into the host cell, AvrRpm1 is directed to the plasma membrane, where it contributes to virulence. This is correlated with phosphorylation of Arabidopsis RIN4 in vivo. The RPM1 nucleotide binding leucine-rich repeat sensor perceives RIN4 perturbation in disease resistant plants, leading to a successful immune response. Here, we demonstrate that AvrRpm1 has a fold homologous to the catalytic domain of poly(ADP-ribosyl)polymerase. Site-directed mutagenesis of each residue in the putative catalytic triad, His63-Tyr122-Asp185 of AvrRpm1 results in loss of both AvrRpm1-dependent virulence and AvrRpm1-mediated activation of RPM1, but, surprisingly, causes a gain of function: the ability to activate the RPS2 nucleotide binding leucine-rich repeat sensor. Additionally, we determined the crystal structure of HopBA1. We were able to show that despite low sequence similarity, HopBA1 shares structural homology to the ChaN/ EreA-like superfamily of proteins. Through structural analysis of HopBA1 we generated several missense mutations that are critical for recognition inside the host. We were also able to putatively classify two additional type III effectors, HopB1 and HopAC1, from P. syringae as additional ChaN/EreA-like superfamily members.