Biology subjects

Guttman, D. S.

Publications and source records attributed to Guttman, D. S..

2 recordsLinked to original sources

bioRxiv2018

Yeast Genomic Screens Identify Kinesins as Potential Targets of the Pseudomonas syringae Type III Effector, HopZ1a

Gram-negative bacterial pathogens inject type III secreted effectors (T3SEs) directly into host cells to promote pathogen fitness by manipulating host cellular processes. Despite their crucial role in promoting virulence, relatively few T3SEs have well-characterized enzymatic activities or host targets. This is in part due to functional redundancy within pathogen T3SE repertoires as well as promiscuous individual T3SEs that can have multiple host targets. To overcome these challenges, we conducted heterologous genetic screens in yeast, a non-host organism, to identify T3SEs that target conserved eukaryotic processes. We screened 75 T3SEs from the plant pathogen Pseudomonas syringae and identified 16 that inhibited yeast growth on rich media and eight that inhibited growth on stress-inducing media, including the acetyltransferase HopZ1a. We focused our further analysis on HopZ1a, which interacts with plant tubulin and alters microtubule networks. We first performed a Pathogenic Genetic Array (PGA) screen of HopZ1a against ~4400 yeast carrying non-essential mutations and found 95 and 10 deletion mutants which reduced or enhanced HopZ1a toxicity, respectively. To uncover putative HopZ1a host targets, we interrogated both the genetic- and physical-interaction profiles of HopZ1a by identifying yeast genes with PGA profiles most similar (i.e. congruent) to that of HopZ1a, performing a functional enrichment analysis of these HopZ1a-congruent genes, and by analyzing previously described HopZ physical interaction datasets. Finally, we demonstrated that HopZ1a can target kinesins by acetylating the plant kinesins HINKEL and MKRP1.\n\nARTICLE SUMMARYBacterial pathogens utilize secretion systems to directly deliver effector proteins into host cells, with the ultimate goal of promoting pathogen fitness. Despite the central role that effectors play in infection, the molecular function and host targets of most effectors remain uncharacterized. We used yeast genomics and protein interaction data to identify putative virulence targets of the effector HopZ1a from the plant pathogen Pseudomonas syringae. HopZ1a is an acetyltransferase that induces plant microtubule destruction. We showed that HopZ1a acetylated plant kinesin proteins known to regulate microtubule networks. Our study emphasizes the power of yeast functional genomic screens to characterize effector functions.

microbiology↗

bioRxiv2017

Recombination of ecologically and evolutionarily significant loci maintains genetic cohesion in the Pseudomonas syringae species complex

Pseudomonas syringae is a highly diverse bacterial species complex capable of causing a wide range of serious diseases on numerous agronomically important crop species. Here, we examine the evolutionary relationships of 391 agricultural and environmental strains from the P. syringae species complex using whole-genome sequencing and evolutionary genomic analyses. Our collection includes strains from 11 of the 13 previously described phylogroups isolated off of over 90 hosts. We describe the phylogenetic distribution of all orthologous gene families in the P. syringae pan-genome, reconstruct the phylogeny of P. syringae using a core genome alignment and a hierarchical clustering analysis of pan-genome content, predict ecologically and evolutionary relevant loci, and establish the forces of molecular evolution operating on each gene family. We find that the common ancestor of the species complex likely carried a Rhizobium-like type III secretion system (TTSS) and later acquired the canonical TTSS. The phylogenetic analysis also showed that the species complex is subdivided into primary and secondary phylogroups based on genetic diversity and rates of genetic exchange. The primary phylogroups, which largely consist of agricultural isolates, are no more divergent than a number of other bacterial species, while the secondary phylogroups, which largely consists of environmental isolates, have levels of diversity more in line with multiple distinct species within a genus. An analysis of rates of recombination within and between phylogroups revealed a higher rate of recombination within primary phylogroups than between primary and secondary phylogroups. We also found that \"ecologically significant\" virulence-associated loci and \"evolutionary significant\" loci under positive selection are over-represented among loci that undergo inter-phylogroup genetic exchange. These results indicate that while inter-phylogroup recombination occurs relatively rarely in the species complex, it is an important force of genetic cohesion, particularly among the strains in the primary phylogroups. This level of genetic cohesion and the shared plant-associated niche argues for considering the primary phylogroups as a true biological species.

genomics↗