AvrRxo1 Is a Bifunctional Type III Secreted Effector and Toxin-Antitoxin System Component with Homologs in Diverse Environmental Contexts.

Lindsay R Triplett,Jianyong Li,Teja Shidore,Hiromichi Ishihara,Changhe Zhou,Jan E Leach,Qian Han,Shuchi Wu,Jiamin Miao,John Long,Bingyu Zhao,Finbarr Hayes

doi:10.1371/journal.pone.0158856

Lindsay R Triplett, Jianyong Li + Show 10 more

Open Access

https://doi.org/10.1371/journal.pone.0158856

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Abstract

Toxin-antitoxin (TA) systems are ubiquitous bacterial systems that may function in genome maintenance and metabolic stress management, but are also thought to play a role in virulence by helping pathogens survive stress. We previously demonstrated that the Xanthomonas oryzae pv. oryzicola protein AvrRxo1 is a type III-secreted virulence factor that has structural similarities to the zeta family of TA toxins, and is toxic to plants and bacteria in the absence of its predicted chaperone Arc1. In this work, we confirm that AvrRxo1 and its binding partner Arc1 function as a TA system when expressed in Escherichia coli. Sequences of avrRxo1 homologs were culled from published and newly generated phytopathogen genomes, revealing that avrRxo1:arc1 modules are rare or frequently inactivated in some species and highly conserved in others. Cloning and functional analysis of avrRxo1 from Acidovorax avenae, A. citrulli, Burkholderia andropogonis, Xanthomonas translucens, and Xanthomonas euvesicatoria showed that some AvrRxo1 homologs share the bacteriostatic and Rxo1-mediated cell death triggering activities of AvrRxo1 from X. oryzae. Additional distant putative homologs of avrRxo1 and arc1 were identified in genomic or metagenomic sequence of environmental bacteria with no known pathogenic role. One of these distant homologs was cloned from the filamentous soil bacterium Cystobacter fuscus. avrRxo1 from C. fuscus caused watersoaking and triggered Rxo1-dependent cell collapse in Nicotiana benthamiana, but no growth suppression in E. coli was observed. This work confirms that a type III effector can function as a TA system toxin, and illustrates the potential of microbiome data to reveal new environmental origins or reservoirs of pathogen virulence factors.

Highlights

Toxin-antitoxin (TA) systems are bacterial stress-management modules consisting of an antibacterial protein toxin and a neutralizing protein or RNA antitoxin
We examine avrRxo1:arc1 –like modules revealed through recent genome and metagenome studies of environmental microorganisms, and demonstrate that an avrRxo1 homolog from the soil myxobacterium Cystobacter fuscus is able to trigger AvrRxo1-like watersoaking and Rxo1-dependent defense responses when expressed in tobacco
AvrRxo1 has a structure and predicted catalytic residues conserved with Zeta toxins, and functions with Arc1 as a toxin-antitoxin system in E. coli

Summary

Introduction

Toxin-antitoxin (TA) systems are bacterial stress-management modules consisting of an antibacterial protein toxin and a neutralizing protein or RNA antitoxin. Analyses of bacterial genomes later revealed that TA systems are ancient and ubiquitous among free-living bacteria. They are diverse, with over 20 families of TA systems in five major classes, and highly abundant, with up to 97 systems in a single genome [2,3]. Bacterial genomes have revealed that the vast majority of TA systems reside on bacterial chromosomes [2], demonstrating that plasmid maintenance is not the only role for these modules. Direct secretion of a TA system toxin into a eukaryotic host has never been demonstrated

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