Pseudomonas syringae pv. tomato DC3000中第六型分泌系統VgrG蛋白之特性分析

Abstract

The type VI secretion system (T6SS), found in most gram-negative bacteria, is a macromolecular structure analogous to bacteriophage tails, with Hcp stacking to form a tail-tube like structure and VgrG, acting as a puncturing device at the tip of this tube. The plant pathogen Pseudomonas syringae pv. tomato DC3000 (Pst DC3000) is known to have two putative T6SS gene clusters, HSI-I and HSI-II. Hcp2, encoded in the HSI-II gene cluster, previously shown to be secreted and playing an essential role in antimicrobial activity against enterobacteriaceae. However, the role of VgrG in this bacterium is still unknown. In this study, a total of seven vgrG genes were predicted to be present in Pst DC3000, three of which are located within the HSI-I and HSI-II gene clusters with additional four others scattered in the bacterial genome. Here, we demonstrate the secretion of VgrG proteins, and also, the contribution of each VgrG protein in secretion and interbacterial competition by systematic deletion of these genes. Among these VgrG homologs, VgrG-2b (PSPTO_5436) was the major contributor in secretion and interbacterial competition activity against E. coli and five different phytopathogenic bacteria followed by a small contribution of VgrG-2a (PSPTO_5415). Both VgrG proteins are encoded within the HSI-II gene cluster. Furthermore, VgrG-2b was shown to directly interact with Hcp2 in vitro. All these further support the notion that HSI-II plays a major role in Pst DC3000 to compete for niches, thus conferring this bacterium significant fitness advantage against competitive microbes. Additionally, eleven putative VgrG-associated genes were targeted for deletion to observe its role in the T6SS results showed a potential effector activity for PSPTO_5413, located downstream of VgrG-2a.