An Interaction between the Inner Rod Protein YscI and the Needle Protein YscF Is Required to Assemble the Needle Structure of the Yersinia Type Three Secretion System

Shi-Yang Cao,Wan-Bin Liu,Ya-Fang Tan,Hui-Ying Yang,Ting-Ting Zhang,Tong Wang,Xiao-Yi Wang,Ya-Jun Song,Rui-Fu Yang,Zong-Min Du

doi:10.1074/jbc.m116.743591

Abstract

The type III secretion system is a highly conserved virulence mechanism that is widely distributed in Gram-negative bacteria. It has a syringe-like structure composed of a multi-ring basal body that spans the bacterial envelope and a projecting needle that delivers virulence effectors into host cells. Here, we showed that the Yersinia inner rod protein YscI directly interacts with the needle protein YscF inside the bacterial cells and that this interaction depends on amino acid residues 83-102 in the carboxyl terminus of YscI. Alanine substitution of Trp-85 or Ser-86 abrogated the binding of YscI to YscF as well as needle assembly and the secretion of effectors (Yops) and the needle tip protein LcrV. However, yscI null mutants that were trans-complemented with YscI mutants that bind YscF still assembled the needle and secreted Yops, demonstrating that a direct interaction between YscF and YscI is critical for these processes. Consistently, YscI mutants that did not bind YscF resulted in greatly decreased HeLa cell cytotoxicity. Together, these results show that YscI participates in needle assembly by directly interacting with YscF.

Highlights

Bacteria have evolved various secretion mechanisms to interact with their external environment to benefit their survival
Electron microscopic analyses of injectisome purified from S. enterica serovar Typhimurium or injectisomes that are embedded in bacterial membranes of Y. enterocolitica or S. flexneri revealed a syringelike structure protruding from the bacterial surface [1, 6]
The structure of the Yersinia T3SS apparatus includes an YscC secretin ring embedded in the outer membrane [10], an MS ring composed of YscD and YscJ in the inner membrane [11, 12], and a needle that is ϳ60 nm long that protrudes from the bacterial envelope [13]

Summary

Edited by Thomas Söllner

The type III secretion system is a highly conserved virulence mechanism that is widely distributed in Gram-negative bacteria It has a syringe-like structure composed of a multi-ring basal body that spans the bacterial envelope and a projecting needle that delivers virulence effectors into host cells. Type III secretion systems (T3SSs) are widely distributed in Gram-negative bacteria, and they are highly conserved among plant and mammalian pathogens as well as some symbiotic bacteria [1] Human pathogens, such as Salmonella enterica, Shigella flexneri, pathogenic Escherichia coli, and the opportunistic pathogen Pseudomonas aeruginosa, use T3SS machines to deliver various virulence effectors into the host cell cytosol to alter the host cell physiology and cause infectious diseases. Point mutants of YscI amino acid residues Trp-85 and Ile-86 completely disrupt the binding of YscI to YscF, leading to defective YscF polymerization and substrate export as well as greatly decreased cytotoxicity to HeLa cells

Results

Strains or plasmids

This study

Discussion

Experimental Procedures