Modulation of Salmonella virulence by a novel SPI-2 injectisome effector that interacts with the dystrophin-associated protein complex.

Abstract

The injectisome encoded by Salmonella pathogenicity island 2 (SPI-2) had been thought to translocate 28 effectors. Here, we used a proteomic approach to characterise the secretome of a clinical strain of invasive non-typhoidal Salmonella enterica serovar Enteritidis, that had been mutated to cause hyper-secretion of the SPI-2 injectisome effectors. Along with many known effectors, we discovered the novel SseM protein. sseM is widely distributed between the five subspecies of Salmonella enterica, is found in many clinically-relevant serovars, and is co-transcribed with pipB2, a SPI-2 effector gene. Translocation of SseM required a functional SPI-2 injectisome. Following expression in human cells, SseM interacted with five components of the dystrophin-associated protein complex (DAPC), namely {beta}-2-syntrophin, utrophin/ dystrophin, -catulin, -dystrobrevin and {beta}-dystrobrevin. The interaction between SseM and {beta}-2-syntrophin and -dystrobrevin was verified in S. Typhimurium-infected cells and relied on the PDZ domain of {beta}-2-syntrophin and a sequence corresponding to a PDZ-binding motif (PBM) in SseM. A {Delta}sseM mutant strain had a small competitive advantage over the wild-type strain in the S. Typhimurium/mouse model of systemic disease. This phenotype was complemented by a plasmid expressing wild type SseM from S. Typhimurium or S. Enteritidis and was dependent on the PBM of SseM. Therefore, a PBM within a Salmonella effector mediates interactions with the DAPC and modulates systemic growth of bacteria in mice. ImportanceIn Salmonella enterica, the injectisome machinery encoded by Salmonella pathogenicity island 2 (SPI-2) is conserved among the five subspecies and delivers proteins (effectors) into host cells that are required for Salmonella virulence. The identification and functional characterisation of SPI-2 injectisome effectors advances our understanding of the interplay between Salmonella and its host(s). Using an optimised method for preparing secreted proteins and a clinical isolate of the invasive non-typhoidal (iNTS) Salmonella enterica serovar Enteritidis strain D24359, we identified 22 known SPI-2 injectisome effectors and one new effector - SseM. SseM modulates bacterial growth during murine infection and has a sequence corresponding to a PDZ-binding motif that is essential for interaction with the PDZ-containing host protein {beta}-2-syntrophin and other components of the dystrophin-associated protein complex (DAPC). To our knowledge, SseM is unique among Salmonella effectors in containing a functional PDZ-binding motif and is the first bacterial protein to target the DAPC.