Modulation ofSalmonellavirulence by a novel SPI-2 injectisome effector that interacts with the dystrophin-associated protein complex

Abstract

The injectisome encoded bySalmonellapathogenicity 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-typhoidalSalmonella entericaserovar 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.sseMis widely distributed between the five subspecies ofSalmonella enterica,is found in many clinically-relevant serovars, and is co-transcribed withpipB2, aSPI-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 β-2-syntrophin, utrophin/ dystrophin, α-catulin, α-dystrobrevin and β-dystrobrevin. The interaction between SseM and β-2-syntrophin and α-dystrobrevin was verified inS.Typhimurium-infected cells and relied on the PDZ domain of β-2-syntrophin and a sequence corresponding to a PDZ-binding motif (PBM) in SseM. A ΔsseMmutant strain had a small competitive advantage over the wild-type strain in theS.Typhimurium/mouse model of systemic disease. This phenotype was complemented by a plasmid expressing wild type SseM fromS.Typhimurium orS.Enteritidis and was dependent on the PBM of SseM. Therefore, a PBM within aSalmonellaeffector mediates interactions with the DAPC and modulates systemic growth of bacteria in mice.ImportanceInSalmonella enterica, the injectisome machinery encoded bySalmonellapathogenicity island 2 (SPI-2) is conserved among the five subspecies and delivers proteins (effectors) into host cells that are required forSalmonellavirulence. The identification and functional characterisation of SPI-2 injectisome effectors advances our understanding of the interplay betweenSalmonellaand its host(s). Using an optimised method for preparing secreted proteins and a clinical isolate of the invasive non-typhoidal (iNTS)Salmonella entericaserovar 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 β-2-syntrophin and other components of the dystrophin-associated protein complex (DAPC). To our knowledge, SseM is unique amongSalmonellaeffectors in containing a functional PDZ-binding motif and is the first bacterial protein to target the DAPC.