Modulation of
Salmonella
virulence by a novel SPI-2 injectisome effector that interacts with the dystrophin-associated protein complex
Authors: Yu Xiu-Jun , Xie Haixia , Li Yan , Liu Mei , Hou Ruhong , Predeus Alexander V. , Perez Sepulveda Blanca M. , Hinton Jay C. D. , Holden David W. , Thurston Teresa L. M. 
Published: June 2024

The injectisome encoded by
Salmonella
pathogenicity island 2 (SPI-2) had been thought to translocate 28 effectors. Here, we used a proteomic approach to characterize 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 among the five subspecies of
Salmonella enterica,
is found in many clinically relevant serovars, and is co-transcribed with
pipB2
, a SPI-2 effector gene. The 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 in
Salmonella
Typhimurium-infected cells and relied on the postsynaptic density-95/discs large/zonula occludens-1 (PDZ) domain of β-2-syntrophin and a sequence corresponding to a PDZ-binding motif (PBM) in SseM. A Δ
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 the systemic growth of bacteria in mice. Furthermore, the Δ
sseM
mutant strain displayed enhanced replication in bone marrow-derived macrophages, demonstrating that SseM restrains intracellular bacterial growth to modulate
Salmonella
virulence.

IMPORTANCE

In
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, which are required for
Salmonella
virulence. The identification and functional characterization of SPI-2 injectisome effectors advance our understanding of the interplay between
Salmonella
and its host(s). Using an optimized method for preparing secreted proteins and a clinical isolate of the invasive non-typhoidal
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 postsynaptic density-95/discs large/zonula occludens-1 (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 among
Salmonella
effectors in containing a functional PDZ-binding motif and is the first bacterial protein to target the DAPC.