Abstract
The intracellular pathogen Salmonella enterica servovar Typhimurium (S.typhimurium) modulates the host cell's phosphoinositide (PI) metabolism to establish its intracellular replicative niche, the Salmonella-containing vacuole (SCV). Upon invasion, phosphoinositide 3-phosphate (PI(3)P) and other early endosomal markers are rapidly recruited to and remain associated with the SCV throughout its early maturation. While the phosphoinositide 3-phosphatase myotubularin 4 (MTMR4) has an established role in regulating autophagy and cellular PI(3)P-content, two processes associated with the intracellular survival of S. typhimurium, a direct role for MTMR4 in Salmonella biology has not been examined. Here we demonstrate that GFP-tagged MTMR4 is recruited to the SCV and infection of cells depleted of endogenous MTMR4 results in a decrease in viable intracellular Salmonella. This reflects a significant increase in the proportion of SCVs with compromised integrity, which targets the compartment for autophagy and consequent bacterial cell death. These findings highlight the importance of PI(3)P regulation to the integrity of the SCV and reveal a novel role for the myotubularins in bacterial pathogenesis.
Highlights
Salmonella is a Gram-negative facultative intracellular pathogen and a major cause of disease in humans (Haraga et al, 2008)
We demonstrated that inhibition of PIKfyve disrupts the formation of Salmonella Induced Filaments (SIFs) and the intracellular replication of S. typhimurium (Kerr et al, 2010), due to the absence of a PI(3,5)P2-specific probe at the time, we were unable to directly monitor the presence of PI(3,5)P2 accumulation on the Salmonella-containing vacuole (SCV)
The final stages of SCV maturation include the formation of extensive LAMP1-positive tubular structures known as SIFs, with which mCherry-2∗ML1N remains associated at 6 h p.i (Figure 1B), validating the model that the SCV transitions from a PI(3)P-positive compartment to become PI(3,5)P2-positive as it matures
Summary
Salmonella is a Gram-negative facultative intracellular pathogen and a major cause of disease in humans (Haraga et al, 2008). The two predominant human pathogenic strains are Salmonella enterica serovar typhi, responsible for typhoid fever, and S. enterica servoar typhimurium, a causative agent of human gastroenteritis (Haraga et al, 2008). Bacterial effector proteins are translocated directly into the host cells by one of two Type 3 Secretion Systems (T3SSs), to manipulate host membrane trafficking and cytoskeletal elements, initiating macropinocytosis and uptake of the pathogen into the cell (Kubori et al, 1998; Zhou and Galan, 2001). Whilst at least 40 effector proteins are translocated by the Salmonella Pathogenicity Island 1 (SPI1)-T3SS, prominent amongst these is SopB, a phosphatidylinositol phosphatase with sequence similarity to both mammalian phosphatidylinositol 4-phosphatase and phosphatidylinositol 5-phosphatase (Norris et al, 1998).
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