Date Published: April 20, 2017
Publisher: Public Library of Science
Author(s): Ciaran E. Finn, Audrey Chong, Kendal G. Cooper, Tregei Starr, Olivia Steele-Mortimer, Denise M. Monack.
Type III secretion system 1 (T3SS1) is used by the enteropathogen Salmonella enterica serovar Typhimurium to establish infection in the gut. Effector proteins translocated by this system across the plasma membrane facilitate invasion of intestinal epithelial cells. One such effector, the inositol phosphatase SopB, contributes to invasion and mediates activation of the pro-survival kinase Akt. Following internalization, some bacteria escape from the Salmonella-containing vacuole into the cytosol and there is evidence suggesting that T3SS1 is expressed in this subpopulation. Here, we investigated the post-invasion role of T3SS1, using SopB as a model effector. In cultured epithelial cells, SopB-dependent Akt phosphorylation was observed at two distinct stages of infection: during and immediately after invasion, and later during peak cytosolic replication. Single cell analysis revealed that cytosolic Salmonella deliver SopB via T3SS1. Although intracellular replication was unaffected in a SopB deletion mutant, cells infected with ΔsopB demonstrated a lack of Akt phosphorylation, earlier time to death, and increased lysis. When SopB expression was induced specifically in cytosolic Salmonella, these effects were restored to levels observed in WT infected cells, indicating that the second wave of SopB protects this infected population against cell death via Akt activation. Thus, T3SS1 has two, temporally distinct roles during epithelial cell colonization. Additionally, we found that delivery of SopB by cytosolic bacteria was translocon-independent, in contrast to canonical effector translocation across eukaryotic membranes, which requires formation of a translocon pore. This mechanism was also observed for another T3SS1 effector, SipA. These findings reveal the functional and mechanistic adaptability of a T3SS that can be harnessed in different microenvironments.
Type III Secretion Systems (T3SSs) are used by a variety of Gram-negative bacteria for interkingdom delivery of proteins (known as effectors) from the bacterial cytosol into eukaryotic cells . For bacterial pathogens, such as Salmonella enterica, Yersinia spp, and pathogenic Escherichia coli, these molecular syringes are key virulence determinants essential for a variety of processes including: adherence; invasion; intracellular survival and cytotoxicity. This broad repertoire is due to the diversified nature of effectors rather than the mechanism of delivery, which is highly conserved [2,3]. T3SS delivery is a contact-dependent process characterized by the formation of a pore, or translocon, at the point of contact with the eukaryotic membrane and through which effectors are delivered into the host cell .
The ability of Salmonella to actively invade and colonize epithelial cells is critical for pathogenesis. Following invasion, the bacteria can survive and replicate within the SCV, a modified phagosome, or in the cytosol. Effectors translocated by the SPI1-encoded T3SS1 play critical roles in invasion. Here, we show a novel role for T3SS1 in the cytosolic subpopulation of intracellular bacteria (See model in Fig 9). Delivery of the effector SopB, via a translocon-independent mechanism, leads to Akt phosphorylation and prolonged survival of epithelial cells containing cytosolic Salmonella.