Abstract
The bacterial type III secretion system (T3SS) delivers virulence proteins, called effectors, into eukaryotic cells. T3SS comprises a transmembrane secretion apparatus and a complex network of specialized chaperones that target protein substrates to this secretion apparatus. However, the regulation of secretion switching from early (needle and inner rod) to middle (tip/filament and translocators) substrates is incompletely understood. Here, we investigated chaperone-mediated secretion switching from early to middle substrates in the T3SS encoded by Salmonella pathogenicity island 2 (SPI2), essential for systemic infection. Our findings revealed that the protein encoded by ssaH regulates the secretion of an inner rod and early substrate, SsaI. Structural modeling revealed that SsaH is structurally similar to class III chaperones, known to associate with proteins in various pathogenic bacteria. The SPI2 protein SsaE was identified as a class V chaperone homolog and partner of SsaH. A pulldown analysis disclosed that SsaH and SsaE form a heterodimer, which interacted with another early substrate, the needle protein SsaG. Moreover, SsaE also helped stabilize SsaH and a middle substrate, SseB. We also found that SsaE regulates cellular SsaH levels to translocate the early substrates SsaG and SsaI and then promotes the translocation of SseB by stabilizing it. In summary, our results indicate that the class III chaperone SsaH facilitates SsaI secretion, and a heterodimer of SsaH and the type V chaperone SsaE then switches secretion to SsaG. This is the first report of a chaperone system that regulates both early and middle substrates during substrate switching for T3SS assembly.
Highlights
The bacterial type III secretion system (T3SS) delivers virulence proteins, called effectors, into eukaryotic cells
Class III chaperones, which form heterodimeric complexes with partner proteins known as class V chaperones, bind and stabilize the monomeric needle protein to prevent self-aggregation in the bacterial cytosol
We found that SsaH functions as a class III chaperone in S. enterica Typhimurium Salmonella pathogenicity island 2 (SPI2)-T3SS
Summary
The bacterial type III secretion system (T3SS) delivers virulence proteins, called effectors, into eukaryotic cells. The regulation of secretion switching from early (needle and inner rod) to middle (tip/filament and translocators) substrates is incompletely understood. Our results indicate that the class III chaperone SsaH facilitates SsaI secretion, and a heterodimer of SsaH and the type V chaperone SsaE switches secretion to SsaG. This is the first report of a chaperone system that regulates both early and middle substrates during substrate switching for T3SS assembly. On the basis of the T3SS secretion hierarchy, secreted proteins are divided into early (needle and inner rod), middle (tip/filament and translocators), and late (effectors) substrates. SctU, a member of the export apparatus, is involved in controlling needle length and secretion switching [3, 7, 9, 10]
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