The export of bacterial toxins across the bacterial envelope requires the assembly of complex, membrane-embedded protein architectures.Pseudomonas aeruginosaemploys type III secretion (T3S) injectisome to translocate exotoxins directly into the cytoplasm of a target eukaryotic cell. This multi-protein channel crosses two bacterial membranes and extends further as a needle through which the proteins travel. We show in this work that PscI, proposed to form the T3S system (T3SS) inner rod, possesses intrinsic properties to polymerize into flexible and regularly twisted fibrils and activates IL-1β production in mouse bone marrow macrophagesin vitro. We also found that point mutations within C-terminal amphipathic helix of PscI alter needle assemblyin vitroand T3SS function in cell infection assays, suggesting that this region is essential for an efficient needle assembly. The overexpression of PscF partially compensates for the absence of the inner rod in PscI-deficient mutant by forming a secretion-proficient injectisome. All together, we propose that the polymerized PscI inP. aeruginosaoptimizes the injectisome function by anchoring the needle within the envelope-embedded complex of the T3S secretome and – contrary to its counterpart inSalmonella –is not involved in substrate switching.
The Pseudomonas aeruginosa T3SS inner rod protein PscI polymerizes into flexible and twisted fibers and induces IL1ß secretion in bone marrow macrophages in vitro through its C terminal region. Polymerized PscI ensures a proper anchoring of the needle to the T3SS basal complex in vivo.