The pilT gene contributes to type III ExoS effector injection into epithelial cells in Pseudomonas aeruginosa

A type IV pilus filament, mainly composed of PilA, is retracted by the driving power generated by PilT and PilU ATPases. pilA is required for injection of type III ExoS effectors into epithelial cells thereby facilitating Pseudomonas aeruginosa penetration through the epithelial barrier by impairing the defense function of tight junctions. Here, we examined whether the pilT and pilU of the P. aeruginosa PAO1 strain are required for ExoS injection into epithelial cells. We measured the quantity of ExoS injected into epithelial cells, and found that within such cells its quantity decreased by 80% (ΔpilA strain), 75% (ΔpilT strain), and 30% (ΔpilU strain) compared with the wild-type strain. pilT deficiency decreased the disruption of human epithelial colorectal adenocarcinoma (Caco-2) cell monolayers to the same extent as that of pilA and exoS deficiency, whereas pilU deficiency decreased disruption of the monolayers less than deficiency of the other genes. pilT and pilU deficiency decreased bacterial penetration of the Caco-2 cell monolayers to the same level as pilA and exoS deficiency. Our data showed that the pilU gene expression level was reduced in the PAO1 strain after adhesion to Caco-2 cell surfaces, but the expression levels of the pilA and pilT genes did not change. We conclude that P. aeruginosa injects ExoS into cells through the function of type IV pilus retraction, and that pilT makes a greater contribution to this process than pilU.