Effector secretion control by the Pseudomonas aeruginosa type III secretion system

P. aeruginosa uses a type three secretion system (T3SS) to inject effectors into host cells. The T3SS is important for the virulence of P. aeruginosa. The T3SS forms a needle protruding from the bacterial surface and a basal apparatus across the envelope. Effector secretion is triggered by host cell contact; however, the mechanism of effector secretion control is still unclear. The needle tip protein, PcrV, and its chaperone, PcrG, are negative regulators of effector secretion. My thesis study focused on how PcrV and PcrG regulate effector secretion. My data demonstrate that PcrV needs to be secreted and properly assembled at the needle tip in order to control effector secretion. I constructed PcrV and PcrG mutants that abolish the PcrG/PcrV interaction and demonstrated that the PcrG/PcrV interaction is dispensable for effector secretion control. I showed that PcrG regulates effector secretion in the cytoplasm in a PcrV independent manner and the C-terminus of PcrG contains most of the regulatory activity. I identified the apparatus component, PcrD, is the interaction partner of the C-terminus of PcrG and PcrG regulates effector secretion by interacting with PcrD. In addition, I found that the center region of PcrG interacts with PscO, a component of the T3SS ATPase complex. The PcrG/PscO interaction may stabilize the PcrG/PcrD interaction and control secretion activity of the apparatus. My data support the model that PcrV and PcrG allosterically regulate effector secretion. Assembly of PcrV at the needle tip and the interaction between PcrG and PcrD stabilize the apparatus in an effector secretion "off" conformation. Once the bacterium contacts with the host cell, PcrV changes its conformation and PcrG is released from the apparatus; therefore, the apparatus switches to an effector secretion "on" conformation. Furthermore, I isolated the mutations in the needle protein, PscF, that cause de-regulated effector secretion, suggesting that the needle can be locked in effector secretion "on" conformation. Therefore, effector secretion is controlled by conformational changes of the apparatus in P. aeruginosa.