Membrane damage induces TagQ/TagR complex-mediated H1-T6SS assembly in Pseudomonas aeruginosa

The type VI secretion system (T6SS) is a large macromolecular complex crucial for inter-bacterial competition. Its expression and localization are tightly regulated, with the anti-bacterial H1-T6SS in Pseudomonas aeruginosa assembling in response to membrane damage. While several proteins are known to regulate this response, the mechanism remains unknown. Here, we use biochemical approaches to show formation of a complex between the membrane lipoprotein TagQ and the periplasmic protein TagR. Furthermore, Co-IP and in silico modeling indicates TagQ/R complex interactions with PpkA, likely dimerizing the kinase to initiate a phosphorylation cascade leading to H1-T6SS assembly. Mass spectrometry analysis of phosphorylated proteins in P. aeruginosa shows that PpkA does not have any targets outside of the previously reported Fha1, suggesting the sensor module acts exclusively to regulate T6SS assembly. Our work reveals the functional basis of the TagQR-PpkA signaling pathway and links membrane damage to localized H1-T6SS assembly, which is critical for efficient retaliation during inter-bacterial interactions.