Surface-Induced cAMP Signaling Requires Multiple Features of the <i>Pseudomonas aeruginosa</i> Type IV Pili

Abstract Pseudomonas aeruginosa type IV pili (TFP) are important for twitching motility and biofilm formation. TFP have been implicated in surface sensing, a process whereby surface-engaged cells up-regulate synthesis of the second messenger cAMP to propagate a signaling cascade leading to biofilm initiation and repression of motility. Here we show that mutations in PilA impairing proteolytic processing of the prepilin into mature pilin as well as the disruption of essential TFP components, including the PilC platform protein and PilB assembly motor protein, fail to induce surface-dependent cAMP signaling. We show that TFP retraction by surface-engaged cells is required to induce signaling, and that the retractile motor PilT is both necessary and sufficient to power surface-specific induction of cAMP. The PilU retraction motor, in contrast, is unable to support full signaling in the absence of PilT. Finally, while we have confirmed that PilA and PilJ interact by bacterial two-hybrid analysis, our data do not support the current model that PilJ-PilA interaction drives cAMP signaling. Importance Surface sensing by P. aeruginosa requires TFP. TFP play a critical role in the induction of the second messenger cAMP upon surface contact; this second messenger is part of a larger cascade involved in the transition from a planktonic to biofilm lifestyle. Here we show that TFP must be deployed and actively retracted by the PilT motor for the full induction of cAMP signaling. Furthermore, the mechanism whereby TFP retraction triggers cAMP induction is not well understood, and our data argues against one of the current models in the field proposed to address this knowledge gap.