Defining the role of <i>Pseudomonas aeruginosa</i> PilY1 in signaling and virulence

<i>Pseudomonas aeruginosa</i> (Pa) is an important opportunistic pathogen that has many virulence factors expressed in a coordinated manner to cause infection. The PilY1 protein is a component of the type IV pili (T4P) but has additional signaling functions outside of the pilus. A major function of PilY1 is to prevent the AlgZ/R two-component system from functioning. However, the complete effects of PilY1 signaling through the control of the AlgZ/R or other regulatory systems are not well understood. In this study, we determine new genes controlled by the AlgZ/R system that are impacted by PilY1. We discovered that PilY1 impacts cAMP by preventing the AlgZ/R system from activating the major adenylate cyclase, <i>cyaB</i>. PilY1 also prevents the AlgZ/R system from activating itself and a putative c-di-GMP phosphodiesterase, <i>PA4781</i>. Furthermore, PilY1 functions in different <i>P. aeruginosa</i> strains, including mucoid strains. Surprisingly, bacterial survival in the lung, liver, and blood did not require PilY1. Overall, these findings suggest that PilY1 is critical for preventing AlgZ/R activity to prevent improper second messenger production. This work identifies new AlgZ/R targets and implicates the AlgZ/R system in the control of cAMP in <i>P. aeruginosa</i>. This work identifies a possible way to use PilY1 to prevent the expression of the major adenylate cyclase using PilY1 which could be used to impact <i>P. aeruginosa</i> virulence.IMPORTANCEUnderstanding PilY1 signaling is important for elucidating how <i>P. aeruginosa</i> adapts to different environments. A major function for PilY1 is to interfere with the AlgZ/R two-component system. Here, we use transcriptomics to determine genes PilY1 affects through the AlgZ/R system. We identified new AlgZ/R targets and established a mechanism for impacting cAMP levels. Our findings further our understanding of PilY1 and the AlgZ/R system and suggest a possible way that PilY1 might be used to prevent cAMP from increasing in <i>P. aeruginosa</i>.