Global Regulatory Pathways Converge To Control Expression of Pseudomonas aeruginosa Type IV Pili

The opportunistic pathogen Pseudomonas aeruginosa relies upon type IV pili (Tfp) for host colonization and virulence. Tfp are retractile surface appendages that promote adherence to host tissue and mediate twitching motility, a form of surface-associated translocation. Tfp are composed of a major structural pilin protein (PilA), several less abundant, fiber-associated pilin-like proteins (FimU, PilV, PilW, PilX, and PilE), and a pilus-associated tip adhesin and surface sensor (PilY1). Several proteins critical for Tfp biogenesis and surface sensing are encoded by the fimU-pilVWXY1Y2E operon. Tfp biogenesis is regulated by the global transcription factor Vfr and its allosteric effector, cyclic AMP (cAMP). Our investigation into the basis for reduced Tfp production in cAMP/vfr mutants revealed a defect in the expression of the fimU operon. We found that cAMP/Vfr activation of the fimU operon occurs via direct binding of Vfr to a specific fimU promoter sequence. We also refined the role of the AlgZ/AlgR two-component system in fimU regulation by demonstrating that phosphorylation of the response regulator AlgR is required for maximal binding to the fimU promoter region in vitro. Vfr also regulates expression of the algZR operon, revealing an indirect regulatory loop affecting fimU operon transcription. Overall, these results demonstrate that two linked but independent regulatory systems couple the expression of Tfp biogenesis and surface sensing genes and highlight the regulatory complexity governing expression of P. aeruginosa virulence factors.