Project Grant F32AI186488

INVESTINGATING THE MECHANISMS CONTROLLING NEISSERIA GONORRHOEAE TYPE IV PILUS DYNAMICS - PROJECT SUMMARY THE NEISSERIA GONORRHOEAE (THE GONOCOCCUS OR GC) TYPE IV PILUS (T4P) IS A CRITICAL COLONIZATION AND VIRULENCE FACTOR DURING PATHOGENESIS. T4P ARE USED TO ADHERE TO HOST CELLS AND PROMOTE COLONIZATION. T4P ARE DYNAMIC STRUCTURES MADE OF PILIN FIBERS THAT EXTEND AND RETRACT FROM THE BACTERIAL CELL. BEYOND HOST CELL ADHESION, T4P ARE INVOLVED IN A VARIETY OF CELLULAR PROCESSES INCLUDING DNA UPTAKE, TWITCHING MOTILITY, AND RESISTANCE TO ANTIMICROBIAL AGENTS. WE PREVIOUSLY IDENTIFIED A NOVEL COMPONENT OF THE GC T4P, TFPC. OUR PUBLISHED DATA ILLUSTRATED THAT TFPC IS REQUIRED FOR FULL PILIATION, AND PILIATION IS RESTORED IN TFPC MUTANTS LACKING THE PILT RETRACTION ATPASE. THERE ARE ELEVEN OTHER SUCH MUTANTS IN THE GC T4P. HERE, WE USE GENETIC, BIOCHEMICAL, AND BIOPHYSICAL TECHNIQUES TO DEFINE HOW TFPC CONTRIBUTES TO T4P DYNAMICS AND ARCHITECTURE. WE SHOW THAT WHILE TFPC IS REQUIRED FOR COMPLETE PILIATION OF GC, REMAINING PILUS STRUCTURES UNDERGO EXTENSION AT RATES SIMILAR TO PARENTAL STRAINS. HOWEVER, RETRACTION IS SLOWER IN STRAINS LACKING TFPC. FURTHERMORE, WE DEMONSTRATE THAT TFPC STABILITY IS DEPENDENT UPON THE PILQ SECRETIN. WE ALSO IDENTIFY AN IMPORTANT MOTIF WITHIN THE FLEXIBLE, DISORDERED, PROLINE-RICH REGION OF TFPC THAT IS IMPORTANT FOR FUNCTION OR STABILITY OF TFPC. I HYPOTHESIZE THAT TFPC INTERACTS WITH THE PILQ SECRETIN OR OTHER COMPONENTS OF THE T4P TO SUPPORT A SUBSET OF T4P. TO TEST THIS HYPOTHESIS, I WILL 1) DETERMINE IF TFPC INTERACTS WITH THE PILQ SECRETIN, 2) PERFORM STRUCTURE-FUNCTION ANALYSES OF TFPC, AND 3) INTERROGATING THE MECHANISM UNDERLYING REDUCED PILIATION IN TWELVE PILUS MUTANTS. THE PROPOSED RESEARCH IS SIGNIFICANT BECAUSE IT WILL DEFINE THE PREVIOUSLY UNCHARACTERIZED TFPC PROTEIN REQUIRED FOR PILIATION AND LINK ITS ACTIVITY TO OTHER PILUS COMPONENTS. IT WILL ALSO PROVIDE IMPORTANT INFORMATION FOR OTHER SPECIES T4P SYSTEMS AND POSSIBLY SOME TYPE II SECRETION SYSTEMS. THESE STUDIES EXPAND OUR CURRENT UNDERSTANDING OF T4P ARCHITECTURE AS WELL AS THE COMPONENTS GOVERNING MECHANISMS OF EXTENSION AND RETRACTION, PROVIDING A FOUNDATION FOR FUTURE STUDIES ON THE T4P DYNAMICS ESSENTIAL FOR COLONIZATION AND VIRULENCE. LONG-TERM, THE PROPOSED RESEARCH COULD PROVIDE NEW TARGETS FOR TREATING GONORRHEA INFECTIONS.