PILP, A PILUS BIOGENESIS LIPOPROTEIN IN NEISSERIA-GONORRHOEAE, AFFECTS EXPRESSION OF PILQ AS A HIGH-MOLECULAR-MASS MULTIMER

Studies of gonococcal pilus biogenesis are fundamental to understandin g organelle structure/function relationships and identifying new appro aches to controlling disease, This area of research is also relevant t o elucidating the basic mechanisms of outer membrane translocation of macromolecules, which requires components highly related to those invo lved in type IV pilus expression. Previous studies have shown that pro ducts of several ancillary pil genes are required for organelle biogen esis but of these only PilQ, a member of the GspD protein family, is a component of the outer membrane, DNA sequencing of the region upstrea m of pilQ revealed the presence of two open reading frames (ORFs) whos e deduced polypeptides shared significant identities with proteins req uired for pilus expression in Pseudomonas aeruginosa and Pseudomonas s yringae, the genes for which are arrayed upstream of a gene encoding a PilQ homologue, Gonococcal mutants bearing transposon insertions in t hese ORFs were non-piliated and failed to express pilus-associated phe notypes, and the corresponding genes were designated pilO and pilP, Th e piliation defects in the mutants could not be ascribed to polarity o n distal pilQ expression as shown by direct measurement of PilQ antige n in those backgrounds and the use of a novel technique to create tand em duplications in the gonococcus (Gc) genome. As predicted by the pre sence of a consensus lipoprotein signal sequence, PilP expressed in bo th Escherichia coli and Gc could be labelled with [H-3]-palmitic acid. PilP(-) as well as PilQ(-) mutants shed PilC, a protein which facilit ates pilus assembly and is implicated in epithelial cell adherence, in a soluble form, Combined with the finding that levels of multimerized PilQ were greatly reduced in PilP(-) mutants, the results suggest tha t PilP is required for PilQ function and that PilQ and PilC may intera ct during the terminal stages of pilus biogenesis. The findings also s upport the hypothesis that the Gc PilQ multimer corresponds to a physi ologically relevant form of the protein required for pilus biogenesis.