Role of the Clp system in stress tolerance, biofilm formation and intracellular invasion in Porphyromonas gingivalis

Departments of Oral Biology, and Center for Molecular Microbiology, College of Dentistry, University of Florida, Gainesville, FL 32610; Department of Periodontics, Endodontics and Dental Hygiene, University of Louisville School of Dentistry, Louisville, KY 40292

^* To whom correspondence should be addressed. Email: rlamont{at}dental.ufl.edu.

	Abstract

Clp proteases and chaperones are ubiquitous among prokaryotes and eukaryotes, and in many pathogenic bacteria the Clp stress response system is also involved in regulation of virulence properties. In this study, the roles of ClpB, ClpC and ClpXP in stress resistance, homotypic and heterotypic biofilm formation, and intracellular invasion in the oral opportunistic pathogen P. gingivalis were investigated. Absence of ClpC and ClpXP, but not ClpB, resulted in diminished tolerance to high temperatures. Response to oxidative stress was not affected by the loss of any of the Clp proteins. The clpC and clpXP mutants demonstrated elevated monospecies biofilm formation, and the absence of ClpXP also enhanced heterotypic P. gingivalis-Streptococcus gordonii biofilm formation. All clp mutants adhered to gingival epithelial cells to the same level as wild type; however, ClpC and ClpXP were found to be necessary for entry into host epithelial cells. ClpB did not play a role in entry, but was required for intracellular replication and survival. ClpXP negatively regulated surface exposure of the minor fimbrial (Mfa) protein subunit of P. gingivalis that stimulates biofilm formation but interferes with epithelial cell entry. Collectively these results show that the Clp protease complex and chaperones control several processes that are important for the colonization and survival of P. gingivalis in the oral cavity.