Porphyromonasgingivalis, a gram-negative anaerobic bacterium, is a recognized periodontopathogen. Although it mainly inhabits anaerobic periodontal pockets it also is present in other niches of the oral cavity enroute to the periodonta pockets as well as it can access distant parts of the body such as cardiovascular system and heart tissues. Finally, it can invade and survive within host cells. In order to adapt to the varios niches the organism is expected to have efficient regulatory mechanisms allowing it to rapidly alter gene and ultimately protein expression in response to the various challenges imposed by the host environments. Riboregulation has emerged as a significant mechanism of post-transciptional regulation in bacteria. The RNA chaperone, Hfq, has been shown to play an indispensable role in this process by binding sRNAs and stabilizing the sRNA-mRNA duplexes. However, at least 50% of bacteria lack Hfq indicating the presence of other novel mechanisms mediating sRNA regulation. Using bioinformatics approaches we identified a candidate for an RNA-binding protein, designated here as Pgr, in P.gingivalis. Our preliminary data demonstrate that the protein binds RNA and plays a role in survival of the bacterium with nitrosative stress as well as in the presence of host cells. Furthermore, we noted significant changes in RNA levels resulting from deletion of Pgr. Based on the preliminary data we hypothesize that Pgr is a novel RNA chaperone and plays a major role in post-transcriptional regulation in P. gingivalis. As Pgr function is relevant for full virulence of the bacterium, it will serve as an excellent target for development of antimicrobial strategies. To gain an insight into the mechanism of regulation by the protein we will define the transcriptome and proteome affected by the absence of the protein as well as identify the RNAs interacting with Pgr. We expect that once completed this work will provide insight into mechanism of action of this protein and ultimately will lead to design of strategies interfering with its action and thus reducing virulence of P. gingivalis. Finally, we predict that this work will shed light on the mechanisms of riboregulation in other Bacteroides- related bacteria such as Prevotella intermedia, Tannerella forsythensis, Bacteroides fragilis, B. thetaiotaomicron and Capnocytophaga hutchinsonii.
The long-term objective of this application is to identify and characterize the riboregulatory mechanisms of anaerobic bacteria. Oral periodontal pathogens have to respond to constantly changing environmental conditions, however, so far the regulatory mechanisms governing the protein expression are poorly understood. Our work is expected to provide clues how the bacterium tolerates various environments and ultimately, we predict that this work will lead to identification of targets for the design of novel and specific therapeutic agents for treatment of periodontal diseases.
|Wunsch, Christopher M; Lewis, Janina P (2015) Porphyromonas gingivalis as a Model Organism for Assessing Interaction of Anaerobic Bacteria with Host Cells. J Vis Exp :e53408|