The Hybrid Cluster Protein Hcp Is Required for Growth with Nitrite and Survival with Host Cells.

Although the periodontal pathogen must withstand high levels of nitrosative stress while in the oral cavity, the mechanisms of nitrosative stress defense are not well understood in this organism. Previously we showed that the transcriptional regulator HcpR plays a significant role in defense, and here we further defined its regulon. Our study shows that (PG0893), a putative nitric oxide (NO) reductase, is the only gene significantly upregulated in response to nitrite (NO) and that this regulation is dependent on HcpR. An isogenic mutant deficient in is not able to grow with 200 μM nitrite, demonstrating that the sensitivity of the HcpR mutant is mediated through Hcp. We further define the molecular mechanisms of HcpR interaction with the promoter through mutational analysis of the inverted repeat present within the promoter. Although other putative nitrosative stress protection mechanisms present on the operon are also found in the genome, we show that their gene products play no role in growth of the bacterium with nitrite. As growth of the -deficient strain was also significantly diminished in the presence of a nitric oxide-producing compound, -nitrosoglutathione (GSNO), Hcp appears to be the primary means by which responds to NO-based stress. Finally, we show that Hcp is required for survival with host cells but that loss of Hcp has no effect on association and entry of into human oral keratinocytes.