Abstract
ABSTRACT Background: Bacteria use two-component signal transduction systems (among others) to perceive and respond to environmental changes. Within the genus Porphyromonas, we observed degeneration of these systems, as exemplified by the loss of RprX, the sensor kinase partner of the RprY. Objective: The purpose of this study was to investigate modulation of RprY function by acetylation. Design: The transcriptional activity of the rprY-pat genes were measured by RT-PCR and 5ʹ-RACE. The acetylation of RprY were detected by western blotting. Electromobility shift and in vitro ChIP assays were used to measure the DNA binding activity of RprY. The expression of RprY target genes was measured by qRT-PCR. Effects of acetylation on phosphorylation of RprY were measured by Phos-tag gels. Results: The rprY gene is cotranscribed with pat. RprY is acetylated in vivo, and autoacetylated in vitro in a reaction that is enhanced by Pat; the CobB sirtuin deacetylates RprY. Acetylation reduced the DNA binding of RprY. Induced oxidative stress decreased production of RprY in vivo, increased its acetylation and increased expression of nqrA. Conclusions: We propose that to compensate for the loss of RprX, P. gingivalis has evolved a novel mechanism to inactivate RprY through acetylation.
Highlights
Porphyromonas gingivalis, a Gram-negative oral anaerobe, is associated with adult periodontitis which affects almost 50% of the US population [1]
In two-component systems, histidine kinase (HK) and cognate regulator protein (RR) genes are most often encoded in an operon; there are examples of the genes located at different loci in the genome
To determine whether rprY and pat are transcribed as a monocistronic message, we performed RT-PCR with RNA from strain ATCC 33277 grown under standard broth culture conditions
Summary
Porphyromonas gingivalis, a Gram-negative oral anaerobe, is associated with adult periodontitis which affects almost 50% of the US population [1]. Anaerobes such as P. gingivalis respond to environmental signals, some yet unknown, which enhance growth of the organism and induce virulence factors that lead to gingivitis and periodontal disease. The ability of bacteria to respond to environmental changes is in part regulated by two-component signal transduction systems [2]. The simplest system involves a sensor histidine kinase (HK), most often a transmembrane protein with its N-terminus located in the periplasm to monitor environmental changes. In a phospho-relay, the HK functions as phosphoryl donor to a conserved aspartate residue in the cognate response regulator protein (RR), inducing change to an active conformation that, when acting as a transcription factor, can bind to target gene promoter sequences. Bacteria use two-component signal transduction systems (among others) to perceive and respond to environmental changes.
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