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Journal of Bacteriology, September 2008, p. 6162-6169, Vol. 190, No. 18
0021-9193/08/$08.00+0     doi:10.1128/JB.00046-08
Copyright © 2008, American Society for Microbiology. All Rights Reserved.

Analysis of a Growth-Phase-Regulated Two-Component Regulatory System in the Periodontal Pathogen Treponema denticola{triangledown}

Jesse R. Frederick,1 Elizabeth A. Rogers,1 and Richard T. Marconi1,2*

Department of Microbiology and Immunology,1 Center for the Study of Biological Complexity, Medical College of Virginia at Virginia Commonwealth University, Richmond, Virginia 23298-06782

Received 10 January 2008/ Accepted 3 July 2008

Nothing is currently known regarding the global regulatory networks of Treponema denticola and other oral spirochetes. In this report, we assess the properties and potential phosphotransfer capability of a putative two-component regulatory system (TCS) of T. denticola that is formed by the products of open reading frames tde0032 (a sensor kinase) and tde0033 (a response regulator), henceforth designated AtcS and AtcR, respectively. Using PCR and DNA sequence analyses, atcS and atcR were demonstrated to be widely distributed and conserved among T. denticola isolates. Reverse transcription-PCR (RT-PCR) analyses revealed that these genes are cotranscribed and may also be expressed as part of a larger operon that includes several flanking genes. Analyses using 5' rapid amplification of cDNA ends identified the transcriptional start sites for these operons and provided evidence that some of these genes may be independently transcribed from internal promoters. Real-time RT-PCR and Western blot analysis revealed significant upregulation of atcRS during late-stage growth, indicating growth-phase-dependent expression. Lastly, the phosphorelay capability of the AtcRS system was assessed and demonstrated using recombinant proteins. AtcS was found to undergo autophosphorylation and to transfer phosphate to AtcR. These analyses represent the first description of a functional TCS in an oral spirochetes and provide insight into the transcriptional regulatory mechanisms of these important bacteria.

* Corresponding author. Mailing address: Department of Microbiology and Immunology, Medical College of Virginia at Virginia Commonwealth University, Richmond, VA 23298-0678. Phone: (804) 828-3779. Fax: (804) 828-9946. E-mail: marconi{at}vcu.edu

{triangledown} Published ahead of print on 11 July 2008.

Journal of Bacteriology, September 2008, p. 6162-6169, Vol. 190, No. 18
0021-9193/08/$08.00+0     doi:10.1128/JB.00046-08
Copyright © 2008, American Society for Microbiology. All Rights Reserved.





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