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J. Biochem, 2003, Vol. 133, No. 3 295-302
© 2003 Japanese Biochemical Society

MOLECULAR BIOLOGY

Involvement of ClpX Protein in the Post-Transcriptional Regulation of a Competence Specific Transcription Factor, ComK Protein, of Bacillus subtilis

Hideaki Nanamiya1, Emiko Shiomi1, Mitsuo Ogura2, Teruo Tanaka2, Kei Asai3 and Fujio Kawamura+,1

1 Laboratory of Molecular Genetics, College of Science, Rikkyo (St. Paul’s) University, Toshima-ku, Tokyo 171-8501; 2 Department of Marine Science and Technology, Tokai University, Shimizu, Shizuoka 424-0902; and 3 Department of Biochemistry and Molecular Biology, Faculty of Science, Saitama University, Saitama, Saitama 338-8570

ComK protein of Bacillus subtilis positively regulates the transcription of several late competence genes as well as comK itself. We constructed a clpX disrupted mutant of B. subtilis and studied its effect on the regulation of ComK activation. When Pspac, which controls the comK gene in a multicopy plasmid, was induced by the addition of IPTG, comK transcripts were detected in both the clpX mutant and the wild type. However, the ComK protein could not be detected in the clpX disrupted mutant. To obtain further information, we constructed several comK-lacZ translational fusions covering different lengths of the comK gene, whose transcription is controlled by an IPTG inducible Pspac promoter. We found that both the expression of comK-lacZ directed ß-galactosidase and the accumulation of ComK-LacZ fused protein, derived from the fusion containing the entire comK open reading frame, were extremely reduced in the clpX mutant compared with the wild type, while the accumulation of comK-lacZ transcripts in the clpX mutant after the addition of IPTG was about half that in the clpX+ background. On the other hand, transcription, translation and activity of comK-lacZ were detected in both the clpX mutant and the wild type when the comK-lacZ fusion lacking the 3' region of the comK gene was induced. These results indicate that ClpX plays an important role in the regulation of ComK at the post-transcriptional level.

+ To whom correspondence should be addressed. Tel/Fax: +81-3-3985-2401, E-mail: kawamura{at}rikkyo.ne.jp


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