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Journal of Bacteriology, September 2005, p. 6341-6353, Vol. 187, No. 18
0021-9193/05/$08.00+0     doi:10.1128/JB.187.18.6341-6353.2005
Copyright © 2005, American Society for Microbiology. All Rights Reserved.

Three Chymotrypsin Genes Are Members of the AdpA Regulon in the A-Factor Regulatory Cascade in Streptomyces griseus

Ayami Tomono, Yisan Tsai, Yasuo Ohnishi, and Sueharu Horinouchi^*

Department of Biotechnology, Graduate School of Agriculture and Life Sciences, University of Tokyo, Bunkyo-ku, Tokyo 113-8657, Japan

Received 1 June 2005/ Accepted 30 June 2005

AdpA is a key transcriptional activator in the A-factor regulatory cascade in Streptomyces griseus, activating a number of genes required for secondary metabolism and morphological differentiation. Of the five chymotrypsin-type serine protease genes, sprA, sprB, and sprD were transcribed in response to AdpA, showing that these protease genes are members of the AdpA regulon. These proteases were predicted to play the same physiological role, since these protease genes were transcribed in a similar time course during growth and the matured enzymes showed high end-to-end similarity to one another. AdpA bound two sites upstream of the sprA promoter approximately at positions –375 and –50 with respect to the transcriptional start point of sprA. Mutational analysis of the AdpA-binding sites showed that both AdpA-binding sites were essential for transcriptional activation. AdpA bound a single site at position –50 in front of the sprB promoter and greatly enhanced the transcription of sprB. The AdpA-binding site at position –40 was essential for transcription of sprD, although there was an additional AdpA-binding site at position –180. Most chymotrypsin activity excreted by S. griseus was attributed to SprA and SprB, because mutant sprAB, having a deletion in both sprA and sprB, lost almost all chymotrypsin activity, as did mutant adpA. Even the double mutant sprAB and triple mutant sprABD grew normally and developed aerial hyphae and spores over the same time course as the wild-type strain.

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* Corresponding author. Mailing address: Department of Biotechnology, Graduate School of Agriculture and Life Sciences, The University of Tokyo, Bunkyo-ku, Tokyo 113-8657, Japan. Phone: 81 3 5841 5123. Fax: 81 3 5841 8021. E-mail: asuhori{at}mail.ecc.u-tokyo.ac.jp.

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Journal of Bacteriology, September 2005, p. 6341-6353, Vol. 187, No. 18
0021-9193/05/$08.00+0     doi:10.1128/JB.187.18.6341-6353.2005
Copyright © 2005, American Society for Microbiology. All Rights Reserved.

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