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The dnrO gene encodes a DNA-binding protein that regulates daunorubicin production in Streptomyces peucetius by controlling expression of the dnrN pseudo response regulator gene

School of Pharmacy¹ and Department of Bacteriology², University of Wisconsin, Madison, WI 53706, USA

Author for correspondence: C. Richard Hutchinson. Tel: +1 510 732 8400 ext. 219. Fax: +1 510 732 8401. e-mail: hutchinson{at}kosan.com

The dnrO gene is located adjacent to and divergently transcribed from the response regulator gene, dnrN, that activates the transcription of the dnrI gene, which in turn activates transcription of the daunorubicin biosynthesis genes in Streptomyces peucetius. Gene disruption and replacement of dnrO produced the dnrO::aphII mutant strain and resulted in the complete loss of daunorubicin biosynthesis. Suppression of the dnrO::aphII mutation by the introduction of dnrN or dnrI on a plasmid suggested that DnrO is required for the transcription of dnrN, whose product is known to be required for dnrI expression. These conclusions were supported by the effects of the dnrO mutation on expression of dnrO, dnrN and dnrI, as viewed by melC fusions to each of these regulatory genes. DnrO was overexpressed in Escherichia coli and the cell-free extract was used to conduct mobility shift DNA-binding assays. The results showed that DnrO binds specifically to the overlapping dnrN/dnrO_p1 promoter region. Thus, DnrO may regulate the expression of both the dnrN and dnrO genes.

Keywords: anthracycline antitumour antibiotic, biosynthesis, melanin gene fusions, transcriptional mapping

Abbreviations: DNR, daunorubicin; DXR, doxorubicin; HTH, helix–turn–helix; RHO, -rhodomycinone

^a Present address: Department of Bacteriology, University of Wisconsin, Madison, WI 53706, USA.

^b Present address: Kosan Biosciences, Inc., 3832 Bay Center Place, Hayward, CA 94545, USA.

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