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Microbiology 146 (2000), 2919-2928
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Microbiology (2000), 146, 2919-2928.
© 2000 Society for General Microbiology

Genetics and Molecular Biology

Identification of DNA-binding proteins involved in regulation of expression of the Streptomyces aureofaciens sigF gene, which encodes sporulation sigma factor {sigma}F

D. Homerová1, B. Sevcíková1, O. Sprusanský1 and J. Kormanec1

Institute of Molecular Biology, Slovak Academy of Sciences, Dubravská cesta 21, 842 51 Bratislava, Slovak Republic1

Author for correspondence: J. Kormanec. Tel: +421 7 5941 2432. Fax: +421 7 5477 2316. e-mail: umbijkor{at}savba.sk

Expression of the sigF gene encoding a sporulation-specific sigma factor, {sigma}F, in Streptomyces aureofaciens is restricted only to sporulation. Gel mobility-shift assays using protein fractions from different developmental stages of S. aureofaciens revealed two different putative proteins specifically bound to the sigF promoter region: a protein (designated RsfA) present in young substrate mycelium, and a protein (designated RsfB) present in the course of sporulation. Based on the characteristic profiles of their appearance during differentiation, RsfA might be a repressor and RsfB an activator of sigF expression. The location of a specific binding site of the repressor-like protein (RsfA) was determined by gel mobility-shift assays of promoter deletion fragments and by DNase I footprinting analysis. The binding site mapped from nucleotides -87 to -25 relative to the transcription start point of the sigF promoter, and overlapped the -35 promoter region. Given the dependence of sigF expression upon whiH, the putative sporulation transcription factor WhiH was overproduced in Escherichia coli and used in the mobility-shift assays with the sigF promoter. However, no specific binding was detected, indicating an indirect dependence of sigF upon whiH.

Keywords: Streptomyces aureofaciens, differentiation, repressor, promoter, sigma factor

Abbreviations: tsp, transcription start point






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