Identification and transcriptional analysis of a cold shock-inducible gene, cspA , in Streptomyces coelicolor A3(2)
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ORIGINAL PAPER
J. Kormanec á B. SÏevcÏõÂ kovaÂ
Identi®cation and transcriptional analysis of a cold shock-inducible gene, cspA, in Streptomyces coelicolor A3(2)
Received: 23 February 2000 / Accepted: 28 May 2000 / Published online: 4 August 2000 Ó Springer-Verlag 2000
Abstract Using the method for identi®cation of promoters recognized by the sporulation-speci®c sigma factor rF, we identi®ed a promoter, cspAp, in Streptomyces coelicolor, which showed similarity to the consensus sequence of Bacillus subtilis promoters recognized by the general stress-response sigma factor rB. cspAp directs expression of the cspA gene, which shows sequence similarity to members of the family of major cold shock proteins (CspA) from bacterial species. S1-nuclease mapping using RNA prepared from Escherichia coli containing a two-plasmid system, and from Streptomyces coelicolor at various developmental stages, identi®ed identical transcription start points in both species, corresponding to cspAp. However, the promoter was also active in a S. coelicolor sigF mutant. Transcriptional studies indicated that cspA is transcribed as a monocistronic mRNA. The level of cspA mRNA remains almost constant in all developmental stages, is dramatically increased after cold shock, and decreased after heat shock. Disruption of the S. coelicolor cspA gene did not aect growth or dierentiation at 11 °C or 30 °C. Key words Streptomyces coelicolor á Cold shock protein á cspA á SigF á S1 mapping
Introduction The cold-shock response is a physiological response of bacteria, as well as eukaryotic cells, to cold shock, which is characterized by the overexpression of speci®c proteins, the so-called cold shock proteins (CSPs; Jones and
Communicated by J. W. Lengeler J. Kormanec (&) á B. SÏevcÏõ kova Institute of Molecular Biology, Slovak Academy of Sciences, Dubravska cesta 21, 842 51 Bratislava, Slovak Republic E-mail: [email protected] Tel.: +421-7-59412432; Fax: +421-7-54772316
Inouye 1994). The major focus of recent work has been on Escherichia coli, where a novel small protein, termed the major cold shock protein, CspA, was identi®ed, that was induced upon cold shock (Goldstein et al. 1990). CspA functions as an RNA chaperone, which prevents the formation of secondary structures in RNA and thus ensures ecient translation of mRNA at low temperatures (Jiang et al. 1997). CspA homologs have been identi®ed in a wide variety of microorganisms (Graumann et al. 1997; Wouters et al. 1998). Streptomycetes undergo morphological dierentiation, involving the development of sporophyte or mycelial colonies (Chater 1993). As soil bacteria, they are exposed to various stress conditions including cold shock. Five proteins that cross-react with antibodies raised against E. coli CspA were identi®ed in Streptomyces aureofaciens; among these, two were inducible by exposure to cold (Mikulik et al. 1999). We have previously described a method for the identi®cation of Streptomyces promoters recognized by a particular Streptomyces r factor (NovaÂkova et al. 1998). The meth
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