Dissection of recognition determinants of Escherichia coli sigma32 suggests a composite -10 region with an 'extended -10' motif and a core -10 element.
Sigma32 controls expression of heat shock genes in Escherichia coli and is widely distributed in proteobacteria. The distinguishing feature of sigma32 promoters is a long -10 region (CCCCATNT) whose tetra-C motif is important for promoter activity. Using alanine-scanning mutagenesis of sigma32 and in vivo and in vitro assays, we identified promoter recognition determinants of this motif. The most downstream C (-13) is part of the -10 motif; our work confirms and extends recognition determinants of -13C. Most importantly, our work suggests that the two upstream Cs (-16, -15) constitute an 'extended -10' recognition motif that is recognized by K130, a residue universally conserved in beta- and gamma-proteobacteria. This residue is located in the alpha-helix of sigmaDomain 3 that mediates recognition of the extended -10 promoter motif in other sigmas. K130 is not conserved in alpha- and delta-/epsilon-proteobacteria and we found that sigma32 from the alpha-proteobacterium Caulobacter crescentus does not need the extended -10 motif for high promoter activity. This result supports the idea that K130 mediates extended -10 recognition. Sigma32 is the first Group 3 sigma shown to use the 'extended -10' recognition motif.
Koo
Byoung-Mo
BM
Department of Microbiology and Immunology, University of California at San Francisco, San Francisco, CA 94158, USA.
Rhodius
Virgil A
VA
Campbell
Elizabeth A
EA
Gross
Carol A
CA
D019943:Amino Acid Substitution; D001483:Base Sequence; D016935:Caulobacter crescentus / Q000235:genetics / Q000378:metabolism; D017124:Conserved Sequence; D004926:Escherichia coli / Q000235:genetics* / Q000378:metabolism; D029968:Escherichia coli Proteins / Q000235:genetics / Q000378:metabolism*; D015964:Gene Expression Regulation, Bacterial; D006360:Heat-Shock Proteins / Q000235:genetics / Q000378:metabolism*; D008969:Molecular Sequence Data; D016296:Mutagenesis; D010957:Plasmids; D011401:Promoter Regions, Genetic; D012808:Sigma Factor / Q000235:genetics / Q000378:metabolism*; D014158:Transcription, Genetic
D029968:Escherichia coli Proteins; D006360:Heat-Shock Proteins; D012808:Sigma Factor; C069774:heat-shock sigma factor 32