Cyclic AMP Receptor Protein Regulates cspE, an Early Cold-Inducible Gene, in Escherichia coli

• Published in: Journal of Bacteriology 2011-11, Vol.193 (22), p.6142-6151
• Main Authors: Uppal, Sheetal, Maurya, Svetlana R, Hire, Ramesh S, Jawali, Narendra
• Format: Article
• Language: English
• Subjects: adenosine monophosphate; Bacterial proteins; Bacteriology; Base Sequence; Binding Sites; Biological and medical sciences; cold; cold shock proteins; Cold Temperature; cyclic AMP; Cyclic AMP Receptor Protein - genetics; Cyclic AMP Receptor Protein - metabolism; DNA-directed RNA polymerase; E coli; Escherichia coli; Escherichia coli - genetics; Escherichia coli - metabolism; Escherichia coli Proteins - genetics; Escherichia coli Proteins - metabolism; Fundamental and applied biological sciences. Psychology; gel electrophoresis; Gene Expression Regulation, Bacterial; Gene Regulation; Genes; Heat-Shock Proteins - genetics; Heat-Shock Proteins - metabolism; melting; metabolism; Microbiology; Miscellaneous; Molecular Sequence Data; Mutation; nucleic acids; Promoter Regions, Genetic; Protein Binding; sequence analysis; sugars; T cell receptors; temperature; transcription (genetics)
• ISSN: 0021-9193; 1098-5530; 1067-8832
• DOI: 10.1128/JB.05728-11

cspE, a member of the cspA family of cold shock proteins in Escherichia coli, is an early cold-inducible protein. The nucleic acid melting ability and transcription antiterminator activity of CspE have been reported to be critical for growth at low temperature. Here, we show that the cyclic AMP receptor protein (CRP), a global regulator involved in sugar metabolism, upregulates cspE in E. coli. Sequence analysis of the cspE upstream region revealed a putative CRP target site centered at –61.5 relative to the transcription start. The binding of CRP to this target site was demonstrated using electrophoretic mobility shift assays. The presence of this site was shown to be essential for PcspE activation by CRP. Mutational analysis of the binding site indicated that the presence of an intact second core motif is more important than the first core motif for CRP-PcspE interaction. Based on the promoter architecture, we classified PcspE as a class I CRP-dependent promoter. This was further substantiated by our data demonstrating the involvement of the AR1 domain of CRP in PcspE transcription. Furthermore, the substitutions in the key residues of the RNA polymerase α-subunit C-terminal domain (α-CTD), which are important for class I CRP-dependent transcription, showed the involvement of 265 and 287 determinants in PcspE transcription. In addition, the deletion of crp led to a growth defect at low temperature, suggesting that CRP plays an important role in cold adaptation.