Transcriptional regulation of the cytR repressor gene of Escherichia coli: autoregulation and positive control by the cAMP/CAP complex

The Escherichia coli cytR‐encoded repressor protein (CytR) controls the expression of several genes involved in nucleoside and deoxynucleoside uptake and metabolism. The cytR promoter was identified by determining the transcriptional initiation site of the cytR gene. A chromosomal cytR‐lacZ+ operon fusion was isolated and used to study the regulation of cytR. We show that cytR expression is negatively controlled by the CytR protein and positively affected by the cAMP/CAP complex. Foot printing studies with purified CAP protein revealed two CAP binding sites upstream of the cytR promoter. A previousty described mutation (cytR*) in the cloned cytR gene, which results in the phenotypic suppression of a CytR operator mutation in the tsx P2 promoter, was analysed. DN A sequence analysis of the cytR* mutation revealed a G‐C to an A‐T base pair transition at position ‐34 bp relative to the translational initiation site of cytR. This point mutation activates a cryptic promoter that is stronger than the wild‐type cytR promoter and leads to overproduction of the CytR repressor.

[1]  J. Sambrook,et al.  Molecular Cloning: A Laboratory Manual , 2001 .

[2]  P. Valentin‐Hansen,et al.  CRP/cAMP‐ and CytR‐regulated promoters in Escherichia coli K12: the cdd promoter , 1989, Molecular microbiology.

[3]  M. Ehrmann,et al.  MalI, a novel protein involved in regulation of the maltose system of Escherichia coli, is highly homologous to the repressor proteins GalR, CytR, and LacI , 1989, Journal of bacteriology.

[4]  J. E. Leclerc,et al.  Mutational analysis of the lac regulatory region: second-site changes that activate mutant promoters. , 1989, Nucleic acids research.

[5]  W. Reznikoff,et al.  Pseudorevertants of a lac promoter mutation reveal overlapping nascent promoters. , 1989, Nucleic acids research.

[6]  B. Matthews,et al.  The helix-turn-helix DNA binding motif. , 1989, The Journal of biological chemistry.

[7]  K. Hammer,et al.  Purification and characterization of the deoR repressor of Escherichia coli. , 1989, The EMBO journal.

[8]  R. Rolfes,et al.  Escherichia coli gene purR encoding a repressor protein for purine nucleotide synthesis. Cloning, nucleotide sequence, and interaction with the purF operator. , 1988, The Journal of biological chemistry.

[9]  G. Weinstock,et al.  Transposition of λplacMu is mediated by the A protein altered at its carboxy-terminal end , 1988 .

[10]  E. D. Hyman.  A new method of sequencing DNA. , 1988, Analytical biochemistry.

[11]  J. Sturtevant,et al.  Scanning calorimetric study of the thermal unfolding of catabolite activator protein from Escherichia coli in the absence and presence of cyclic mononucleotides. , 1988, Biochemistry.

[12]  V. de Lorenzo,et al.  Fur (ferric uptake regulation) protein and CAP (catabolite-activator protein) modulate transcription of fur gene in Escherichia coli. , 1988, European journal of biochemistry.

[13]  A. Middendorf,et al.  Double negative and positive control of tsx expression in Escherichia coli , 1988, Journal of bacteriology.

[14]  C. Harley,et al.  Analysis of E. coli promoter sequences. , 1987, Nucleic acids research.

[15]  S T Cole,et al.  Inducible cephalosporinase production in clinical isolates of Enterobacter cloacae is controlled by a regulatory gene that has been deleted from Escherichia coli. , 1986, The EMBO journal.

[16]  P. Valentin‐Hansen,et al.  DNA‐protein recognition: demonstration of three genetically separated operator elements that are required for repression of the Escherichia coli deoCABD promoters by the DeoR repressor. , 1986, The EMBO journal.

[17]  P. Højrup,et al.  Nucleotide sequence of the CytR regulatory gene of E. coli K-12. , 1986, Nucleic acids research.

[18]  K. Hammer,et al.  Two operator sites separated by 599 base pairs are required for deoR repression of the deo operon of Escherichia coli. , 1985, The EMBO journal.

[19]  J. Singer,et al.  Identification of the Escherichia coli deoR and cytR gene products , 1985, Journal of bacteriology.

[20]  G. Winter,et al.  Improved oligonucleotide site-directed mutagenesis using M13 vectors. , 1985, Nucleic acids research.

[21]  R. Ebright,et al.  Molecular basis of DNA sequence recognition by the catabolite gene activator protein: detailed inferences from three mutations that alter DNA sequence specificity. , 1984, Proceedings of the National Academy of Sciences of the United States of America.

[22]  S. Short,et al.  Studies on deo operon regulation in Escherichia coli: cloning and expression of the cytR structural gene. , 1984, Gene.

[23]  L. Enquist,et al.  Experiments With Gene Fusions , 1984 .

[24]  G. Weinstock,et al.  Lambda placMu: a transposable derivative of bacteriophage lambda for creating lacZ protein fusions in a single step , 1984, Journal of bacteriology.

[25]  H. Buc,et al.  Cyclic AMP receptor protein: role in transcription activation. , 1984, Science.

[26]  Robert Entriken,et al.  Escherichia coli promoter sequences predict in vitro RNA polymerase selectivity , 1984, Nucleic Acids Res..

[27]  J. Messing,et al.  Construction of improved M13 vectors using oligodeoxynucleotide-directed mutagenesis. , 1983, Gene.

[28]  G. Hong,et al.  Buffer gradient gels and 35S label as an aid to rapid DNA sequence determination. , 1983, Proceedings of the National Academy of Sciences of the United States of America.

[29]  B. Bachmann.  Linkage map of Escherichia coli K-12, edition 7 , 1983, Microbiological reviews.

[30]  P. Valentin‐Hansen,et al.  Tandem CRP binding sites in the deo operon of Escherichia coli K‐12. , 1982, The EMBO journal.

[31]  H. Aiba,et al.  The structure of tandem regulatory regions in the deo operon of Escherichia coli K12 , 1982, The EMBO journal.

[32]  T. Silhavy,et al.  Genetic analysis of the ompB locus in Escherichia coli K-12. , 1981, Journal of molecular biology.

[33]  D. Galas,et al.  DNAse footprinting: a simple method for the detection of protein-DNA binding specificity. , 1978, Nucleic acids research.

[34]  F. Sanger,et al.  DNA sequencing with chain-terminating inhibitors. , 1977, Proceedings of the National Academy of Sciences of the United States of America.

[35]  F. Bolivar,et al.  Construction and characterization of new cloning vehicles. I. Ampicillin-resistant derivatives of the plasmid pMB9. , 1977, Gene.

[36]  W. Gilbert,et al.  A new method for sequencing DNA. , 1977, Proceedings of the National Academy of Sciences of the United States of America.

[37]  M. Casadaban,et al.  Regulation of the regulatory gene for the arabinose pathway, araC. , 1976, Journal of molecular biology.

[38]  M. Casadaban,et al.  Transposition and fusion of the lac genes to selected promoters in Escherichia coli using bacteriophage lambda and Mu. , 1976, Journal of molecular biology.

[39]  U. K. Laemmli,et al.  Cleavage of Structural Proteins during the Assembly of the Head of Bacteriophage T4 , 1970, Nature.

[40]  M. Ashburner.  A Laboratory manual , 1989 .

[41]  S. Takeshita,et al.  High-copy-number and low-copy-number plasmid vectors for lacZ alpha-complementation and chloramphenicol- or kanamycin-resistance selection. , 1987, Gene.

[42]  P. Valentin‐Hansen,et al.  Long-range cooperativity between gene regulatory sequences in a prokaryote , 1987, Nature.

[43]  P. Valentin-Hansen.  DNA Sequences Involved in Expression and Regulation of deoR -, cytR - and cAMP/CRP-Controlled Genes in Escherichia Coli , 1985 .

[44]  C. Blanco,et al.  The use of gene fusions to study the expression of uidR, a negative regulatory gene of Escherichia coli K-12. , 1985, Gene.

[45]  M. Schwartz,et al.  Positive control of transcription initiation in bacteria. , 1984, Annual review of genetics.

[46]  J. Messing.  New M13 vectors for cloning. , 1983, Methods in enzymology.

[47]  J. Messing.  [2] New M13 vectors for cloning , 1983 .

[48]  J. Reeve.  Use of minicells for bacteriophage-directed polypeptide synthesis. , 1979, Methods in enzymology.

[49]  M. Schulman.  Purines and Pyrimidines , 1961 .

[50]  W. Fearon.  PURINES AND PYRIMIDINES , 1940 .