Cloning and in vivo and in vitro regulation of cyclic AMP-dependent carbon starvation genes from Escherichia coli

P. H. Blum, S. B. Jovanovich, M. P. McCann, J. E. Schultz, S. A. Lesley, R. R. Burgess, A. Matin

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Abstract

The regulation of three Escherichia coli carbon starvation (cst) genes fused to lacZ was examined. Expression of these genes is induced by starvation for a carbon source. The role of carbon and cyclic AMP (cAMP) availability and of an altered-function crp mutation were investigated for their effect on cst expression in vivo. The experiments indicated that cAMP concentrations controlled the absolute expression of one cst fusion, but the other two cst fusions were dependent upon some component not present in exponentially growing cells under conditions of glucose excess, even when cAMP was added. To examine the regulation of these genes in further detail, the three cst::lacZ fusions were cloned on multicopy plasmids. All three cst::lacZ fusions retained their inducible regulatory phenotype in the multicopy state. Analysis of the expression of the cloned cst::lacZ fusions in an in vitro-coupled transcription-translation cell-free system demonstrated that the predominant promoter(s) present on each cloned DNA was dependent on σ70 for expression. In vitro cAMP titration curves indicated that this molecule was necessary and sufficient for the expression of one fusion but not sufficient for the second fusion, while the third fusion exhibited constitutive levels of expression in vitro. The results are discussed in the context of the E. coli carbon starvation response.

Original languageEnglish (US)
Pages (from-to)3813-3820
Number of pages8
JournalJournal of bacteriology
Volume172
Issue number7
DOIs
StatePublished - Jan 1 1990

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Starvation
Cyclic AMP
Organism Cloning
Carbon
Escherichia coli
Genes
In Vitro Techniques
Cell-Free System
Plasmids
Phenotype
Gene Expression
Glucose
Mutation
DNA

ASJC Scopus subject areas

  • Microbiology
  • Molecular Biology

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Cloning and in vivo and in vitro regulation of cyclic AMP-dependent carbon starvation genes from Escherichia coli. / Blum, P. H.; Jovanovich, S. B.; McCann, M. P.; Schultz, J. E.; Lesley, S. A.; Burgess, R. R.; Matin, A.

In: Journal of bacteriology, Vol. 172, No. 7, 01.01.1990, p. 3813-3820.

Research output: Contribution to journalArticle

Blum, P. H. ; Jovanovich, S. B. ; McCann, M. P. ; Schultz, J. E. ; Lesley, S. A. ; Burgess, R. R. ; Matin, A. / Cloning and in vivo and in vitro regulation of cyclic AMP-dependent carbon starvation genes from Escherichia coli. In: Journal of bacteriology. 1990 ; Vol. 172, No. 7. pp. 3813-3820.
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