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A regulatory cascade in the induction of rhaBAD.
J Mol Biol. 1993 Nov 05; 234(1):87-98.JM

Abstract

The RhaS and RhaR regulatory proteins are encoded in the Escherichia coli L-rhamnose gene cluster. We used complementation analysis and DNA mobility shift assays to show that RhaR is not the direct activator of the L-rhamnose catabolic operon, rhaBAD. An in-frame deletion of rhaS (rhaS-rhaR+) eliminated expression from the rhaBAD promoter, pBAD, while overexpression of rhaS greatly speeded the normally slow induction of transcription from pBAD. Expression from pBAD in a coupled transcription-translation assay was only detected when rhaS+ DNA was added to allow synthesis of RhaS protein. RhaS thus appears to be the direct L-rhamnose-specific activator of rhaBAD expression. Deletion mapping located the binding site for the L-rhamnose-specific regulator to a region overlapping position -70 relative to the rhaBAD transcription start site. Deletion mapping and DNA mobility shift assays located a CRP binding site just upstream from the binding site for the L-rhamnose-specific regulator. Quantitative primer extension analysis showed that induction of both the rhaBAD and rhaSR messages was unusually slow, requiring 40 to 50 minutes to reach a steady-state level. Induction of rhaBAD apparently involves a regulatory cascade in which RhaR first induces rhaSR expression, then RhaS accumulates and induces rhaBAD expression.

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Publisher Full Text

Authors+Show Affiliations

Egan SM
Biology Department, Johns Hopkins University, Baltimore, MD 21218.
Schleif RF
No affiliation info available

MeSH

Bacterial ProteinsBase SequenceCarrier ProteinsCyclic AMP Receptor ProteinDNA PrimersDNA, BacterialDNA-Binding ProteinsEscherichia coli ProteinsGene Expression Regulation, BacterialGenes, BacterialGenes, RegulatorMolecular Sequence DataOperonPromoter Regions, GeneticRhamnoseTrans-ActivatorsTranscription FactorsTranscription, Genetic

Pub Type(s)

Journal Article
Research Support, U.S. Gov't, P.H.S.

Language

eng

PubMed ID

8230210

Citation

Egan, S M., and R F. Schleif. "A Regulatory Cascade in the Induction of RhaBAD." Journal of Molecular Biology, vol. 234, no. 1, 1993, pp. 87-98.
Egan SM, Schleif RF. A regulatory cascade in the induction of rhaBAD. J Mol Biol. 1993;234(1):87-98.
Egan, S. M., & Schleif, R. F. (1993). A regulatory cascade in the induction of rhaBAD. Journal of Molecular Biology, 234(1), 87-98.
Egan SM, Schleif RF. A Regulatory Cascade in the Induction of RhaBAD. J Mol Biol. 1993 Nov 5;234(1):87-98. PubMed PMID: 8230210.
* Article titles in AMA citation format should be in sentence-case
TY - JOUR T1 - A regulatory cascade in the induction of rhaBAD. AU - Egan,S M, AU - Schleif,R F, PY - 1993/11/5/pubmed PY - 1993/11/5/medline PY - 1993/11/5/entrez SP - 87 EP - 98 JF - Journal of molecular biology JO - J Mol Biol VL - 234 IS - 1 N2 - The RhaS and RhaR regulatory proteins are encoded in the Escherichia coli L-rhamnose gene cluster. We used complementation analysis and DNA mobility shift assays to show that RhaR is not the direct activator of the L-rhamnose catabolic operon, rhaBAD. An in-frame deletion of rhaS (rhaS-rhaR+) eliminated expression from the rhaBAD promoter, pBAD, while overexpression of rhaS greatly speeded the normally slow induction of transcription from pBAD. Expression from pBAD in a coupled transcription-translation assay was only detected when rhaS+ DNA was added to allow synthesis of RhaS protein. RhaS thus appears to be the direct L-rhamnose-specific activator of rhaBAD expression. Deletion mapping located the binding site for the L-rhamnose-specific regulator to a region overlapping position -70 relative to the rhaBAD transcription start site. Deletion mapping and DNA mobility shift assays located a CRP binding site just upstream from the binding site for the L-rhamnose-specific regulator. Quantitative primer extension analysis showed that induction of both the rhaBAD and rhaSR messages was unusually slow, requiring 40 to 50 minutes to reach a steady-state level. Induction of rhaBAD apparently involves a regulatory cascade in which RhaR first induces rhaSR expression, then RhaS accumulates and induces rhaBAD expression. SN - 0022-2836 UR - https://www.unboundmedicine.com/medline/citation/8230210/A_regulatory_cascade_in_the_induction_of_rhaBAD_ L2 - https://linkinghub.elsevier.com/retrieve/pii/S0022-2836(83)71565-2 DB - PRIME DP - Unbound Medicine ER -