| 1. |
- O'Neill, E, et al.
(författare)
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Aromatic ligand binding and intramolecular signalling of the phenol-responsive sigma54-dependent regulator DmpR.
- 1998
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Ingår i: Molecular Microbiology. - 0950-382X .- 1365-2958. ; 28:1
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Tidskriftsartikel (refereegranskat)abstract
- The Pseudomonas-derived sigma54-dependent regulator DmpR has an amino-terminal A-domain controlling the specificity of activation by aromatic effectors, a central C-domain mediating an ATPase activity essential for transcriptional activation and a carboxy-terminal D-domain involved in DNA binding. In the presence of aromatic effectors, the DmpR protein promotes transcription from the -24, -12 Po promoter controlling the expression of specialized (methyl)phenol catabolic enzymes. Previous analysis of DmpR has led to a model in which the A-domain acts as an interdomain repressor of DmpR's ATPase and transcriptional promoting property until specific aromatic effectors are bound. Here, the autonomous nature of the A-domain in exerting its biological functions has been dissected by expressing portions of DmpR as independent polypeptides. The A-domain of DmpR is shown to be both necessary and sufficient to bind phenol. Analysis of phenol binding suggests one binding site per monomer of DmpR, with a dissociation constant of 16 microM. The A-domain is also shown to have specific affinity for the C-domain and to repress the C-domain mediated ATPase activity in vitro autonomously. However, physical uncoupling of the A-domain from the remainder of the regulator results in a system that does not respond to aromatics by its normal derepression mechanism. The mechanistic implications of aromatic non-responsiveness of autonomously expressed A-domain, despite its demonstrated ability to bind phenol, are discussed.
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| 2. |
- Shingler, V, et al.
(författare)
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Direct regulation of the ATPase activity of the transcriptional activator DmpR by aromatic compounds.
- 1995
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Ingår i: Molecular Microbiology. - 0950-382X .- 1365-2958. ; 17:3
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Tidskriftsartikel (refereegranskat)abstract
- The NtrC-like regulator DmpR controls transcription from the dmp operon that encodes the enzymes for catabolism of phenol and some related aromatic compounds. DmpR activates transcription from the sigma 54-dependent dmp-operon promoter in the presence of pathway substrates or structural analogues in the growth medium. Using affinity-purified DmpR and a truncated derivative, we show here that aromatic compounds directly activate the ATPase activity of this protein in vitro, and that the amino-terminal domain represses this activity in the absence of an aromatic ligand. In order to dissect the activation process, derivatives of DmpR exhibiting single amino acid changes were isolated and their effector-dependence and specificity profiles were analysed in vivo. The mechanistic implications of the phenotypes of these mutants are discussed.
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| 3. |
- Shingler, V
(författare)
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Signal sensing by sigma 54-dependent regulators : derepression as a control mechanism.
- 1996
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Ingår i: Molecular Microbiology. - 0950-382X .- 1365-2958. ; 19:3
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Tidskriftsartikel (refereegranskat)abstract
- Transcription by RNA polymerase utilizing the alternative sigma factor beta 54 is regulated by a distinct class of positive activators designated the sigma 54-dependent family. The activities of these regulators are themselves modulated in response to a wide variety of environmental signals. Factors that modulate the expression or the activity of the regulatory protein in response to chemical and metabolic changes are ultimately responsible for determining the level of expression of sigma 54-dependent genes and hence the diverse bacterial functions that they encode. Many members of the sigma 54-dependent family are part of two-component sensor-response systems. This MicroReview emphasizes recent data concerning the activities of a distinct subgroup of the sigma 54-dependent regulators that directly sense and respond with transcriptional activation to the presence of small effector molecules in their environment. The functional consequences of effector activation in terms of regulation of the enzymatic (ATPase) activity of these transcriptional activators and interdomain interactions are discussed.
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| 4. |
- Sze, C C, et al.
(författare)
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The alarmone (p)ppGpp mediates physiological-responsive control at the sigma 54-dependent Po promoter.
- 1999
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Ingår i: Molecular Microbiology. - 0950-382X .- 1365-2958. ; 31:4
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Tidskriftsartikel (refereegranskat)abstract
- Transcription from the Pseudomonas-derived sigma 54-dependent Po promoter of the dmp operon is mediated by the aromatic-responsive regulator DmpR. However, physiological control is superimposed on this regulatory system causing silencing of the DmpR-mediated transcriptional response in rich media until the transition between exponential and stationary phase is reached. Here, the positive role of the nutritional alarmone (p)ppGpp in DmpR regulation of the Po promoter has been identified and investigated in vivo. Overproduction of (p)ppGpp in a Pseudomonas reporter system was found to allow an immediate transcriptional response under normally non-permissive conditions. Conversely (p)ppGpp-deficient Escherichia coli strains were found to be severely defective in DmpR-mediated transcription, demonstrating the requirement for this metabolic signal. A subset of mutations in the beta, beta' and sigma 70 subunits of RNA polymerase, which confer prototrophy on ppGpp0 E. coli, was also found to restore specific DmpR-mediated transcription from Po, suggesting that the metabolic signal is mediated directly through the sigma 54-RNA polymerase. These data provide a direct mechanistic link between the physiological status of the cell and expression from sigma 54 promoters.
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