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PMID: 10940009 Published · ppublish English Journal Article Research Support, Non-U.S. Gov't

In vivo and in vitro effects of (p)ppGpp on the sigma(54) promoter Pu of the TOL plasmid of Pseudomonas putida.

Journal of bacteriology ·Vol. 182 ·No. 17 ·2000-09-00 ·Pages 4711-8

Carmona M, Rodríguez MJ, Martínez-Costa O, De Lorenzo V

Abstract

The connection between the physiological control of the sigma(54)-dependent Pu promoter of the TOL plasmid pWW0 of Pseudomonas putida and the stringent response mediated by the alarmone (p)ppGpp has been examined in vivo an in vitro. To this end, the key regulatory elements of the system were faithfully reproduced in an Escherichia coli strain and assayed as lacZ fusions in various genetic backgrounds lacking (p)ppGpp or overexpressing relA. Neither the responsiveness of Pu to 3-methyl benzylalcohol mediated by its cognate activator XylR nor the down-regulation of the promoter by rapid growth were affected in relA/spoT strains to an extent which could account for the known physiological control that governs this promoter. Overexpression of the relA gene [predicted to increase intracellullar (p)ppGpp levels] did, however, cause a significant gain in Pu activity. Since such a gain might be the result of indirect effects, we resorted to an in vitro transcription system to assay directly the effect of ppGpp on the transcriptional machinery. Although we did observe a significant increase in Pu performance through a range of sigma(54)-RNAP concentrations, such an increase never exceeded twofold. The difference between these results and the behavior of the related Po promoter of the phenol degradation plasmid pVI150 could be traced to the different promoter sequences, which may dictate the type of metabolic signals recruited for the physiological control of sigma(54)-systems.

MeSH Terms
Amino Acids/pharmacology DNA, Bacterial DNA-Binding Proteins DNA-Directed RNA Polymerases/genetics Escherichia coli/drug effects Escherichia coli Proteins Gene Deletion Gene Expression Gene Expression Regulation, Bacterial Genes, Reporter Guanosine Pentaphosphate/genetics,metabolism Lac Operon Ligases/genetics Plasmids Promoter Regions, Genetic Pseudomonas putida/genetics,physiology RNA Polymerase Sigma 54 Sigma Factor/genetics Transcription, Genetic
Chemicals
Amino Acids DNA, Bacterial DNA-Binding Proteins Escherichia coli Proteins Sigma Factor casamino acids rpoN protein, E coli Guanosine Pentaphosphate DNA-Directed RNA Polymerases RNA Polymerase Sigma 54 Ligases guanosine 3',5'-polyphosphate synthetases
Authors & Affiliations
4 authors, click to expand affiliations / ORCID
Carmona M
Department of Environment, Universidad Europea CEES, Villaviciosa de Odón, 28670 Madrid, Spain.
Rodríguez M J
Martínez-Costa O
De Lorenzo V
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Article Info
Journal
Journal of bacteriology
Abbr.
J Bacteriol
ISSN
0021-9193
DOI
10.1128/JB.182.17.4711-4718.2000
Published
2000-09-00
Pages
4711-8
Language
English
Region
United States
NLM ID
2985120R
PMCID
PMC111345
Subset
IM
External Links
PubMed source DOI: 10.1128/JB.182.17.4711-4718.2000 PMC Full Text View journal details
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