Characterization of the transcriptional stimulatory properties of the Pseudomonas putida RapA protein
2013 (English)In: Biochimica et Biophysica Acta. Gene Regulatory Mechanisms, ISSN 1874-9399, Vol. 1829, no 2, 219-230 p.Article in journal (Refereed) Published
RNA polymerase-associated factors can significantly affect its performance at specific promoters. Here we identified a Pseudomonas putida RNA polymerases-associated protein as a homolog of Escherichia coli RapA. We found that P. putida RapA stimulates the transcription from promoters dependent on a variety of sigma-factors (sigma(70), sigma(S), sigma(54), sigma(32), sigma(E)) in vitro. The level of stimulation varied from 2- to 10-fold, with the maximal effect observed with the sigma(E)-dependent PhtrA promoter. Stimulation by RapA was apparent in the multi-round reactions and was modulated by salt concentration in vitro. However, in contrast to findings with E. coli RapA, P. putida RapA-mediated stimulation of transcription was also evident using linear templates. These properties of P. putida RapA were apparent using either E. coli- or P. putida-derived RNA polymerases. Analysis of individual steps of transcription revealed that P. putida RapA enhances the stability of competitor-resistant open-complexes formed by RNA polymerase at promoters. In vivo, P. putida RapA can complement the inhibitory effect of high salt on growth of an E. coli RapA null strain. However, a P. putida RapA null mutant was not sensitive to high salt. The in vivo effects of lack of RapA were only detectable for the sigma(E)-PhtrA promoter where the RapA-deficiency resulted in lower activity. The presented characteristics of P. putida RapA indicate that its functions may extend beyond a role in facilitating RNA polymerase recycling to include a role in transcription initiation efficiency.
Place, publisher, year, edition, pages
2013. Vol. 1829, no 2, 219-230 p.
RapA, Pseudomonas putida, Transcription factors, RNA polymerase
Cell and Molecular Biology
IdentifiersURN: urn:nbn:se:umu:diva-67395DOI: 10.1016/j.bbagrm.2012.11.008ISI: 000315063100004OAI: oai:DiVA.org:umu-67395DiVA: diva2:615889