PrfA mediates specific binding of RNA polymerase of Listeria monocytogenes to PrfA-dependent virulence gene promoters resulting in a transcriptionally active complex.
There is accumulating evidence that the coordinate transcription of the virulence genes in Listeria monocytogenes constitutes a very complex regulation mechanism which might require other factors in addition to PrfA. We previously described an unknown proteinaceous component from crude bacterial cell extracts, which, together with PrfA, formed a specific complex (CI) in electrophoretic mobility shift assays (EMSA) with an hly promoter probe. Here we identify the RNA polymerase (RNAP) of L. monocytogenes as an essential component of the CI complex. Addition of purified RNAP plus PrfA to the hly promoter probe allowed reconstitution of a complex migrating at the same height as CI. By using EMSA and DNaseI footprint experiments it could be shown that PrfA leads to an enhanced and specific binding of RNAP. Transcriptional activity of RNAP in vitro, using the actA promoter, was strictly dependent on PrfA.
Böckmann
R
R
Lehrstuhl für Mikrobiologie, Theodor-Boveri-Institut für Biowissenschaften der Universität Würzburg, Am Hubland, 97074 Würzburg, Germany.
Dickneite
C
C
Goebel
W
W
Bohne
J
J
D001426:Bacterial Proteins / Q000737:chemistry / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism*; D002847:Chromatography, Agarose; D012321:DNA-Directed RNA Polymerases / Q000378:metabolism*; D003850:Deoxyribonuclease I / Q000378:metabolism; D004586:Electrophoresis / Q000379:methods; D015964:Gene Expression Regulation, Bacterial; D006459:Hemolysin Factors / Q000235:genetics; D008089:Listeria monocytogenes / Q000235:genetics / Q000254:growth & development / Q000378:metabolism / Q000472:pathogenicity*; D008565:Membrane Proteins; D010454:Peptide Termination Factors; D011401:Promoter Regions, Genetic; D015534:Trans-Activators / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism*; D014158:Transcription, Genetic; D014774:Virulence / Q000235:genetics
D001426:Bacterial Proteins; D008565:Membrane Proteins; D010454:Peptide Termination Factors; D015534:Trans-Activators; C072980:actA protein, Listeria monocytogenes; D012321:DNA-Directed RNA Polymerases; D003850:Deoxyribonuclease I