title: The dynamic protein partnership of RNA polymerase in Bacillus subtilis.
abstract: In prokaryotes, transcription results from the activity of a 400 kDa RNA polymerase (RNAP) protein complex composed of at least five subunits (2α, β, β', ω). To ensure adequate responses to changing environmental cues, RNAP activity is tightly controlled by means of interacting regulatory proteins. Here, we report the affinity-purification of the Bacillus subtilis RNAP complexes from cells in different growth states and stress conditions, and the quantitative assessment by mass spectrometry of the dynamic changes in the composition of the RNAP complex. The stoichiometry of RNA polymerase was determined by a comparison of two mass spectrometry-based quantification methods: a label-based and a label-free method. The validated label-free method was then used to quantify the proteins associated with RNAP. The levels of sigma factors bound to RNAP varied during growth and exposure to stress. Elongation factors, helicases such as HelD and PcrA, and novel unknown proteins were also associated with RNAP complexes. The content in 6S RNAs of purified RNAP complexes increased at the onset of the stationary phase. These quantitative variations in the protein and RNA composition of the RNAP complexes well correlate with the known physiology of B. subtilis cells under different conditions.
authors.affiliation: INRA,UMR 1319 Micalis, Jouy-en-Josas, France. olivier.delumeau@jouy.inra.fr
authors.forename: Olivier
authors.initials: O
authors.lastname: Delumeau
authors.forename: François
authors.initials: F
authors.lastname: Lecointe
authors.forename: Jan
authors.initials: J
authors.lastname: Muntel
authors.forename: Alain
authors.initials: A
authors.lastname: Guillot
authors.forename: Eric
authors.initials: E
authors.lastname: Guédon
authors.forename: Véronique
authors.initials: V
authors.lastname: Monnet
authors.forename: Michael
authors.initials: M
authors.lastname: Hecker
authors.forename: Dörte
authors.initials: D
authors.lastname: Becher
authors.forename: Patrice
authors.initials: P
authors.lastname: Polard
authors.forename: Philippe
authors.initials: P
authors.lastname: Noirot
mesh_terms: D000345:Affinity Labels; D001412:Bacillus subtilis / Q000201:enzymology* / Q000235:genetics; D001426:Bacterial Proteins / Q000032:analysis / Q000235:genetics / Q000378:metabolism*; D002846:Chromatography, Affinity; D012321:DNA-Directed RNA Polymerases / Q000032:analysis / Q000378:metabolism*; D004591:Electrophoresis, Polyacrylamide Gel; D046912:Multiprotein Complexes / Q000032:analysis / Q000378:metabolism; D021122:Protein Subunits / Q000032:analysis / Q000378:metabolism; D040901:Proteomics; D012329:RNA, Bacterial / Q000032:analysis / Q000378:metabolism; D022661:RNA, Untranslated; D012808:Sigma Factor / Q000032:analysis / Q000378:metabolism; D053719:Tandem Mass Spectrometry
chemical_list: C411065:6S RNA; D000345:Affinity Labels; D001426:Bacterial Proteins; D046912:Multiprotein Complexes; D021122:Protein Subunits; D012329:RNA, Bacterial; D022661:RNA, Untranslated; D012808:Sigma Factor; D012321:DNA-Directed RNA Polymerases