Replacement of vegetative sigmaA by sporulation-specific sigmaF as a component of the RNA polymerase holoenzyme in sporulating Bacillus subtilis.
Soon after asymmetric septation in sporulating Bacillus subtilis cells, sigmaF is liberated in the prespore from inhibition by SpoIIAB. To initiate transcription from its cognate promoters, sigmaF must compete with sigmaA, the housekeeping sigma factor in the predivisional cell, for binding to core RNA polymerase (E). To estimate the relative affinity of E for sigmaA and sigmaF, we made separate mixtures of E with each of the two sigma factors, allowed reconstitution of the holoenzyme, and measured the concentration of free E remaining in each mixture. The affinity of E for sigmaF was found to be about 25-fold lower than that for sigmaA. We used quantitative Western blotting to estimate the concentrations of E, sigmaA, and sigmaF in sporulating cells. The cellular concentrations of E and sigmaA were both about 7.5 microM, and neither changed significantly during the first 3 h of sporulation. The concentration of sigmaF was extremely low at the beginning of sporulation, but it rose rapidly to a peak after about 2 h. At its peak, the concentration of sigmaF was some twofold higher than that of sigmaA. This difference in concentration cannot adequately account for the replacement of sigmaA holoenzyme by sigmaF holoenzyme in the prespore, and it seems that some further mechanism-perhaps the synthesis or activation of an anti-sigmaA factor-must be responsible for this replacement.
Lord
M
M
Microbiology Unit, Department of Biochemistry, University of Oxford, Oxford OX1 3QU, United Kingdom.
Barillà
D
D
Yudkin
M D
MD
D001412:Bacillus subtilis / Q000502:physiology*; D001426:Bacterial Proteins / Q000032:analysis / Q000378:metabolism*; D012321:DNA-Directed RNA Polymerases / Q000032:analysis / Q000378:metabolism*; D020035:Holoenzymes; D011485:Protein Binding; D012808:Sigma Factor / Q000032:analysis / Q000378:metabolism*; D013171:Spores, Bacterial
D001426:Bacterial Proteins; C064971:FliA protein, Bacteria; D020035:Holoenzymes; D012808:Sigma Factor; C063505:RNA polymerase sigma A; D012321:DNA-Directed RNA Polymerases