Replacement of vegetative sigma(A) by sporulation-specific sigma(F) as a component of the RNA polymerase holoenzyme in sporulating Bacillus subtilis

Soon after asymmetric septation in sporulating Bacillus subtilis cells, sig ma(F) is liberated in the prespore from inhibition by SpoIIAB. To initiate transcription from its cognate promoters, sigma(F) must compete with sigma( A), the housekeeping sigma factor in the predivisional cell, for binding to core RNA polymerase (E). To estimate the relative affinity of E for sigma( A) and sigma(F), we made separate mixtures of E with each of the two sigma factors, allowed reconstitution of the holoenzyme, and measured the concent ration of free E remaining in each mixture. The affinity of E for sigma(F) was found to be about 25-fold lower than that for sigma(A). We used quantit ative Western blotting to estimate the concentrations of E, sigma(A), and s igma(F) in sporulating cells. The cellular concentrations of E and sigma(A) were both about 7.5 mu M, and neither changed significantly during the fir st 3 h of sporulation. The concentration of sigma(F) was extremely low at t he beginning of sporulation, but it rose rapidly to a peak after about 2 h. At its peak, the concentration of sigma(F) was some twofold higher than th at of sigma(A). This difference in concentration cannot adequately account for the replacement of sigma(A) holoenzyme by sigma(F) holoenzyme in the pr espore, and it seems that some further mechanism-perhaps the synthesis or a ctivation of an anti-sigma(A) factor-must be responsible for this replaceme nt.