Binding of σ(A) and σ(B) to Core RNA Polymerase after Environmental Stress in Bacillus subtilis

In Bacillus subtilis, the alternative sigma factor σ(B) is activated in response to environmental stress or energy depletion. The general stress regulon under the control of σ(B) provides the cell with multiple stress resistance. Experiments were designed to determine how activated σ(B) replaces σ(A) as a constituent of the RNA polymerase holoenzyme. Studies of the transcription of the σ(A)-dependent stress gene clpE under σ(B)-inducing conditions showed that expression was higher in a sigB mutant background than in the wild type. The relative affinities of σ(A) and σ(B) for binding to the core RNA polymerase (E) were determined by means of indirect surface plasmon resonance. The results showed that the affinity of σ(B) for E was 60-fold lower than that of σ(A). Western blot analyses with antibodies against σ(A), σ(B), and E showed that, after exposure to ethanol stress, the concentration of σ(B) was only twofold higher than those of σ(A) and E. Thus, the concentration of σ(B) after stress is not high enough to compensate for its relatively low affinity for E, and it seems that additional mechanisms must be invoked to account for the binding of σ(B) to E after stress.