The RNA polymerase sigma subunit, σ(H), of Bacillus subtilis is required for the transcription of genes that are induced in late-growth cultures at high cell density, including genes that function in sporulation. The expression of σ(H)-controlled genes is repressed when nutrient broth sporulation medium (Difco sporulation medium [DSM]) is supplemented with high concentrations of glucose and glutamine (DSM-GG), preferred carbon and nitrogen sources of B. subtilis. Under these conditions, the pH of the DSM- GG medium decreases to ≃5. Raising the pH by the addition of morpholinepropanesulfonic acid (MOPS) or Tris-HCl (pH 7.5) results in a dramatic increase in the expression of lacZ fusions to σ(H)-dependent promoters. Correspondingly, the level of σ(H) protein was higher in cells of late-growth DSM-GG cultures treated with a pH stabilizer. When σ(H)-de- pendent gene expression was examined in cells bearing a mutation in abrB, encoding the transition state regulator that negatively controls genes transcribed by the σ(H) form of RNA polymerase, derepression was observed as well as an increase in medium pH. Reducing the pH with acetic acid resulted in repression, suggesting that AbrB was not functioning directly in pH- dependent repression but was required to maintain the low medium pH in DSM- GG. AbrB protein levels were high in late-growth, DSM-GG cultures but significantly lower when the pH was raised by Tris-HCl addition. An active tricarboxylic acid (TCA) cycle was required to obtain maximum derepression of σ(H)-dependent transcription, and transcription of the TCA cycle enzyme gene citB was repressed in DSM-GG but derepressed when the pH was artificially raised. The negative effect of low pH on σ(H)-dependent lacZ expression was also observed in unbuffered minimal medium and appeared to be exerted posttranslationally with respect to spo0H expression. However, the addition of amino acids to the medium caused pH-independent repression of both σ(H)- dependent transcription and spo0H-lacZ expression. These results suggest that spo0H transcription or translation is repressed by a mechanism responding to the availability of amino acids whereas spo0H is posttranslationally regulated in response to external pH.
ASJC Scopus subject areas
- Molecular Biology