Osmoregulation of the opuE proline transport gene from Bacillus subtilis:: contributions of the sigma A- and sigma B-dependent stress-responsive promoters

The opuE gene from Bacillus subtilis encodes a transport system (Opu E) for osmoprotective proline uptake and is expressed from two osmoregulated promoters: opuE P-l recognized by the vegetative sigma factor A (sigma(A)) and opuE P-2 dependent on the stress-induced transcription factor sigma B (sigma(B)). The contributions of these two promoters to osmoregulation of opuE were analysed. Genetic studies using chromosomal opuE-treA operon fusions revealed that opuE transcription is rapidly induced after an osmotic upshock. The strength of opuE expression is proportionally linked to the osmolarity of the growth medium. Deletion analysis of the opuE regulatory region identified a 330 bp DNA segment carrying all sequences required in cis for full and osmoregulated transcription. The proper rotational orientation of the upstream region present within this fragment was essential for the functioning of both opuE promoters. Mutant opoE-treA fusions with defects in either the sigma(A)- or the sigma(B)- dependent promoters revealed different contributions of these sequences to the overall osmoregulation of opuE. opuE P-2 (sigma(B)) activity increased transiently after an osmotic upshock and did not significantly contribute to the level of opuE expression in cells subjected to long-term osmotic stress. In contrast, transcription initiating from opuE P-l (sigma(A)) rose in proportion to the external osmolarity and was maintained at high levels. Moreover, both promoters exhibited a different response to the osmoprotectant glycine betaine in the medium. Our results suggest that at least two different signal transduction pathways operate in B. subtilis to communicate osmotic changes in the environment to the transcription apparatus of the cell.