Development of a two-part transcription probe to determine the completeness of temporal and spatial compartmentalization of gene expression during bacterial development

We have developed a two-part test, using the Bacillus subtilis sacB/SacY transcription antitermination system, to evaluate the completeness of temporal and spatial compartmentalization of gene expression during bacterial cell development. Transcription of sacY(1-55) (encoding a constitutively active form of the antiterminator, SacY) is directed by one promoter, whereas transcription of sacB'-'IacZ(the target of SacY action) is directed by the same or another promoter. To obtain beta -galactosidase activity, SacY(1-55) needs to be present when sacB'-'IacZ is being transcribed. We tested the system by analyzing the spatial compartmentalization of the activities of RNA polymerase a factors, which are tightly regulated during sporulation of B. subtilis: sigma (F) and then sigma (G) in the prespore, sigma (E) and then sigma (K) in the mother cell. We have confirmed that the activities of sigma (F) and sigma (E) are spatially compartmentalized. We have demonstrated that there is also sharp temporal compartmentalization, with little or no overlap in the activities of ar and sigma (G) or of sigma (E) and sigma (K). In contrast, we found no compartmentalization of the activity of the main vegetative factor, sigma (A), which continued to be active alongside all of the sporulation-specific or factors. We also found no temporal compartmentalization of expression of loci that are activated during the development of competent cells of B. subtilis, a developmental program distinct from spore formation. A possible mechanism to explain the temporal compartmentalization of sigma (F) and sigma (G) activities is that the anti-sigma factor SpollAB transfers from sigma (G) to sigma (F).