Transcriptional regulation of a promoter in the men gene cluster of Bacillus subtilis

Abstract

The control of men gene expression during growth and sporulation of Bacillus subtilis was examined at the transcriptional level. Two different approaches were used. (i) Steady-state levels of men-specific mRNA were measured directly. (ii) A men'-lacZ gene fusion was constructed. In both cases, it was observed that men promoter activity was maximal at the onset of sporulation and declined soon thereafter. These kinetics were similar to the pattern of menaquinone accumulation previously observed. Expression from the men promoter was independent of the presence of the products of the spoOA and spoOH genes and was enhanced by addition of glucose and glutamine to the culture medium. DNA sequence analysis of the promoter region revealed a potential recognition site for the principal vegetative form of RNA polymerase but not for any of the known minor polymerase forms. The (Fig. 1). A men'-lacZ transcriptional gene fusion was created by cloning this fragment into the integrable lacZ fusion vector pDEB1 (31). The structure of this plasmid, pAI103, is shown in Fig. 2. Strains containing chromosomally integrated single copies of pAI103 were generated by transformation of appropriate recipients and selection for chloramphenicol resistance.

Sequence organization and regulation of the Bacillus subtilis menBE operon

“…A rapid decline in menB transcript levels began at T2 and continued through T4. The kinetics were similar to those found for menCDp-induced transcripts (21) and to the kinetics of MK accumulation during batch culture growth (10). Read-through transcription originating upstream from menBp also was detected, represented by a high-molecular-weight fragment corresponding to complete protection of men-specific sequences in the cRNA probe corresponding to a size of approximately 400 nucleotides (nt) (Fig.…”

“…However, the pathway in B. subtilis appears to be similar if not identical (20,29 deficient. This has permitted cloning of men-specific sequences and assignment of gene functions to specific segments of DNA.The cloning of menCDBE has been previously reported, along with preliminary analysis of a promoter (menCDp) (13,21,22). In subsequent experiments, we discovered that the previously cloned material was unable to rescue the menE310 mutation (11).…”

“…This could provide a system for control of MK formation. Previous work on menCDp included measurement of transcript accumulation during growth and sporulation (21). To map the approximate 5' termini of menB and menE transcripts and to compare the timing of expression from menB and menE promoters to that from menCDp, RNase protection assays were performed.…”

“…The cloning of menCDBE has been previously reported, along with preliminary analysis of a promoter (menCDp) (13,21,22). In subsequent experiments, we discovered that the previously cloned material was unable to rescue the menE310 mutation (11).…”

Four additional genes in the sigB operon of Bacillus subtilis that control activity of the general stress factor sigma B in response to environmental signals

“…One such example is the menCD operon, which is involved in menaquinone biosynthesis. Like the sigB operon, menCD is transcribed during stationary phase from a glucoseinducible, A -like promoter (37). One consequence of growth in glucose is the progressive acidification of the medium due to the production of acetate and pyruvate (47), and the menCD promoter has been shown to be responsive to changes in extracellular pH (20).…”