Developmental gene expression in Bacillus subtilis crsA47 mutants reveals glucose-activated control of the gene for the minor sigma factor sigma(H) - PubMed (original) (raw)

Developmental gene expression in Bacillus subtilis crsA47 mutants reveals glucose-activated control of the gene for the minor sigma factor sigma(H)

L G Dixon et al. J Bacteriol. 2001 Aug.

Abstract

The presence of excess glucose in growth media prevents normal sporulation of Bacillus subtilis. The crsA47 mutation, located in the gene for the vegetative phase sigma factor (sigma(A)) results in a glucose-resistant sporulation phenotype. As part of a study of the mechanisms whereby the mutation in sigma(A) overcomes glucose repression of sporulation, we examined the expression of genes involved in sporulation initiation in the crsA47 background. The crsA47 mutation had a significant impact on a variety of genes. Changes to stage II gene expression could be linked to alterations in the expression of the sinI and sinR genes. In addition, there was a dramatic increase in the expression of genes dependent on the minor sigma factor sigma(H). This latter change was paralleled by the pattern of spo0H gene transcription in cells with the crsA47 mutation. In vitro analysis of RNA polymerase containing sigma(A47) indicated that it did not have unusually high affinity for the spo0H gene promoter. The in vivo pattern of spo0H expression is not predicted by the known regulatory constraints on spo0H and suggests novel regulation mechanisms that are revealed in the crsA47 background.

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Figures

FIG. 1

The effect of the crsA47 mutation on expression of kinA and spoVG. β-Galactosidase activities in strains carrying kinA-lacZ (A) or spo0VG-lacZ (B) fusions were measured as described in Materials and Methods. _T_0 represents the onset of sporulation. Strains contained the wild-type ςA gene (squares) or the crsA47 mutation (diamonds) and were grown in either SSM (open symbols) or SSMG (filled symbols). (A) JH12664 (kinA-lacZ) and GBS107 (crsA47 kinA-lacZ); (B) GBS110 (spoVG-lacZ) and GBS109 (crsA47 spoVG-lacZ).

FIG. 2

The effect of the crsA47 mutation on expression of spo0F. β-Galactosidase activities in strains carrying spo0F-lacZ fusions were measured as described in Materials and Methods. _T_0 represents the onset of sporulation. Strains contained the wild-type ςA gene (JH12862; squares) or the crsA47 mutation (GBS105; diamonds) and were grown in either SSM (open symbols) or SSMG (filled symbols).

FIG. 3

The effect of the crsA47 mutation on expression of spoIIA and spoIIG. β-Galactosidase activities in strains carrying spoIIA-lacZ (A) or spoIIG-lacZ (B) fusions were measured as described in Materials and Methods. _T_0 represents the onset of sporulation. Strains contained the wild-type ςA gene (squares) or the crsA47 mutation (diamonds), and were grown in either SSM (open symbols) or SSMG (filled symbols). (A) JH16124 (spoIIA-lacZ) and GBS106 (crsA47 spoIIA-lacZ); (B) JH16304 (spoIIG-lacZ) and GBS101 (crsA47 spoIIG-lacZ).

FIG. 4

The effect of the crsA47 mutation on expression of sinI and sinR. β-Galactosidase activities in strains carrying sinI-lacZ (A) or sinR-lacZ (B) fusions were measured as described in Materials and Methods. _T_0 represents the onset of sporulation. Strains contained the wild-type ςA gene (squares) or the crsA47 mutation (diamonds) and were grown in either SSM (open symbols) or SSMG (filled symbols). (A) GBS114 (sinI-lacZ) and GBS113 (crsA47 sinI-lacZ); (B) GBS116 (sinR-lacZ) and GBS115 (crsA47 sinR-lacZ).

FIG. 5

Effect of the crsA47 mutation on the expression of spo0H. β-Galactosidase activities in a strain carrying spo0H-lacZ fusions were measured as described in Materials and Methods. _T_0 represents the onset of sporulation. Strains contained the wild-type ςA gene (GBS151; squares) or the crsA47 mutation (GBS150; diamonds) and were grown in either SSM (open symbols) or SSMG (filled symbols).

FIG. 6

Effect of the crsA47 mutation on the expression of rapA and rapB. β-Galactosidase activities in strains carrying rapA-lacZ (A) or rapB-lacZ (B) fusions were measured as described in Materials and Methods. _T_0 represents the onset of sporulation. Strains contained the wild-type ςA gene (squares) or the crsA47 mutation (diamonds) and were grown in either SSM (open symbols) or SSMG (filled symbols). (A) JH12961 (rapA-lacZ) and GBS104 (crsA47 rapB-lacZ); (B) JH12866 (rapB-lacZ) and GBS103 (crsA47 rapB-lacZ).

FIG. 7

In vitro transcription activity of RNA polymerase isolated from JH642 and GBS10. In vitro transcription reactions were carried out using a control promoter (pUCA2trpA) (A) or a DNA fragment containing the spo0H gene promoter (B). Reaction mixtures contained a constant amount of RNA polymerase from either JH642 (wild type; squares) or GBS10 (crsA; circles) and increasing amounts of template. Transcription products were separated by electrophoresis through polyacrylamide gels containing 7 M urea. The level of product produced was determined using a Molecular Dynamics PhosphorImager and ImageQuant 1.0 software and is reported in arbitrary units.

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Developmental gene expression in Bacillus subtilis crsA47 mutants reveals glucose-activated control of the gene for the minor sigma factor sigma(H) - PubMed (original) (raw)