Changes in the association between Bacillus subtilis RNA polymerase core and two specificity-determining subunits during transcription

The Bacillus subtilis RNA polymerase sigma subunit and the phage SPO1-coded gene 28 protein are responsible for selective binding of RNA polymerase to early and middle SPO1 promoters, respectively. The association of the RNA polymerase core with each of these subunits weakens during the elongation of RNA chains. Similar changes are known to be an essential part of the Escherichia coli RNA polymerase sigma cycle.     

Competition between sigma factors for core RNA polymerase.

The switch of RNA polymerase specificity from early to late promoters of bacteriophage T4 is achieved by substitution of host sigma factor, sigma 70, with the T4 induced factor, sigma gp55. However, overproduction of sigma gp55 from an expression vector is not detrimental to Escherichia coli growth. Direct competition binding assays demonstrate that sigma 70 readily displaces sigma gp55 from RNA polymerase and thereby reverses the promoter specificity of the enzyme. The displacement also occurs with the core enzyme modified by bacteriophage T4 infection. We postulate that an antagonist of sigma 70 should be formed in T4-infected cells to aid sigma gp55 in the early/late switch.     

Roles of genes 44, 50, and 51 in regulating gene expression and host takeover during infection of Bacillus subtilis by bacteriophage SPO1

We show that the products of SPO1 genes 44, 50, and 51 are required for the normal transition from early to middle gene expression during infection of Bacillus subtilis by bacteriophage SPO1; that they are also required for control of the shutoff of host DNA, RNA, and protein synthesis; and that their effects on host shutoff could be accounted for by their effects on the regulation of gene expression. These three gene products had four distinguishable effects in regulating SPO1 gene expression: (i) gp44-50-51 acted to restrain expression of all SPO1 genes tested, (ii) gp44 and/or gp50-51 caused additional specific repression of immediate-early genes, (iii) gp44 and/or gp50-51 stimulated expression of middle genes, and (iv) gp44 and/or gp50-51 stimulated expression of some delayed-early genes. Shutoff of immediate-early gene expression also required the activity of gp28, the middle-gene-specific sigma factor. Shutoff of host RNA and protein synthesis was accelerated by either the 44- single mutant or the 50(-)51(-) double mutant and more so by the 44(-)50(-)51(-) triple mutant. Shutoff of host DNA synthesis was accelerated by the mutants early in infection but delayed by the 44(-)50(-)51(-) triple mutant at later times. Although gp50 is a very small protein, consisting almost entirely of an apparent membrane-spanning domain, it contributed significantly to each activity tested. We identify SPO1 genes 41 to 51 and 53 to 60 as immediate-early genes; genes 27, 28, and 37 to 40 as delayed-early genes; and gene 52 as a middle gene.     

Interactions of Bacillus subtilis RNA polymerase with subunits determining the specificity of initiation. Sigma and delta peptides can bind simultaneously to core

The Bacillus subtilis RNA polymerase sigma 43 subunit and the phage SP82 encoded 28-kDa peptide are responsible for the binding of RNA polymerase to early and middle SP82 promoters, respectively. The delta peptide enhances the specificity of the interaction of B. subtilis RNA polymerase with these promoters. We have used sedimentation experiments to determine the effect of each of the three specificity factors, delta, sigma, and the 28-kDa peptide, on the binding of the other two factors to RNA polymerase core and the effect of NaCl on these binding equilibria. We show that sigma 43 and the 28-kDa peptide can each bind to RNA polymerase core at the same time as delta. Sigma 43 and the 28-kDa peptide have similar affinities to core at 0.1 M NaCl, but the 28-kDa peptide binds to core-delta more strongly than sigma 43. The implications of these findings with respect to the replacement of sigma 43 by the 28-kDa peptide and the mechanism of promoter search by B. subtilis RNA polymerase are discussed.     

Substitution of RNA-polymerase sigma-subunits and transcription regulation

Recent data on regulation of gene activity in bacteria by substitution of RNA polymerase sigma subunits are reviewed. The htpR gene which controls the switch-on of the Escherichia coli heat-shock protein synthesis codes for sigma 32 subunit. sigma 32-containing RNA polymerase transcribes the heat-shock genes in vitro from specific promoters of no use for RNA polymerase containing the major sigma 70 subunit. Several minor sigma subunits have been found in Bacillus subtilis vegetative cells, in addition to the major sigma 55 subunit, differing in the specificity of promoter recognition. Many B. subtilis genes are controlled by tandemly located promoters recognized by RNA polymerases carrying different sigma subunits. sigma 29 subunit is encoded by spoIIG gene and is probably involved in the regulation of sporulation. Specific sigma subunits for transcribing "middle" or "late" genes are encoded by a number of phages.