Sigma Factor Activity Research Articles

The sigma factors of Bacillus subtilis.

The specificity of DNA-dependent RNA polymerase for target promotes is largely due to the replaceable sigma subunit that it carries. Multiple sigma proteins, each conferring a unique promoter preference on RNA polymerase, are likely to be present in all bacteria; however, their abundance and diversity have been best characterized in Bacillus subtilis, the bacterium in which multiple sigma factors were first discovered. The 10 sigma factors thus far identified in B. subtilis directly contribute to the bacterium's ability to control gene expression. These proteins are not merely necessary for the expression of those operons whose promoters they recognize; in many instances, their appearance within the cell is sufficient to activate these operons. This review describes the discovery of each of the known B. subtilis sigma factors, their characteristics, the regulons they direct, and the complex restrictions placed on their synthesis and activities. These controls include the anticipated transcriptional regulation that modulates the expression of the sigma factor structural genes but, in the case of several of the B. subtilis sigma factors, go beyond this, adding novel posttranslational restraints on sigma factor activity. Two of the sigma factors (sigma E and sigma K) are, for example, synthesized as inactive precursor proteins. Their activities are kept in check by "pro-protein" sequences which are cleaved from the precursor molecules in response to intercellular cues. Other sigma factors (sigma B, sigma F, and sigma G) are inhibited by "anti-sigma factor" proteins that sequester them into complexes which block their ability to form RNA polymerase holoenzymes. The anti-sigma factors are, in turn, opposed by additional proteins which participate in the sigma factors' release. The devices used to control sigma factor activity in B, subtilis may prove to be as widespread as multiple sigma factors themselves, providing ways of coupling sigma factor activation to environmental or physiological signals that cannot be readily joined to other regulatory mechanisms.

The dacF-spoIIA operon of Bacillus subtilis, encoding sigma F, is autoregulated

The spoIIA operon of Bacillus subtilis encodes sigma F and two proteins that may regulate sigma factor activity. High level induction of the tricistronic spoIIA operon occurs early during spore formation. At later times, the locus is cotranscribed with the upstream gene dacF, which encodes a putative DD-carboxypeptidase. In this study, the regulation of dacF-spoIIA transcription has been analyzed. Expression of a dacF-lacZ transcriptional fusion during sporulation required sigma F but not the later-expressed sporulation-associated sigma factors. Induction of sigma F synthesis during vegetative growth caused expression of dacF-lacZ fusions. The dacF-spoIIA promoter sequence is similar to sequences of previously identified sigma F promoters. It is concluded that dacF-spoIIA is transcribed by E sigma F. We present evidence that dacF-spoIIA is also transcribed by E sigma G, as is the case for the three other promoters known to be transcribed by E sigma F.

TheStaphylococcus aureuschromsomal geneplacC, indentified by mutations amplifying plasmid pT181, encodes a sigma factor

The Staphylococcus aureus chromosomal gene plaC, identified by mutations such as plaC1 that lead to the amplification of plasmid pT181, has been cloned and sequenced. The plaC gene encodes a protein with high similarity (79% identity) with the vegetative sigma factor of Bacillus subtilis, sigA, suggesting that it acts as an RNA polymerase sigma factor in S.aureus. The plaC1 mutation was found to be a C to T transition leading to a proline to serine substitution at amino acid residue 209 of the protein. In other sigma factors this region of the protein is involved in specific recognition of the -10 promoter sequence. The change in sigma factor activity due to this mutation is characterized by its strict specificity for a limited number of promoters and the rather high amplitude of the effect.

MicroReview Cascades of sigma factors revisited

Programmed gene expression during the process of endospore formation in Bacillus subtilis is governed by the successive appearance of five developmental sigma factors. These sigma factors are encoded by genes in which mutations arrest sporulation at a defined stage. These genes are turned on sequentially and depend for their own transcription on the activity of a previously synthesized sigma factor. Superimposed on the regulation of synthesis of the sigma factors are post-transcriptional control mechanisms that couple the activation of the developmental sigma factors to the course of sporulation. Here we review evidence indicating that these developmental transcription factors comprise a regulatory cascade in the order sigma H----sigma F----sigma E----sigma G----sigma K in which the activity of each sigma factor depends on the action of the preceding sigma factor in the cascade.

Effects of antibiotics on synthesis and persistence of sigma E in sporulating Bacillus subtilis

A potential regulatory link between the activation of a sporulation-specific sigma factor (sigma E) and forespore septum formation was investigated by treating Bacillus subtilis with inhibitors of protein or peptidoglycan synthesis and monitoring the consequences of these treatments on sigma E activation and septation. Western blot (immunoblot) and electron microscopic analyses revealed that both the formation of sigma E and septation were inhibited to a similar degree when either rifampin or chloramphenicol was added at different times before the second hour into sporulation but that penicillin preferentially blocked septation. We interpret these results as indicating that the syntheses of the gene products for both septation and sigma E activation occur at approximately the same time in development but that synthesis of an intact septum is unlikely to be a prerequisite for the formation of sigma E. We observed that penicillin could not only block septation but, depending on the time of its addition, could also inhibit both the activation of sigma E and the synthesis of its precursor. The basis of this effect is unknown, but it is not due to an overall disruption of protein synthesis. The incorporation of [35S] methionine by the sporulating cultures was unaffected by penicillin treatment. A time course study of the effects of rifampin and chloramphenicol treatments on sigma E levels revealed that both the synthesis of sigma E and its disappearance from sporulating cultures is inhibited by these antibiotics. This suggests that ongoing macromolecular synthesis is required for the turnover of sigma E.

Evolution of the transcription complex during sporulation of Bacillus subtilis.

Ribonucleic acid polymerase activity in partially purified extract of cells of Bacillus subtilis harvested at different times (t-1, to, t1, and t2) was studied by zone centrifugation. During the course of sporulation, vegetative sigma-factor activity decreased and the transcription complex lost some of its affinity for active sigma factor. The complex underwent a two-stage change in sedimentation value, from 14.5S in vegetative growth phase to a 13S species very early in sporulation to a 16S species at later times. Two SpoO mutants have been studied by zone centrifugation. One strain, a rifampin-resistant (RfmR) mutant, failed to show any modification of the transcription complex, whereas the other, a Rfms strain, underwent a partial evolution of the transcription complex after to.

Direct assay for sigma factor activity and demonstration of the loss of this activity during sporulation in Bacillus subtilis.

Summary. The change in template specificity of RNA polymerase, which in Bacillus subtilis takes place during the two hours following cessation of exponential growth (to--t2), has been confirmed with enzyme extracted in the presence of several protease inhibitors. With these enzyme preparations no alteration of the fl subunits could be detected. It is concluded that such an alteration cannot account for the loss by the enzyme of the ability to transcribe phage ~e DNA. An assay specific for vegetative sigma factor activity is described. When applied to crude extracts prepared in the presence of protease inhibitors, it was possible to demonstrate directly a loss in this activity, occurring between t o and t 2. This loss seems to account for the change in RNA polymerase template specificity which takes place during the same time period.