Mutations in the precursor region of a Bacillus subtilis sporulation sigma factor

Rong, Sing; Sonenshein, Abraham L.

doi:10.1128/jb.174.11.3812-3817.1992

Cited by 9 publications

(6 citation statements)

References 26 publications

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“…However, sequences immediately upstream from the spoIIM coding sequence establish the likelihood that the gene is transcribed by RNA polymerase associated with &--, as are spolID, spolIA, and spoIIID (17,23,43,53,58). Thus, spoIIM apparently belongs to a family of genes expressed in the mother cell compartment whose products are required for full activity of crG-associated RNA polymerase in the forespore compartment.…”

Section: Methodsmentioning

confidence: 99%

Physical and functional characterization of the Bacillus subtilis spoIIM gene

1993

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The spoIIM locus ofBacilus subtilis is the most recently discovered of six genetic loci in which mutations can prevent the synthesis of a normal asymmetric septum or prevent migration of the septal structure to engulf the forespore compartment of the sporangium. Ultrastructure studies of a spoIIM mutant confirmed a block prior to the completion of engulfment. Introduction of a spoIIM mutation into a panel of strains containing lacZ fusions belonging to different regulatory classes allowed us to determine that the spoIIM gene product is required for the efficient expression of genes transcribed by rG-associated RNA polymerase but is not required for the expression of Mr-controlled genes, including spoIIIG, which encodes crG. The results of complementation studies, gene disruption analysis, and DNA sequencing revealed that the spoIIM locus contains a single sporulation-essential gene encoding a polypeptide with a predicted molecular mass of 24,850 Da. The predicted spolIM gene product is highly hydrophobic and very basic, and it does not exhibit significant homology to sequence files in several major data bases.

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Section: Methodsmentioning

confidence: 99%

Physical and functional characterization of the Bacillus subtilis spoIIM gene

1993

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“…The region of the putative helix in which the changes were made was also chosen because it was an area of the pro sequence in which one of the Leu-+Pro mutations (Fig. 1, 202) which substantially destabilized pro-er' occurred (5,15). Having constructed the first insertion mutation, we then incorporated additional residues into the mutant sequence wheel.…”

Section: Constructionmentioning

confidence: 99%

“…The arrows indicate the residues altered by mutations and illustrate the amino acid changes that accompany the mutations. The sigE201 and sigE202 alleles (isolated by S. Rong and A. L. Sonenshein) were previously described (5,15). Each of the insertion mutations (Ti in sigE12 and Ti and T2 in sigE13) was generated by placing codons for the indicated amino acids within sigE at the sites depicted.…”

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confidence: 99%

Mutational analysis of the precursor-specific region of Bacillus subtilis sigma E

1992

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crE is a sporulation-specific sigma factor ofBacilus subtilis that is formed from an inactive precursor protein (pro-cE) by the removal of 27 to 29 amino acids from the pro-oE amino terminus. By using oligonucleotidedirected mutagenesis, sequential deletions were constructed in the precursor-specific region of sigE and analyzed for their effect on the gene product's activity, ability to accumulate, and susceptibility to conversion into mature &E. The results demonstrated that the first 17 residues of the pro sequence contribute to silencing the r-like activity of pro-cr and that the amino acids between positions 12 and 17 are also important for its conversion into cE. Deletions that remove 21 or more codons from sigE reduce crE activity in cells which carry it, presumably by affecting prodrE stability. A 26-codon deletion results in a gene whose product is not detectable in B. subtilis by either reporter gene activity or Western blot (immunoblot) assay. The primary structure as well as the size of the pro region of c1r contributes to the protein's stability. The placement of additional amino acids into the pro region reduces the cell's ability to accumulate pro-oE. Additional sigE mutations revealed that the amino acids normally found at the putative processing site(s) of pro-cr are not essential to the processing reaction; however, a Glu residue upstream of these sites (position 25) was found to be important for processing.These last results suggest that the pro-IrF processing apparatus does not recognize the actual site within procr.E at which cleavage occurs but rather sequence elements that are upstream of this site.Bacillus subtilis synthesizes at least four sporulationspecific sigma factors (yEs, oF, cG, and ci) that are critical for orchestrating the temporal and compartmental patterns of gene expression that occur throughout endospore formation (12). &rE is the most abundant of these regulatory proteins during the early stages of spore formation (3). If a cell fails to synthesize o--, its development proceeds to the point where the cell partitions itself into forespore and mother cell compartments (stage II) but fails to progress beyond that stage (11). Transcription of the operon that encodes &E (spoIIG [22]) begins at the onset of sporulation and requires the sporulation gene activator, SpoOA, for its expression (7,8, 171. In addition to transcriptional regulation, synthesis of oe is also controlled posttranslationally. The initial protein product of the or-structural gene (sigEl spoIIGB) is an inactive precursor form of cre (pro-&e) (11,23). Pro-oe is converted into &e by proteolytic activity that removes 27 to 29 amino acids from the pro-&e amino terminus (11,12). Mutants that fail to convert pro-&e into cE have the same Spo-phenotype as those with null mutations in &e itself (4). The activity that cleaves the pro sequence from aE is itself developmentally regulated (24). It appears approximately 1 h after the onset of pro-o-synthesis if the products of at least six spo genes (4) and the cell divisi...

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“…Two leucine residues are conserved in the deduced clostridial sequence (positions 11 and 16 in B. subtilis, 8 and 12 in C. acetobutylicum; Fig. 2), which were shown to be critical for preventing partial activity of unprocessed pro-o" E in B. subtilis [41]. In contrast to the extensive amino acid conservation in the protease processing site of the pro<r E proteins from bacilli and C. acetobutylicum, only V20, Y22, N25, and A27 of B. subtilis pro-o-r: are conserved in the processing sites of all three organisms [18,40,48].…”

Section: Organismmentioning

confidence: 99%

“…N-terminal amino acid sequences as determined by Edman degradation of the purified ~ E proteins are underlined in the respective proteins. Two leucine residues, known to allow o -E activity without proteolysis [41], are marked above the sequence by asterisks. Promoter recognition regions 2.4 and 4.2 are marked by thick lines above the sequence.…”

Section: Introductionmentioning

confidence: 99%

Sigma factor and sporulation genes in Clostridium

Sauer

1995

FEMS Microbiology Reviews

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The genus Clostridium, represented by Gram-positive, anaerobic, spore-forming bacteria, is well known for its clinical importance and considerable biotechnological potential. Recently, evidence for a functional role of the transcription factors sigma A, sigma E, sigma G, and sigma K in this genus was provided by cloning and sequencing these genes from C. acetobutylicum. In C. kluyveri, a partially sequenced open reading frame was found to encode the N terminus of the putative sigma factor L with significant similarity to members of the sigma 54 family. The identification of sequences with high similarity to the Bacillus sigma F (C. acetobutylicum), sigma H (several clostridial species), and sigma D (C. thermocellum)-controlled consensus promoters renders the existence of these transcription factors in clostridia very likely. These data are in agreement with information obtained by RNA transcript mapping (sigma A, sigma H), heterologous DNA hybridization (sigma D, sigma H), and immuno characterization of purified proteins (sigma A) from various clostridial species. Thus, the picture emerges that a fundamental similarity exists at the genetic level between the regulation of various cellular responses, in particular sporulation, in the genera Bacillus and Clostridium. The different induction patterns of sporulation in Bacillus spp. (nutrient starvation) and many clostridial species (cessation of growth or exposure to oxygen in the presence of excess nutrients) are most interestingly not reflected in the general regulatory features of this developmental process.

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Mutations in the precursor region of a Bacillus subtilis sporulation sigma factor

Cited by 9 publications

References 26 publications

Physical and functional characterization of the Bacillus subtilis spoIIM gene

Physical and functional characterization of the Bacillus subtilis spoIIM gene

Mutational analysis of the precursor-specific region of Bacillus subtilis sigma E

Sigma factor and sporulation genes in Clostridium

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