Developmental expression of three proteins from the first gene of the RNA polymerase sigma 43 operon of Bacillus subtilis

Wang, Lin‐Fa; Doi, Roy H.

doi:10.1128/jb.169.9.4190-4195.1987

Cited by 17 publications

(11 citation statements)

References 29 publications

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“…However, the relationships between growth, cell division, and sporulation are well established (4,8,17,25,30). In addition, we have found that two sporulation mutants (spoOB and spoOE) of B. subtilis are apparently defective in the tumover of the methyl group(s) on P-40 (K. J.…”

Section: Discussionmentioning

confidence: 86%

“…Sporulation is initiated in bacilli during the last generation of growth (4,24) and is characterized by a cascade of sigma factors or transcriptional regulators or both, but the mechanism by which this cascade is initiated is unknown (16,17,30,31). The regulation of these early events in sporulation is under the control of a set of genes (the spoO genes in Bacillus subtilis) whose products have not been characterized, although at least one (SpoOH) is thought to be a sigma factor (7).…”

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confidence: 99%

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Nutrient-stimulated methylation of a membrane protein in Bacillus licheniformis

et al. 1988

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When nitrogen-starved vegetative cells of Bacillus licheniformis A5 were presented with a good nitrogen source in the presence of chloramphenicol and methyl-labeled methionine, a 40-kilodalton (kDa) protein was found to be reversibly methylated, with a half-life of approximately 10 to 15 min. The 40-kDa protein was strongly methylated in response to the addition of ammonia, glutamine, or sodium glutamate (nitrogen sources that produce generation times of <90 min) but was very poorly methylated in the absence of a nitrogen source or in the presence of potassium glutamate or histidine (generation times of >150 min). The methylated protein was found to be membrane associated, but the methylation reaction did not appear to be related to chemotaxis, because the spectrum of nutrients that promoted methylation was different from that which prompted a chemotactic response. In addition, the methyl residue on the 40-kDa protein was found to be alkali stable. Approximately 180 to 640 molecules of the methylated protein were found per cell. The characteristics of this methylated protein were consistent with the hypothesis that the reversible methylation of the protein functions in nutrient sensing to regulate growth, cell division, and the initiation of sporulation.

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Section: Discussionmentioning

confidence: 86%

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confidence: 99%

Nutrient-stimulated methylation of a membrane protein in Bacillus licheniformis

et al. 1988

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“…Swarm plates were spotted with fresh bacterial colonies and incubated at room (29). The filters were probed with antisera directed against sheared B. subtilis flagella.…”

Section: Methodsmentioning

confidence: 99%

Transposon Tn917lacZ mutagenesis of Bacillus subtilis: identification of two new loci required for motility and chemotaxis

et al. 1990

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We have used Tn9171acZ to mutagenize the Bacillus subtilis chromosome and have isolated mutants that are defective in chemotaxis and motility. Mapping of the transposon inserts identified two new loci. Mutations in one of these loci generated mutants that had paralyzed flagella. Accordingly, we designate this a mot locus. The other locus is closely linked to the first and encodes proteins specifying chemotaxis functions. This locus is designated the cheX locus. Both the mot and cheX loci map close to ptsl. An additional transposon insert that maps in the hag locus was obtained. The pattern of P-galactosidase expression from some of the transposons suggested that the mot locus is regulated by sigD, a minor sigma factor of B. subtllis. The cheX locus appeared to be under the control of vegetative sigA. Four transposon inserts were mapped to a previously characterized che locus near spcB. These mutants did not produce flagellin and were defective in the methylation of the methyl-accepting chemotaxis proteins. This locus probably encodes proteins required for flagellum biosynthesis and other proteins that are required for the methylation response.The mechanism of bacterial chemotaxis has been the focus of an intense study over the last decade, and recently a model has been proposed for Escherichia coli that appears to be consistent with most genetic, biochemical, and physiological data (2). Over this same period, we have shown that the mechanism of chemotaxis in Bacillus subtilis appears to differ from that in E. coli in several respects (23-25).Methanol does not appear to be produced from a direct hydrolysis of methyl groups from the methyl-accepting chemotaxis proteins (MCPs) (24). We also observe that methanol is released in response to the addition or removal of attractants or repellents (24). In E. coli, negative stimuli (removal of attractants or addition of repellents) cause methanol formation but positive stimuli (addition of attractants or removal of repellents) prevent even basal levels of methanol formation (6,22,26). Methanol release in response to all stimuli has also been reported for Halobacterium halobium (1). B. subtilis motile cells move toward all amino acids (16), whereas cells of E. coli have a neutral response to some amino acids and are repelled by others (12,27). This more elaborate methylation mechanism and the greater variety of chemoeffectors detected should be reflected in the existence of additional gene products that are not found in E. coli.There are only six proteins that are known to be required for chemotaxis in E. coli and Salmonella typhimurium. These are encoded by the cheA, cheB, cheR, cheW, cheY, and cheZ genes (8). Other proteins are also involved, but mutants lacking these proteins have specific defects in flagellum biosynthesis, flagellum structure, motility (rotation of the flagellum), or switching (flagellum reversal) or are unable to respond to specific chemoeffectors.

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“…The results of Wang and Doi (13) suggested that three polypeptides were produced from the P23 gene when the products were expressed as fusion proteins to 3-galactosidase from plasmids in B. subtilis and E. coli. Their results indicated that these polypeptides were all in the same phase and so would share the same C-terminal amino acids.…”

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A mutation in P23, the first gene in the RNA polymerase sigma A (sigma 43) operon, affects sporulation in Bacillus subtilis

Zuberi

Doi

1990

J Bacteriol

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Mutations within P23, the first gene of the Bacillus subtilis SA operon, were not detrimental to vegetative growth or sporulation. One deletion of P23 resulted in a strain that sporulated earlier than the wild type. This aberrant phenotype may be due to the simultaneous deletion of a CH promoter from the FA operon.The Bacillus subtilis major vegetative RNA polymerase u factor (uA) is encoded by the sigA gene. The sigA gene has been cloned and sequenced (3,8; GenBank/EMBL accession no. X03897). It was found to lie within an operon with the dnaE (encodes DNA primase) and the P23 genes, the gene order being P23, dnaE, sigA (11).The organization of the B. subtilis CA operon is similar to that of the Escherichia coli Cr70 operon (11). The genes and gene products of dnaE and sigA are very homologous to their E. coli counterparts, dnaG and rpoD, respectively. In contrast, the deduced 23-kilodalton polypeptide product encoded by the P23 gene has no homology to E. coli ribosomal protein S21, which is the product of rpsU, the first gene in the r70 operon. A computer search of the Georgetown protein data bank did not reveal any significant homology of the P23 product to any known protein.The results of Wang and Doi (13) suggested that three polypeptides were produced from the P23 gene when the products were expressed as fusion proteins to 3-galactosidase from plasmids in B. subtilis and E. coli. Their results indicated that these polypeptides were all in the same phase and so would share the same C-terminal amino acids. The molecular masses of these polypeptides were deduced from sequence data as 23, 19, and 9 kilodaltons. Expression from the P23 and P19 ribosome-binding sites (RBSs) was observed during vegetative growth, but expression from the P9 RBS was observed only after the onset of sporulation. This pattern of expression was attributed to promoter switching between the three major promoters of the CrA operon (12). The operon is expressed during vegetative growth from the 9A-dependent P1 and P2 promoters located upstream from the P23 RBS. An additional promoter is located within the P23 gene between the P19 and P9 RBSs. Expression from this P3 promoter is induced at T2 and is dependent upon cH, the gene product of spoOH (2).As part of a continuing genetic characterization of the B. subtilis crA operon, we have mutated the P23 gene in order to attempt to identify its role during growth and sporulation.Both Figure 1 shows the predicted structure in the P23 gene region for the three different B. subtilis Cmr mutant derivatives DB226, DB227, and DB228. DB226 contains the cat gene inserted at the position of the XmnI site of P23. DB227 contains an XmnI-RsaI deletion of P23, replaced with the cat gene. DB228 contains a cat gene replacement of the HaeIIIRsaI fragment at the 3' end of the P23 gene. The integration was confirmed in each case by Southern hybridization. Hindlll-digested chromosomal DNA from each transformant was probed with plasmid pUC19-P23 (L. F. Wang and R. H. Doi, unpublished). This plasmid carries the com...

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Developmental expression of three proteins from the first gene of the RNA polymerase sigma 43 operon of Bacillus subtilis

Cited by 17 publications

References 29 publications

Nutrient-stimulated methylation of a membrane protein in Bacillus licheniformis

Nutrient-stimulated methylation of a membrane protein in Bacillus licheniformis

Transposon Tn917lacZ mutagenesis of Bacillus subtilis: identification of two new loci required for motility and chemotaxis

A mutation in P23, the first gene in the RNA polymerase sigma A (sigma 43) operon, affects sporulation in Bacillus subtilis

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