Complex character of senS, a novel gene regulating expression of extracellular-protein genes of Bacillus subtilis

Wang, Lin‐Fa; Doi, Roy H.

doi:10.1128/jb.172.4.1939-1947.1990

Cited by 33 publications

(20 citation statements)

References 53 publications

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“…Similar organizations have been reported in at least three other cases, i.e., sacB (32) and senS (38) in B. subtilis and bgl (26) in E. coli. The transcription terminator-like structure has been demonstrated to function as a regulatory device in controlling the induction of sacB, the structural gene for levansucrase (28,31,48).…”

Section: Resultssupporting

confidence: 60%

“…In the third case, senS, a homolog of senN, also carries an upstream terminatorlike structure. If the terminatorlike structure is deleted, it is lethal for B. subtilis (38). For the ten operon, the presence of the terminatorlike structure reduces the expression of the operon.…”

Section: Resultsmentioning

confidence: 99%

“…Furthermore, the cloning of several regulatory genes from B. subtilis, B. natto, and B. stearothermophilus has been shown to enhance the production of extracellular enzymes when these genes are in a multicopy plasmid. These genes are sacV (17), degR (19,47), senS (38), senN (39,42), and degT (33). The size of the protein products derived from these genes varies from small polypeptides (46 to 65 amino acids for the degR, sacV, senS, and senN products) to larger polypeptides (372 amino acids for the degT product).…”

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Cloning and characterization of a pair of novel genes that regulate production of extracellular enzymes in Bacillus subtilis

1991

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Two novel Bacillus subtilis genes that regulate the production of several extracellular enzymes were cloned and characterized. These two genes are organized as part of an operon. When cloned in a multicopy plasmid, the first gene (tenA, transcription enhancement) stimulates alkaline protease production at the transcriptional level. The second gene (tenI) exerts an opposite effect to reduce alkaline protease production. The production of neutral protease, levansucrase, and alkaline protease can be stimulated up to 11-to Bacillus subtilis is capable of secreting a wide variety of extracellular enzymes (proteases, a-amylase, levansucrase, and P-glucanases) directly into the medium (22). Several mutants that can stimulate the production of extracellular enzymes have been isolated. These mutants carry mutations in degQ and sacU (12,14). Structural genes encoding degQ and sacU have been cloned (1,8,11,34,36,45). Furthermore, the cloning of several regulatory genes from B. subtilis, B. natto, and B. stearothermophilus has been shown to enhance the production of extracellular enzymes when these genes are in a multicopy plasmid. These genes are sacV (17), degR (19, 47), senS (38), senN (39,42), and degT (33). The size of the protein products derived from these genes varies from small polypeptides (46 to 65 amino acids for the degR, sacV, senS, and senN products) to larger polypeptides (372 amino acids for the degT product). Although these gene products show no sequence homology with each other, they all stimulate the production of extracellular enzymes. The target genes encoding extracellular enzymes such as aprE (alkaline protease) (30, 41), nprE (neutral protease) (46), amyE (oa-amylase) (44), and sacB (levansucrase) (32) have also been cloned. With these mutants and the cloned genes available, the mechanism for the enhanced production of extracellular enzymes can be studied in detail. degR, senN, degQ, and sacU have been shown to exert their stimulatory effects at the transcriptional level (28,35,42). However, in the sacU minus mutant strain, no enhanced production of extracellular enzymes can be observed with cells carrying either degR or degQ on a multicopy plasmid (1,34).In this report, we describe in detail the cloning, nucleotide sequence, genetic mapping, and organization of a set of two novel genes from B. subtilis that can regulate the production of several extracellular enzymes. They were isolated by using a shotgun cloning approach and were selected by their abilities to stimulate the production of alkaline protease. The * Corresponding author. characterization of these genes and their roles in regulating gene expression are discussed. MATERIALS AND METHODSPlasmids. Plasmid pUB18 (42), a derivative of pUB110, was used for routine subcloning in B. subtilis. pPQ is a pUB18 derivative carrying degQ (40). Bluescribe plasmid from Escherichia coli was used for cloning and doublestranded DNA sequencing. Standard DNA transformation was performed by the competent cell method in B. subtilis (29) and E. coli (16).Med...

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Section: Resultssupporting

confidence: 60%

Section: Resultsmentioning

confidence: 99%

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Cloning and characterization of a pair of novel genes that regulate production of extracellular enzymes in Bacillus subtilis

1991

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“…Several regulatory genes encoding small polypeptides of 46 to 65 amino acid residues which cause a higher level of expression of a class of extracellular proteins in B. subtilis have been reported, i.e., senS (45), degR (25,42,49), and degQ (3,48). The pleiotropic effects of these genes suggest that they are part of a global control mechanism affecting the synthesis of degradative enzymes.…”

Section: Discussionmentioning

confidence: 99%

DegS-DegU and ComP-ComA modulator-effector pairs control expression of the Bacillus subtilis pleiotropic regulatory gene degQ

et al. 1991

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Production of a class of both secreted and intracellular degradative enzymes in Bacillus subtilis is regulated at the transcriptional level by a signal transduction pathway which includes the DegS-DegU two-component system and at least two additional regulatory genes, degQ and degR, encoding polypeptides of 46 and 60 amino acids, respectively. Expression of degQ was shown to be controlled by DegS-DegU. This Production of a class of degradative enzymes in Bacillus subtilis, including an intracellular protease and several secreted enzymes (levansucrase, alkaline and metalloproteases, a-amylase, 3-glucanase[s], and xylanase) (3,5,11,29), is controlled at the transcriptional level by the DegSDegU two-component system and by at least two additional regulatory genes, degQ and degR, which encode small polypeptides of 46 and 60 amino acids, respectively (3,25,42,48,49).Although the two-component system is required for degradative enzyme production, the products of degQ and degR appear to be dispensable (48,49). Mutations were identified in both degS and degU leading to either deficiency of degradative enzyme production or a pleiotropic (Hy) phenotype, which includes hyperproduction of degradative enzymes, ability to sporulate in the presence of glucose, decreased genetic competence, and loss of flagella (6,12,17,23).Since degradative enzyme production requires both the DegS protein kinase and the DegU effector, we postulated that the phosphorylated form of DegU may be necessary for this process. On the other hand, we postulate that the nonphosphorylated form of DegU may be required for competence (23, 28; this report). This hypothesis is supported by the following observations: (i) the degUJ46 mutation abolishing the putative site of phosphorylation of the DegU effector did not abolish competence (23), and (ii) deletion of the degS gene did not strongly reduce competence (this report).The degQ36 mutation, a single base change at position -10 leading to overexpression of the degQ gene, gave a Hy phenotype similar to that of the degS(Hy) and degU(Hy) * Corresponding author. mutations (3, 48). The increase of degradative enzyme production due to overproduction of DegQ is, however, strictly dependent upon the presence of a functional DegSDegU two-component system (3; this report).We previously reported that expression of degQ was decreased in strains carrying either a deletion of degS and degU or a degU32(Hy) mutation (23). In this report, we show that degQ expression is regulated by a second twocomponent system controlling competence in B. subtilis: 47). Several findings had suggested that this could be the case: (i) DegS-DegU and ComP-ComA show strong amino acid sequence similarities, (ii) both of these two-component systems control the expression of competence, and (iii) the degQ, comP, and comA genes are located at adjacent positions on the B. subtilis chromosome (47).The DegS, DegU, and DegQ proteins are produced at different times during growth. The degS-degU operon is expressed throughout the exponential growth pha...

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“…The authors later identified the B. subtilis counterpart of senN named senS that also functions as a positive regulator of aprE expression. SenS is a positively charged, 65-amino-acid protein with a helix-turn-helix motif in the molecule (25). It was demonstrated that SenS exerts its positive effect by acting on the region between nt Ϫ177 and Ϫ415 upstream of the transcription start site of aprE (5), but further details, including the target of SenS and the relationship with the other transcriptional regulators, have not been investigated.…”

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Inhibition of Bacillus subtilis scoC Expression by Multicopy senS

2005

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The Bacillus subtilis aprE gene, which encodes the extracellular alkaline protease, is regulated by many positive and negative transcriptional regulators. SenS is one such positive regulator consisting of 65 amino acids. We found that the senS gene on a multicopy plasmid, pSEN24, caused an increase in aprE expression in strains carrying the upstream region of aprE up to ؊340 with respect to the transcription initiation site but not in a strain carrying the region up to ؊299, which is within the binding site of the negative regulator ScoC (Hpr). Epistatic analysis showed that the pSEN24 effect was lost in a scoC-deleted mutant. In accordance with these results, the scoC transcription level as assayed by a scoC-lacZ fusion and Northern analysis was greatly reduced in the cells carrying pSEN24. From these results we conclude that multicopy senS enhances aprE expression by suppressing the transcription of scoC.Bacillus subtilis secretes degradative enzymes after the end of logarithmic growth, apparently for degrading high-molecular-weight materials around the cell to cope with adverse nutritional conditions (17,23). Among the extracellular proteases produced by this organism, the alkaline and neutral proteases constitute the major part of the protease activities. Since these enzymes are produced on the order of grams per liter, expression of the genes coding for the enzymes (aprE and nprE, respectively) has to be strictly controlled to avoid extravagant use of energy and materials. Thus, the aprE gene is regulated by many positive and negative regulators. The positive regulators include Spo0A, the DegS-DegU two-component system, DegR, DegQ, ProB, TenA, RelA, SalA, and SenS, while the negative regulators include AbrB, ScoC (previously called Hpr), SinR, and Pai (2,9,10,20). Among these factors, AbrB, DegU, ScoC, and SinR are known to regulate aprE expression directly by binding to upstream regions of the aprE coding sequence (1,4,11,22). Therefore, it can be summarized that the major regulatory pathways controlling aprE expression are the routes via AbrB, ScoC, SinR, and DegU. The positive effect of Spo0A is through inhibition of abrB expression (20), while those of DegR, DegQ, ProB, and TenA are through functional DegU (6,8,9,15). It was shown recently that disruption of salA caused a decrease in aprE expression, and this was attributed to the enhanced synthesis of ScoC (10). In addition, the stringent factor RelA was shown to be required for the efficient expression of aprE (2). The mode of action of the pai gene product remains to be studied.The DNA-binding transcription regulators, such as ScoC, SinR, DegU, and AbrB, have their own specific target sites in the control region of aprE, i.e., for ScoC, these are nucleotides (nt) spanning Ϫ324 to Ϫ267 and Ϫ79 to Ϫ14; for SinR, Ϫ268 to Ϫ220; for DegU, Ϫ164 to Ϫ113 and/or Ϫ70 to Ϫ27; and for AbrB, Ϫ59 to ϩ25 (3,4,19,22 (5), but further details, including the target of SenS and the relationship with the other transcriptional regulators, have not been investigated. In th...

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Complex character of senS, a novel gene regulating expression of extracellular-protein genes of Bacillus subtilis

Cited by 33 publications

References 53 publications

Cloning and characterization of a pair of novel genes that regulate production of extracellular enzymes in Bacillus subtilis

Cloning and characterization of a pair of novel genes that regulate production of extracellular enzymes in Bacillus subtilis

DegS-DegU and ComP-ComA modulator-effector pairs control expression of the Bacillus subtilis pleiotropic regulatory gene degQ

Inhibition of Bacillus subtilis scoC Expression by Multicopy senS

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