Nucleotide sequence of a promoter recognized by Bacillus subtilis RNA polymerase

Lee, Gloria; Talkington, Carol; Pero, Janice

doi:10.1007/bf00267352

Cited by 72 publications

(36 citation statements)

References 24 publications

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“…As can be seen belongs to the family of IS elements, and consequently we proin Figure 2, this putative promoter, located exactly upstream of pose to denote this RS as IS231. The IS231-encoded protein was the putative transposase (Figure 1) (Lee and Pero, 1981;Lee et al, 1980); line 4, B. subtilis phage <29 G2 promoter (Yoshikawa et al, 1981); line 5, consensus sequence recognized by the B. subtilis vegetative u43 factor (Moran et al, 1982). Figure 1.…”

Section: Resultsmentioning

confidence: 99%

Nucleotide sequence and structural organization of an insertion sequence element (IS231) from Bacillus thuringiensis strain berliner 1715.

Mahillon¹,

Seurinck²,

Rompuy³

et al. 1985

The EMBO Journal

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Section: Resultsmentioning

confidence: 99%

Nucleotide sequence and structural organization of an insertion sequence element (IS231) from Bacillus thuringiensis strain berliner 1715.

Mahillon¹,

Seurinck²,

Rompuy³

et al. 1985

The EMBO Journal

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“…The penicillinase and SPO-1 promoters were used to construct the hybrid promoters primarily because the nucleotide sequences and sites of transcription initiation of both had been determined (12)(13)(14). The two hybrid promoters differ slightly in the relationship of the RNA polymerase recognition site and the lac operator sequence.…”

Section: Discussionmentioning

confidence: 99%

Use of the Escherichia coli lac repressor and operator to control gene expression in Bacillus subtilis.

Yansura¹,

Henner²

1984

Proc. Natl. Acad. Sci. U.S.A.

298

219

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The Escherichia coli lac operator has been placed on the 3' side of the promoter for the penicillinase gene of Bacillus licheniformis, creating a hybrid promoter controllable by the E. coli lac repressor. The E. coli lac repressor gene has been placed under the control of a promoter and ribosomebinding site that allows expression in Bacillus subtilis. When the penicillinase gene that contains the lac operator is expressed in B. subtilis on a plasmid that also produces the lac repressor, the expression of the penicillinase gene can be modulated by isopropyl 13-D-thiogalactoside (IPTG), an inducer of the lac operon in E. coli. A similar system was constructed from a promoter of the B. subtilis phage SPO-1 and the leukocyte interferon A gene, which allowed the controlled expression of interferon in B. subtilis. These two examples show that a functional control system can be introduced into B. subtilis from E. coli.The ability to regulate transcription of a gene in Escherichia coli by using a number of easily controllable promoter systems has been important for the development of foreign protein expression systems as well as for the study of expression of native E. coli proteins. The lac repressor operator system is one such system. Repression of lac operon transcription is a result of binding of the lac repressor to the DNA sequence comprising the lac operator, thus preventing RNA polymerase from binding to the promoter (1). Induction occurs through the binding of an inducer to the lac repressor, which causes a conformational shift in the repressor, decreasing the affinity of the repressor for the operator (1). In Bacillus subtilis (or other Gram-positive organisms), no analogous system has been described. The only transcriptionally controlled promoters that have been characterized use a different mechanism. Gene regulation in these systems is controlled by a factors, which are proteins that bind to the RNA polymerase and determine the recognition site for RNA initiation (2). Such systems cannot be used for the controlled expression of another gene easily.To take advantage of the desirable characteristics of an operon whose transcription is easily induced, we have transferred the regulatory elements of the lac operon into B. subtilis. We report here that the lac operator-repressor control system can be transferred into B. subtilis and that the lac repressor and operator function as transcriptional regulatory elements in this microorganism. The penicillinase gene of Bacillus licheniformis system was used as a model system. In this system, the expression of penicillinase is modulated by isopropyl f3-D-thiogalactoside (IPTG), an inducer of the lac operon in E. coli. To further illustrate the use of this transferred control system, we demonstrated IPTG-modulated expression of leukocyte interferon in B. subtilis. MATERIALS AND METHODSStrains and Plasmids. E. coli strain MM294 (F-supE44 endAl thi-J hsdR4) was used for all constructions unless otherwise indicated (3). E. coli strain 3300 (Hfr thi-1 lacI22 relAl sp...

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“…Intrinsically curved DNA upstream of the RNA polymerase binding site strongly influences transcription in B. subtilis. Indeed, deletions which eliminate the upstream A tracts have been shown to dramatically reduce expression from several promoters in B. subtilis (4,19,21). Furthermore, the optimal positioning of the curvature relative to the position of the promoter is essential for efficient promoter utilization in B. subtilis (22).…”

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Role of curved DNA in binding of Escherichia coli RNA polymerase to promoters

Nickerson

Achberger

1995

J Bacteriol

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The ability of curved DNA upstream of the ؊35 region to affect the interaction of Escherichia coli RNA polymerase and promoter DNA was examined through the use of hybrid promoters. These promoters were constructed by substituting the curved DNA from two Bacillus subtilis bacteriophage SP82 promoters for the comparable DNA of the bacteriophage promoters p R and p L . The SP82 promoters possessed intrinsic DNA curvature upstream of their ؊35 regions, as characterized by runs of adenines in phase with the helical repeat. In vitro, the relative affinities of purified 70 -RNA polymerase for the promoters were determined in a competition binding assay. Hybrid promoters derived from p R that contained curved DNA were bound by E. coli RNA polymerase more efficiently than was the original p R . Binding of E. coli RNA polymerase to these hybrid promoters was favored on superhelical DNA templates according to gel retardation analysis. Both the supercoiled and relaxed forms of the hybrid p L series were better competitors for E. coli RNA polymerase binding than was the original p L . The results of DNase I footprinting analysis provided evidence for the wrapping of the upstream curved DNA of the hybrid p R promoters around the E. coli RNA polymerase in a tight, nucleosomal-like fashion. The tight wrapping of the upstream DNA around the polymerase may facilitate the subsequent steps of DNA untwisting and strand separation.Curved DNA upstream of the Ϫ35 region has been implicated in the function of numerous promoters (11,21,27). Studies of promoters associated with upstream curved DNA have demonstrated a strong correlation between promoter function and the presence of altered DNA structure, such as sequence-dependent curved DNA (4, 6, 13, 16-18, 21, 26, 27). The presence of curved DNA, as characterized by in-phase adenine tracts, has been found upstream of several promoters from Escherichia coli, including the ompF promoter (23), the his promoter, and the lpp promoters (29) and a subset of rRNA promoters (8) and tRNA promoters (5,6,16,18,25). Deletion analysis of the upstream curved DNA has suggested that promoter activation is intimately related to sequence-dependent curvature (16,21). In addition, sequence-dependent DNA curvature has been demonstrated to stimulate transcription when curved DNA is used to replace the upstream binding sites for the catabolite activator protein (CAP) (7,14). This suggests that protein-mediated DNA bending is functionally related to sequence-dependent DNA curvature in transcription initiation.Some of the most profound effects of curved DNA have been documented in promoters utilized by the Bacillus subtilis RNA polymerase. Intrinsically curved DNA upstream of the RNA polymerase binding site strongly influences transcription in B. subtilis. Indeed, deletions which eliminate the upstream A tracts have been shown to dramatically reduce expression from several promoters in B. subtilis (4,19,21). Furthermore, the optimal positioning of the curvature relative to the position of the promoter is essenti...

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Nucleotide sequence of a promoter recognized by Bacillus subtilis RNA polymerase

Cited by 72 publications

References 24 publications

Nucleotide sequence and structural organization of an insertion sequence element (IS231) from Bacillus thuringiensis strain berliner 1715.

Nucleotide sequence and structural organization of an insertion sequence element (IS231) from Bacillus thuringiensis strain berliner 1715.

Use of the Escherichia coli lac repressor and operator to control gene expression in Bacillus subtilis.

Role of curved DNA in binding of Escherichia coli RNA polymerase to promoters

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