DNA Supercoiling and transcription in Escherichia coli: Influence of RNA polymerase mutations

Mirkin, Sergei M.; Gorlenko, Zh. M.; Gragerov, Alexander; Larionov, O. A.

doi:10.1007/bf00267267

Cited by 23 publications

(12 citation statements)

References 28 publications

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“…Two-dimensional protein gel electrophoresis of E. coli gyrB mutants, which have a 25% steady-state decrease in the level of supercoiling, showed that 45 of 90 proteins had altered synthesis rates (7). Most proteins affected by the lowered levels of supercoiling showed an increase in abundance, consistent with our findings and those of Mirkin et al (21). In further agreement, Menzel and Gellert have observed that 4 of 20 random promoters shotgun cloned into a pKO1 plasmid were strongly induced by the addition of coumermycin A1 (19).…”

Section: Discussionsupporting

confidence: 93%

Estimation of the effect of coumermycin A1 on Salmonella typhimurium promoters by using random operon fusions

Jovanovich

Lebowitz

1987

J Bacteriol

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We have estimated the extent to which relaxation of supercoiling by the DNA gyrase inhibitor coumermycin A1 affects gene expression in vivo in Salmonella typhimurium. We isolated a set of Mu dl-8 Lac+ operon fusions to random promoters and measured the effect of coumermycin A1 on the expression of 67 fusions. The differential rate of synthesis was increased for 70% of the fusions and decreased for 16%, and 13% of the fusions had less than a 25% change in expression. The coumermycin Al response was found to correlate well (P = 0.067) with the basal level of expression such that coumermycin Al tended to stimulate fusions with low expression and inhibit those with high expression. Since the vast majority of the fusions were sensitive to coumermycin Al addition and, therefore, to the level of supercoiling, these results indicate that if the level of supercoiling were to vary under physiological conditions, then major readjustments in the cellular economy would occur.In vivo, the DNA of all procaryotes is believed to be in the supercoiled state (for reviews, see references 1, 3, 9, 31, and 35). The degree of supercoiling is maintained in vivo at a level characteristic of the host organism by an apparently homeostatic balance of the actions of DNA gyrase and DNA topoisomerase I (2,18,25,34). DNA gyrase is an essential enzyme in Escherichia coli and is composed of two subunits, one encoded by gyrA and the other encoded by gyrB. These subunits are the targets of resistance to the antibiotics nalidixic acid and oxolinic acid for gyrA and coumermycin A1 and novobiocin for gyrB (9). Inhibition of gyrase can result in a total cessation of DNA replication (10) along with a decreased capacity for repair (27) and recombination (12). DNA topoisomerase I, the topA product in E. coli (32, 33), decreases the amount of underwinding, and therefore supercoils, in the DNA (25). Significantly, mutations in topA have pleiotropic cellular effects, including increasing the specific activity of alkaline phosphatase (5) and the expression of lacLI (6) and leu (8), as well as conferring increased sensitivity to UV light damage (23).The transcription of many genes and operons is affected by the degree of supercoiling. Hayashi and Hayashi (11) first demonstrated that in vitro transcription of 4X replicativeform DNA is enhanced about 20-fold when supercoiled DNA is compared to nicked DNA. Subsequently, trp, lambda PL, the three maltose operons (28), lac, an rRNA gene, the colicin gene of ColEl, and gal (36) were all shown to have increased transcription when supercoiled or, conversely, to have decreased transcription in the presence of gyrase inhibitors. Genes known to have increased expression in the presence of DNA gyrase inhibitors are lacUVS, lacI (28), his (26), gyrA and gyrB (18), and genes controlled by either the heat shock system (22) or the SOS repair system (16).In the present study we estimated the percentage of S. typhimurium genes which are affected by the addition of the DNA gyrase inhibitor coumermycin A1, which causes the loss ...

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Section: Discussionsupporting

confidence: 93%

Estimation of the effect of coumermycin A1 on Salmonella typhimurium promoters by using random operon fusions

Jovanovich

Lebowitz

1987

J Bacteriol

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“…Contradictory results concerning the effects of these inhibitors on the differential rate of tryptophanase synthesis have been reported (Shuman and Schwartz, 1975;Sanzey, 1979). Inactivation of gyrase enhanced promoter activities in several other cases too (Mirkin et al, 1979;Fairweather et al, 1980;DiNardo et al, 1982). Inactivation of gyrase enhanced promoter activities in several other cases too (Mirkin et al, 1979;Fairweather et al, 1980;DiNardo et al, 1982).…”

Section: Introductionmentioning

confidence: 99%

Gene expression in a temperature-sensitive gyrB mutant of Escherichia coli.

Wahle¹,

Mueller²,

Orr³

1984

The EMBO Journal

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The effect of gyrase inactivation on gene expression was studied by examining the activities of different promoters in a temperature‐sensitive gyrB mutant of Escherichia coli. The relative activities of promoters affected by cAMP‐binding protein (CAP), e.g., the lac promoter, are not reduced by gyrase inactivation but can, on the contrary, be enhanced. This stimulation depends on the promoter location or its structure. The tnaA promoter is activated when located near the origin of replication, suggesting a differential effect of gyrase inactivation on various chromosomal domains. Only silent or mutant promoters such as the non‐functional wild‐type bgl or the lacIq can be activated. No differential effect of gyrase inactivation on the lambda pL and the trp promoters carried by the phage can be detected.

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“…Following the initial indications that superhelical viral DNA can be more efficiently transcribed than relaxed DNA both in vitro (25) and in vivo (42), much evidence that DNA superhelicity affects transcription initiation has accumulated; most promoters are best transcribed from supercoiled templates, while some are best transcribed from relaxed ones (for reviews, see references 10, 18, 53, and 54). A related observation is that mutations in the gene for the d subunit of RNA polymerase (rpoB) can suppress the defective initiation of transcription and replication produced by a number of DNA gyrase mutations (11,13,35). Furthermore, rpoB mutations and treatment of wild-type strains with rifampin (which binds to the 1 subunit of RNA polymerase) can alter the degree of dependence of transcription initiation on superhelicity (11,13,35).…”

mentioning

confidence: 99%

“…A related observation is that mutations in the gene for the d subunit of RNA polymerase (rpoB) can suppress the defective initiation of transcription and replication produced by a number of DNA gyrase mutations (11,13,35). Furthermore, rpoB mutations and treatment of wild-type strains with rifampin (which binds to the 1 subunit of RNA polymerase) can alter the degree of dependence of transcription initiation on superhelicity (11,13,35).…”

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confidence: 99%

Regulation of DNA superhelicity by rpoB mutations that suppress defective Rho-mediated transcription termination in Escherichia coli

Arnold

Tessman

1988

J Bacteriol

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The highly defective rho-IS mutant of Escherichia coli produces plasmid DNA that is 22% less negatively supercoiled than DNA from an isogenic wild-type strain (J. S. Fassler, G. F. Arnold, and I. Tessman, Mol.Gen. Genet. 204:424-429, 1986 The ability of the Rho protein of Escherichia coli to affect DNA superhelicity in a substantial way provides the protein with a potentially powerful regulatory function and suggests how some of the pleiotropy of the rho-15 gene might arise (12). The superhelicity of DNA, determined for the pBR327-related plasmid pJSF6, is reduced by 22 + 2% in rho-15 cells, which is comparable to the reduction of 32 + 2% in gyrB203(Ts) gyrB22J(Cour) cells (12). The mechanism by which Rho affects DNA superhelicity is not known. An indirect effect through the regulation of E. coli topoisomerase synthesis (12) now appears unlikely because cellular levels of gyrase A, gyrase B, and topoisomerase I are not significantly altered in rho-15 cells (G. F. Arnold, Ph.D. thesis, Purdue University, West Lafayette, Ind., 1987). We focus on the possibility of a more direct effect of the Rho protein on superhelicity, which could conceivably occur through its activity in transcription termination.The aim of the current work was to see how DNA superhelicity is affected by mutations that suppress defective Rho-mediated transcription termination. Some of the suppressor mutations studied were in the rho gene itself, but the most striking results were provided by suppressor mutations that were in the rpoB gene, which encodes the ,B subunit of RNA polymerase. Transcription termination and the pausing that precedes it appear to be mediated by the ,B subunit of RNA polymerase (14,37,56). Indirect genetic evidence suggests that the Rho protein interacts with this subunit of RNA polymerase

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DNA Supercoiling and transcription in Escherichia coli: Influence of RNA polymerase mutations

Cited by 23 publications

References 28 publications

Estimation of the effect of coumermycin A1 on Salmonella typhimurium promoters by using random operon fusions

Estimation of the effect of coumermycin A1 on Salmonella typhimurium promoters by using random operon fusions

Gene expression in a temperature-sensitive gyrB mutant of Escherichia coli.

Regulation of DNA superhelicity by rpoB mutations that suppress defective Rho-mediated transcription termination in Escherichia coli

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