Compromised Factor-Dependent Transcription Termination in a
            <i>nusA</i>
            Mutant of Escherichia coli: Spectrum of Termination Efficiencies Generated by Perturbations of Rho, NusG, NusA, and H-NS Family Proteins

Saxena, Shivalika; Gowrishankar, J.

doi:10.1128/jb.00221-11

Cited by 17 publications

(24 citation statements)

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“…Thus, expression of any one of several dominant-negative variants of H-NS, including ⌬64, L26P, and E53G/T55P, restores the efficiency of transcription termination (that is, suppresses polarity relief) in rho and nusG strains, as does overexpression of YdgT. As reported in an accompanying study (51), a nusA mutation associated with defective Rho-dependent termination is also suppressed by H-NS⌬64 and YdgT. These effects are manifest in strains that express wild-type H-NS, StpA or both proteins but not in strains that are simultaneously deficient for H-NS and StpA.…”

Section: Discussionmentioning

confidence: 67%

“…These results indicate that the effect of hns mutations on modulation of Rho-dependent termination is not completely correlated with that on transcription initiation or gene silencing. A search for genetic suppressors of the effects of H-NS on Rho-dependent termination, as reported in an accompanying study (51), also did not identify any H-NS-repressed gene as being responsible for such modulation. An alternative model is accordingly proposed below.…”

Section: Discussionmentioning

confidence: 99%

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Modulation of Rho-Dependent Transcription Termination in Escherichia coli by the H-NS Family of Proteins

Saxena

Gowrishankar

2011

J Bacteriol

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Nascent transcripts in Escherichia coli that fail to be simultaneously translated are subject to a factordependent mechanism of termination (also termed a polarity) that involves the proteins Rho and NusG. In this study, we found that overexpression of YdgT suppressed the polarity relief phenotypes and restored the efficiency of termination in rho or nusG mutants. YdgT and Hha belong to the H-NS and StpA family of proteins that repress a large number of genes in Gram-negative bacteria. Variants of H-NS defective in one or the other of its two dimerization domains, but not those defective in DNA binding alone, also conferred a similar suppression phenotype in rho and nusG mutants. YdgT overexpression was associated with derepression of proU, a prototypical H-NS-silenced locus. Polarity relief conferred by rho or nusG was unaffected in a derivative completely deficient for both H-NS and StpA, although the suppression effects of YdgT or the oligomerizationdefective H-NS variants were abolished in this background. Transcription elongation rates in vivo were unaffected in any of the suppressor-bearing strains. Finally, the polarity defects of rho and nusG mutants were exacerbated by Hha and YdgT deficiency. A model is proposed that invokes a novel role for the polymeric architectural scaffold formed on DNA by H-NS and StpA independent of the gene-silencing functions of these nucleoid proteins, in modulating Rho-dependent transcription termination such that interruption of the scaffold (as obtained by expression either of the H-NS oligomerization variants or of YdgT) is associated with improved termination efficiency in the rho and nusG mutants.Translation is a cotranscriptional process in both eubacteria and archaebacteria (1,9,25,45,50), and it has been proposed that such coupling is a defining characteristic of prokaryotic life (34, 64). The maintenance of transcription-translation coupling in a prokaryotic cell would require dynamic inter-regulation between the binding and progression of a pioneer ribosome on the nascent transcript on the one hand and the rate of transcription elongation on the other (9, 45); however, the detailed mechanisms by which such regulation is achieved are not known. Furthermore, in bacteria such as Escherichia coli, nascent transcripts that are not simultaneously translated are subject to a mechanism of factor-dependent transcription termination (also referred to as transcriptional polarity) (reviewed in references 1, 6, 15, 40, 44, 47, and 52), so that the occurrence of translation-uncoupled transcription is minimized within the cells (11, 43).Factor-dependent (also called Rho-dependent) transcription termination is mediated by, among others, the Rho and NusG proteins (1,6,15,39,40,44,47,52). Although the structures of the two proteins have been determined and several of their biochemical properties have been characterized, the precise mechanisms of their action in transcription termination remain unclear, even controversial. Rho is an RNAbinding protein and possesses RNA-dependent ATPa...

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Section: Discussionmentioning

confidence: 67%

Section: Discussionmentioning

confidence: 99%

Modulation of Rho-Dependent Transcription Termination in Escherichia coli by the H-NS Family of Proteins

Saxena

Gowrishankar

2011

J Bacteriol

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“…Genome-wide gene expression patterns of NusA-deleted strains showed same pattern as that of the NusG-deleted ones, where the latter is a bona fide partner of Rho (6). More recently, a NusA mutant was observed to produce a termination defect at a Rho-dependent terminator, H-19B t R1 (17).…”

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confidence: 69%

“…In particular, the hairpin-dependent pauses are enhanced by NusA via stabilization of the hairpin-RNAP interaction (12). NusA improves the termination efficiency at many hairpin-dependent terminators by facilitating the hairpin folding (13) and also by stabilizing the latter's interactions in the RNA exit channel of the transcription elongation complex (14).Previously, different studies have indicated the involvement of NusA in the Rho-dependent termination process (15)(16)(17). A…”

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confidence: 99%

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Transcription Elongation Factor NusA Is a General Antagonist of Rho-dependent Termination in Escherichia coli

Qayyum

Dey

Sen

2016

Journal of Biological Chemistry

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NusA is an essential protein that binds to RNA polymerase and also to the nascent RNA and influences transcription by inducing pausing and facilitating the process of transcription termination/antitermination. Its participation in Rho-dependent transcription termination has been perceived, but the molecular nature of this involvement is not known. We hypothesized that, because both Rho and NusA are RNA-binding proteins and have the potential to target the same RNA, the latter is likely to influence the global pattern of the Rho-dependent termination. Analyses of the nascent RNA binding properties and consequent effects on the Rho-dependent termination functions of specific NusA-RNA binding domain mutants revealed an existence of Rho-NusA direct competition for the overlapping nut (NusA-binding site) and rut (Rho-binding site) sites on the RNA. This leads to delayed entry of Rho at the rut site that inhibits the latter's RNA release process. High density tiling microarray profiles of these NusA mutants revealed that a significant number of genes, together with transcripts from intergenic regions, are up-regulated. Interestingly, the majority of these genes were also up-regulated when the Rho function was compromised. These results provide strong evidence for the existence of NusA-binding sites in different operons that are also the targets of Rho-dependent terminations. Our data strongly argue in favor of a direct competition between NusA and Rho for the access of specific sites on the nascent transcripts in different parts of the genome. We propose that this competition enables NusA to function as a global antagonist of the Rho function, which is unlike its role as a facilitator of hairpin-dependent termination.The bacterial transcription terminator, Rho, functions as a hexameric RNA-dependent ATPase that is capable of translocating along the RNA. It dislodges the elongating RNA polymerase (RNAP) 4 once it is loaded onto the nascent RNA (1, 2). Its termination function is facilitated upon interaction with the transcription elongation factor, NusG (3-5). Recent genomics studies have revealed that Rho-dependent termination occurs at more than one-third of the operons of a dividing Escherichia coli (6, 7). This mode of transcription termination is involved in many physiological processes, like control of translation (2), riboswitch formation (8), inhibition of unwanted antisense transcription, etc. (7). Rho is capable of loading onto unstructured stretches of RNA (the rut ([R]ho [ut]ilization) sites), and once it is bound to the RNA, it translocates in a processive manner along the 5Јto 3Ј direction, which may induce unwanted termination of transcription. Interestingly, in vivo negative regulation by cellular factors or any control switch of the Rho function has not been documented. In principle, any RNA-binding protein that has overlapping binding sites with the rut sites on the nascent RNA could influence the Rho activity by a direct competition for occupancy of the same sites.NusA, a ubiquitous transcription elo...

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2020

Structure and Function of the Bacterial Genome

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Compromised Factor-Dependent Transcription Termination in a nusA Mutant of Escherichia coli: Spectrum of Termination Efficiencies Generated by Perturbations of Rho, NusG, NusA, and H-NS Family Proteins

Cited by 17 publications

References 58 publications

Modulation of Rho-Dependent Transcription Termination in Escherichia coli by the H-NS Family of Proteins

Modulation of Rho-Dependent Transcription Termination in Escherichia coli by the H-NS Family of Proteins

Transcription Elongation Factor NusA Is a General Antagonist of Rho-dependent Termination in Escherichia coli

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