RNA-binding specificity of E. coli NusA

Prasch, Stefan J.; Jurk, Marcel; Washburn, Robert S.; Gottesman, Max E.; Wöhrl, Birgitta M.; Rösch, Paul

doi:10.1093/nar/gkp452

Cited by 31 publications

(34 citation statements)

References 45 publications

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“…This indicates that the intrinsic affinity of the NusA RNA binding domain for the nut site increased due to the R258C mutation. The obtained binding constant of the WT SKK domain is consistent with what has been reported earlier (10).…”

Section: Nut Site-dependent Inhibition Of Rho Function By Nusasupporting

confidence: 92%

“…2B). Among these terminators, only the t R1 terminators have an overlapping NusA binding site, nutR (10) (Fig. 1C); hence, the termination defects caused by these two NusA mutants were only observed with the terminators having nut sites.…”

Section: Nut Site-dependent Inhibition Of Rho Function By Nusamentioning

confidence: 99%

“…6G). Both the SKK domain and the NusA devoid of AR1-2 domains are capable of binding to RNA in trans (10), and the latter construct was used because the presence of AR1 and AR2 domains inhibits NusA from binding to the RNA (30). To avoid the translocase activity of Rho, instead of ATP, its non-hydrolyzable analogue, AMP-PNP, was used.…”

Section: Nusa Binding To Nut Site(s) Delays the Loading Of Rho At Thementioning

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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Section: Nut Site-dependent Inhibition Of Rho Function By Nusasupporting

confidence: 92%

Section: Nut Site-dependent Inhibition Of Rho Function By Nusamentioning

confidence: 99%

Section: Nusa Binding To Nut Site(s) Delays the Loading Of Rho At Thementioning

confidence: 99%

See 1 more Smart Citation

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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“…This hypothesis needs further study. (2) NusA modulates transcriptional pausing, termination, and antitermination (Paliy et al, 2008;Prasch et al, 2009;Cohen et al, 2009; Fig. 3 Identification of protein of Spot 5 by mass spectrometry Probability-based Mowse score of Spot 5 is shown.…”

Section: Discussionmentioning

confidence: 99%

Comparative proteome analysis of Helicobacter pylori clinical strains by two-dimensional gel electrophoresis

Zhang

Ding

Huang³

et al. 2011

J. Zhejiang Univ. Sci. B

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Abstract:Objective: To investigate the pathogenic properties of Helicobacter pylori by comparing the proteome map of H. pylori clinical strains. Methods: Two wild-type H. pylori strains, YN8 (isolated from biopsy tissue of a gastric cancer patient) and YN14 (isolated from biopsy tissue of a gastritis and duodenal ulcer patient), were used. Proteomic analysis, using a pH range of 3-10 and 5-8, was performed. The individual proteins were identified by quadrupole time-of-flight (Q-TOF) mass spectrometer and protein database search. Results: Variation in spot patterns directed towards differential protein expression levels was observed between the strains. The gel revealed prominent proteins with several protein "families". The comparison of protein expressions of the two strains reveals a high variability. Differentially present or absent spots were observed. Nine differentially expressed protein spots identified by Q-TOF included adenosine triphosphate (ATP)-binding protein, disulfide oxidoreductase B (DsbB)-like protein, N utilization substance A (NusA), ATP-dependent protease binding subunit/heat shock protein, hydantoin utilization protein A, seryl-tRNA synthetase, molybdenum ABC transporter ModD, and hypothetical proteins. Conclusions: This study suggests that H. pylori strains express/repress protein variation, not only in terms of the virulence proteins, but also in terms of physiological proteins, when they infect a human host. The difference of protein expression levels between H. pylori strains isolated from gastric cancer and gastritis may be the initiator of inflammation, and result in the different clinical presentation. In this preliminary study, we report seven differential proteins between strains, with molecule weights from approximately 10 kDa to approximately 40 kDa. Further studies are needed to investigate those proteins and their function associated with H. pylori colonization and adaptation to host environment stress.

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“…The λ N protein further binds with E. coli proteins (NusA, NusB, NusG, and NusE), inducing a conformational change in RNAP that leads to a terminationresistant complex. [20][21][22] It must be noted here that evolutionary changes in the N-protein contributes to the diversity of phages such as HK022, in which Nun protein acts an anti-termination factor for early transcripts.…”

Section: Gene Transcription Regulation Of Phage λmentioning

confidence: 95%