Transcription elongation: structural basis and mechanisms

Nudler, Evgeny

doi:10.1006/jmbi.1999.2641

Cited by 99 publications

(69 citation statements)

References 92 publications

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“…There are several models proposed to explain such functional differences (4,46,(57)(58)(59). All of the models suggest that hydrolytic cleavage of the nascent transcript within an arrested or stalled ternary complex generates a new transcript 3Ј-end at the active site.…”

Section: Discussionmentioning

confidence: 99%

“…Mammalian RNA polymerase II U138 complexes, like the yeast ternary complexes, are also susceptible to intrinsic transcript cleavage (data not shown). Perhaps the efficiency with which ternary complexes carry out intrinsic transcript cleavage correlates with particular types of structural alterations, such as the mobility of the catalytic center forward or back on the DNA template (46), or the positioning of the RNA transcript within the proposed RNA exit channel (66), or extruded from the ternary complex (58). Clearly all these models make testable predictions for future work.…”

Section: Discussionmentioning

confidence: 99%

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Intrinsic Transcript Cleavage in Yeast RNA Polymerase II Elongation Complexes

Weilbaecher

Awrey

Edwards

et al. 2003

Journal of Biological Chemistry

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Transcript elongation can be interrupted by a variety of obstacles, including certain DNA sequences, DNAbinding proteins, chromatin, and DNA lesions. Bypass of many of these impediments is facilitated by elongation factor TFIIS through a mechanism that involves cleavage of the nascent transcript by the RNA polymerase II/TFIIS elongation complex. Highly purified yeast RNA polymerase II is able to perform transcript hydrolysis in the absence of TFIIS. The "intrinsic" cleavage activity is greatly stimulated at mildly basic pH and requires divalent cations. Both arrested and stalled complexes can carry out the intrinsic cleavage reaction, although not all stalled complexes are equally efficient at this reaction. Arrested complexes in which the nascent transcript was cleaved in the absence of TFIIS were reactivated to readthrough blocks to elongation. Thus, cleavage of the nascent transcript is sufficient for reactivating some arrested complexes. Small RNA products released following transcript cleavage in stalled ternary complexes differ depending upon whether the cleavage has been induced by TFIIS or has occurred in mildly alkaline conditions. In contrast, both intrinsic and TFIIS-induced small RNA cleavage products are very similar when produced from an arrested ternary complex. Although ␣-amanitin interferes with the transcript cleavage stimulated by TFIIS, it has little effect on the intrinsic cleavage reaction. A mutant RNA polymerase previously shown to be refractory to TFIIS-induced transcript cleavage is essentially identical to the wild type polymerase in all tested aspects of intrinsic cleavage.Gene expression can be controlled by regulating any step of the transcription cycle: promoter binding, initiation, promoter escape, elongation, or termination. After transcript synthesis begins, the ability to suppress or complete the synthesis of an RNA transcript is vital to the cell, and a substantial and growing number of genes have been reported to be regulated during transcript elongation (1).The ternary elongation complex consists of the RNA polymerase, the template DNA, and the nascent RNA transcript. Surratt et al. (2) discovered that bacterial RNA polymerase ternary complexes have a mechanism for transcript shortening, endonucleolytic cleavage near the 3Ј-end of the transcript. After transcript cleavage, the 3Ј-fragment is released while the 5Ј-fragment is retained in an active ternary complex. A single cleavage event can liberate from 1 to 17 nt 1 of RNA bearing a 5Ј-monophosphate (3-6). Remarkably, transcription accurately resumes from the site of transcript cleavage to re-synthesize the excised RNA.Transcript cleavage activity is conserved in many DNA-dependent RNA polymerases, including bacterial RNA polymerases, vaccinia virus RNA polymerase (7), RNA polymerase I (8, 9), RNA polymerase II (10 -14), and RNA polymerase III (15). Accessory factors that stimulate transcript cleavage in ternary complexes have been identified in both prokaryotes (GreA and GreB) and eukaryotes (reviewed in Refs. 3,9,[16][17]...

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Section: Discussionmentioning

confidence: 99%

Section: Discussionmentioning

confidence: 99%

Intrinsic Transcript Cleavage in Yeast RNA Polymerase II Elongation Complexes

Weilbaecher

Awrey

Edwards

et al. 2003

Journal of Biological Chemistry

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“…Backtracking of RNAPIIo might be one of the key mechanisms to allow repair and/or transcription restart. During backtracking, RNAPIIo and the transcription-associated DNA bubble shifts backward along the RNA [86] and restart of transcription depends on cleavage of the extruded mRNA to reposition the 3' end of the RNA to the active center of RNAPIIo [87]. The transcription cleavage factor TFIIS implicated in TC-NER [54,88] can stimulate the cleavage activity of RNAPIIo, allowing arrested RNAPIIo to restart elongation.…”

Section: Transcription Restart: a Crucial Role For Tfiis?mentioning

confidence: 99%

Transcription-coupled nucleotide excision repair in mammalian cells: molecular mechanisms and biological effects

2008

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The encounter of elongating RNA polymerase II (RNAPIIo) with DNA lesions has severe consequences for the cell as this event provides a strong signal for P53-dependent apoptosis and cell cycle arrest. To counteract prolonged blockage of transcription, the cell removes the RNAPIIo-blocking DNA lesions by transcription-coupled repair (TC-NER), a specialized subpathway of nucleotide excision repair (NER). Exposure of mice to UVB light or chemicals has elucidated that TC-NER is a critical survival pathway protecting against acute toxic and long-term effects (cancer) of genotoxic exposure. Deficiency in TC-NER is associated with mutations in the CSA and CSB genes giving rise to the rare human disorder Cockayne syndrome (CS). Recent data suggest that CSA and CSB play differential roles in mammalian TC-NER: CSB as a repair coupling factor to attract NER proteins, chromatin remodellers and the CSA-E3-ubiquitin ligase complex to the stalled RNAPIIo. CSA is dispensable for attraction of NER proteins, yet in cooperation with CSB is required to recruit XAB2, the nucleosomal binding protein HMGN1 and TFIIS. The emerging picture of TC-NER is complex: repair of transcription-blocking lesions occurs without displacement of the DNA damage-stalled RNAPIIo, and requires at least two essential assembly factors (CSA and CSB), the core NER factors (except for XPC-RAD23B), and TC-NER specific factors. These and yet unidentified proteins will accomplish not only efficient repair of transcription-blocking lesions, but are also likely to contribute to DNA damage signalling events.

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“…T ranscription elongation is a processive but discontinuous process, with active synthesis of mRNA punctuated by transient pauses (1)(2)(3)(4). Both bacteria and eukaryotes control gene expression in vivo by regulating pausing at specific sites on the DNA template (5,6).…”

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confidence: 99%

Single molecule analysis of RNA polymerase elongation reveals uniform kinetic behavior

Adelman

Porta

Santangelo

et al. 2002

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

188

203

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By using single-molecule measurements, we demonstrate that the elongation kinetics of individual Escherichia coli RNA polymerase molecules are remarkably homogeneous. We find no evidence of distinct elongation states among RNA polymerases. Instead, the observed heterogeneity in transcription rates results from statistical variation in the frequency and duration of pausing. When transcribing a gene without strong pause sites, RNA polymerase molecules display transient pauses that are distributed randomly in both time and distance. Transitions between the active elongation mode and the paused state are instantaneous within the resolution of our measurements (<1 s). This elongation behavior is compared with that of a mutant RNA polymerase that pauses more frequently and elongates more slowly than wild type.T ranscription elongation is a processive but discontinuous process, with active synthesis of mRNA punctuated by transient pauses (1-4). Both bacteria and eukaryotes control gene expression in vivo by regulating pausing at specific sites on the DNA template (5, 6). Several regulatory pauses lead to long-lived isomerizations of the elongation complex, allowing transcription factors to bind the paused RNA polymerase (RNAP) and modify subsequent elongation (6-8). However, not all hesitation by the RNAP is thought to be regulatory; transcription of any naturally occurring template reveals numerous pauses with short half-lives, reflecting a distinct type of pausing that is inherent to the RNAP enzyme mechanism. At present, little is known about the RNAP configurations during these brief pauses or what governs the transition from active elongation to a paused state in the absence of specific regulatory signals.These dynamic aspects of elongation are obscured when studying a population of RNAP molecules in which the asynchronous behavior of individual RNAPs is smeared into an ensemble-average. Analyzing the motion of single RNAP molecules in real time eliminates the complication of population dynamics, revealing the kinetic interplay between active elongation and nonproductive states (9-12). Furthermore, continuous tracking of individual RNAPs exposes any variation in the behavior of single RNAP molecules as well as the differences that exist between RNAPs.There is more asynchrony during transcription elongation than would be generated by an enzymatic reaction with a single rate-limiting step, but the mechanisms leading to the additional asynchrony have yet to be clearly defined (13). Transcriptional pausing, where a fraction of RNAP stop at discrete sites for a variable duration, is known to induce dispersion of the population. However, whether or not the stochastic behavior of a structurally homogeneous population is sufficient to generate the observed levels of asynchrony, or if one must also invoke alternate stable RNAP conformations is a subject of debate. A recent study of single RNAP molecules suggests that an elongating RNAP population is composed of RNAPs in distinct states that elongate at different intrinsic ...

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Transcription elongation: structural basis and mechanisms

Cited by 99 publications

References 92 publications

Intrinsic Transcript Cleavage in Yeast RNA Polymerase II Elongation Complexes

Intrinsic Transcript Cleavage in Yeast RNA Polymerase II Elongation Complexes

Transcription-coupled nucleotide excision repair in mammalian cells: molecular mechanisms and biological effects

Single molecule analysis of RNA polymerase elongation reveals uniform kinetic behavior

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