Kinetic and mechanistic analysis of the RNA polymerase II transcription reaction at the human interleukin-2 promoter

Ferguson, Heather A.; Kugel, Jennifer F.; Goodrich, James A.

doi:10.1006/jmbi.2000.5215

Cited by 21 publications

(14 citation statements)

References 70 publications

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“…Even in the simplest phage systems, this transition involves a complicated structural reorganization of the template-bound transcription machinery (65,77). On some RNA polymerase II promoters in vitro, promoter escape, and not preinitiation complex formation, is the slow step in transcript synthesis (14,33,34). The data in this work indicate that in vivo, functional TFIIH expands the zone at the endogenous c-myc promoter in which nascent transcript extension is regulated.…”

Section: What Does Tfiih Do In Transcription?mentioning

confidence: 76%

“…After initiation, the transcription apparatus, without proper TFIIH, supports incremental transcript growth with pauses occurring at several, possibly variable, positions until promoter escape occurs early, well within the first 50 nucleotides. On some promoters in vitro, multiple sequential postinitiation, prepromoter escape transcription pauses have been observed; the exact number and sites of pausing are sequence dependent (14,51,52). If preinitiation complex formation is rapid on the c-myc promoter, as all evidence suggests, then promoter escape may be imagined as a queue of promoter-specific pauses.…”

Section: What Does Tfiih Do In Transcription?mentioning

confidence: 99%

See 1 more Smart Citation

TFIIH Operates through an Expanded Proximal Promoter To Fine-Tune c-myc Expression

Weber

Liu

Collins

et al. 2005

Molecular and Cellular Biology

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Section: What Does Tfiih Do In Transcription?mentioning

confidence: 76%

Section: What Does Tfiih Do In Transcription?mentioning

confidence: 99%

TFIIH Operates through an Expanded Proximal Promoter To Fine-Tune c-myc Expression

Weber

Liu

Collins

et al. 2005

Molecular and Cellular Biology

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“…We previously found that at the interleukin-2 promoter the rate at which complexes assembled on promoter-bound TFIID-TFIIA also limited the rate of transcription; however, in these experiments, increasing the concentration of Pol II-TFIIF increased the rate of complex formation, indicating that a recruitment event as opposed to an isomerization was rate limiting. 50 Hence, it seems likely that the core promoter elements and/or sequences that TAFs contact affect the rate at which TAFs can release those contacts. Our studies reveal that TAFs kinetically repress the formation of active complexes at promoters, and suggest another means with which to finely tune the transcriptional output.…”

Section: Discussionmentioning

confidence: 99%

RNA Polymerase II and TAFs Undergo a Slow Isomerization after the Polymerase Is Recruited to Promoter-Bound TFIID

Yakovchuk

Gilman

Goodrich

et al. 2010

Journal of Molecular Biology

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“…all steps that occur after the addition of nucleotides to preinitiation complexes) was significantly slower at the AdMLP than at the IL-2P in a transcription system reconstituted from TFIIA, TFIID, TFIIB, TFIIF, Pol II, TFIIE, and TFIIH (23). Our goals were to understand what properties of the IL-2P and AdMLP cause different rates of transcript synthesis and to further decipher the mechanism of promoter escape.…”

Section: The Early Transcribed Regions Of the Il-2p And Admlp Set Difmentioning

confidence: 99%

“…We previously found promoter escape to be the slowest of the transcript synthesis steps in vitro at both the adenovirus major late promoter (AdMLP) (9,13) and the interleukin-2 promoter (IL-2P) (23). There was, however, a significant difference in the rate constants for transcript synthesis at the two promoters.…”

mentioning

confidence: 99%

The Sequence at Specific Positions in the Early Transcribed Region Sets the Rate of Transcript Synthesis by RNA Polymerase II in Vitro

Weaver¹,

Kugel

Goodrich

2005

Journal of Biological Chemistry

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To further understand the mechanism of promoter escape by RNA polymerase II, we have systematically investigated the effect of core promoter sequence on the rate of transcript synthesis in vitro. Chimeric and mutant promoters were made by swapping sequences between the human interleukin-2 promoter and the adenovirus major late promoter, which exhibit different rates of transcript synthesis. Kinetic studies at these promoters revealed that sequences downstream of the start sites set the rate of transcript synthesis. Specifically, the sequences at ؉2 and ؉7/؉8 are critical for determining the rate; when either ؉2 is a C (nontemplate strand) or ؉7/؉8 is a TT (nontemplate strand), transcript synthesis is slow. At ؉7/؉8, the thermodynamic stability of the RNA:DNA hybrid controls the overall rate of transcript synthesis. Our data support a model in which the rate-limiting step during transcript synthesis by RNA polymerase II in vitro occurs at the point in the reaction at which early ternary complexes transform into elongation complexes.Eukaryotic mRNA transcription is a multistep reaction involving numerous protein factors and DNA elements. RNA polymerase II (Pol II) 4 catalyzes mRNA synthesis; however, promoter-specific initiation of transcription requires additional proteins known as the general transcription factors, including TFIIA, TFIIB, TFIID, TFIIE, TFIIF, and TFIIH (1). In addition, a multitude of other factors, including regulators, co-regulators, chromatin, and chromatin modifying factors, work in concert with Pol II and the general transcription factors to regulate levels of transcription (2-4). The DNA elements that control mRNA transcription are contained in the promoters of genes, which include regulatory elements that bind activators and repressors and core promoter elements that bind Pol II and the general transcription factors.During the early stages of transcript synthesis unstable initiated complexes that synthesize 2-and 3-nt RNAs (5-9) transform into stable elongation complexes that complete synthesis of the transcript (9, 10). This transition occurs during promoter escape, and involves a complex series of molecular transformations including changes in the melted region of DNA, the release of general transcription factors, and formation of the full-length RNA:DNA hybrid (6, 9, 11-15). The primary transformations in the DNA involve the formation and initial movement of the transcription bubble. Permanganate footprinting studies showed that the DNA melts from Ϫ9 to ϩ2 upon assembly of the preinitiation complex (6). Early transcript synthesis then results in continuous extension of the downstream edge of the melted region. By synthesis of an 11-nt RNA the upstream edge of the transcription bubble re-anneals through at least Ϫ2 (6). Recent studies indicate that this re-annealing cannot occur until the bubble is 18 residues in length and a 7-nt RNA is synthesized (15). In addition to the DNA, the protein components of the preinitiation complex change en route to formation of elongation complexes. Fo...

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Kinetic and mechanistic analysis of the RNA polymerase II transcription reaction at the human interleukin-2 promoter

Cited by 21 publications

References 70 publications

TFIIH Operates through an Expanded Proximal Promoter To Fine-Tune c-myc Expression

TFIIH Operates through an Expanded Proximal Promoter To Fine-Tune c-myc Expression

RNA Polymerase II and TAFs Undergo a Slow Isomerization after the Polymerase Is Recruited to Promoter-Bound TFIID

The Sequence at Specific Positions in the Early Transcribed Region Sets the Rate of Transcript Synthesis by RNA Polymerase II in Vitro

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