Roles of RNA:DNA hybrid stability, RNA structure, and active site conformation in pausing by human RNA polymerase II

2001 ◽  
Vol 311 (2) ◽  
pp. 265-282 ◽  
Author(s):  
Murali Palangat ◽  
Robert Landick
Keyword(s):  
Rna Polymerase ◽  
Rna Polymerase Ii ◽  
Active Site ◽  
Rna Structure ◽  
Rna:Dna Hybrid ◽  
Active Site Conformation

Characterization of the active site of yeast RNA polymerase II by DFT and ReaxFF calculations

2008 ◽  
Vol 120 (4-6) ◽  
pp. 479-489 ◽  
Author(s):  
Rui Zhu ◽  
Florian Janetzko ◽  
Yue Zhang ◽  
Adri C. T. van Duin ◽  
William A. Goddard ◽  
...  
Keyword(s):  
Rna Polymerase ◽  
Rna Polymerase Ii ◽  
Active Site

scholarly journals The Non-Coding B2 RNA Binds to the DNA Cleft and Active-Site Region of RNA Polymerase II

2013 ◽  
Vol 425 (19) ◽  
pp. 3625-3638 ◽  
Author(s):  
Steven L. Ponicsan ◽  
Stephane Houel ◽  
William M. Old ◽  
Natalie G. Ahn ◽  
James A. Goodrich ◽  
...  
Keyword(s):  
Rna Polymerase ◽  
Rna Polymerase Ii ◽  
Active Site ◽  
B2 Rna ◽  
Active Site Region

scholarly journals mRNA adenosine methylase (MTA) deposits m6A on pri-miRNAs to modulate miRNA biogenesis inArabidopsis thaliana

2020 ◽  
Vol 117 (35) ◽  
pp. 21785-21795 ◽  
Author(s):  
Susheel Sagar Bhat ◽  
Dawid Bielewicz ◽  
Tomasz Gulanicz ◽  
Zsuzsanna Bodi ◽  
Xiang Yu ◽  
...  
Keyword(s):  
Secondary Structure ◽  
Rna Polymerase ◽  
Rna Polymerase Ii ◽  
Rna Structure ◽  
Methylation Status ◽  
Mirna Biogenesis ◽  
Stem Loop ◽  
Structure Probing ◽  
Plant Transcriptome ◽  
Loop Regions

InArabidopsis thaliana, the METTL3 homolog, mRNA adenosine methylase (MTA) introducesN6-methyladenosine (m6A) into various coding and noncoding RNAs of the plant transcriptome. Here, we show that an MTA-deficient mutant (mta) has decreased levels of microRNAs (miRNAs) but accumulates primary miRNA transcripts (pri-miRNAs). Moreover, pri-miRNAs are methylated by MTA, and RNA structure probing analysis reveals a decrease in secondary structure within stem–loop regions of these transcripts inmtamutant plants. We demonstrate interaction between MTA and both RNA Polymerase II and TOUGH (TGH), a plant protein needed for early steps of miRNA biogenesis. Both MTA and TGH are necessary for efficient colocalization of the Microprocessor components Dicer-like 1 (DCL1) and Hyponastic Leaves 1 (HYL1) with RNA Polymerase II. We propose that secondary structure of miRNA precursors induced by their MTA-dependent m6A methylation status, together with direct interactions between MTA and TGH, influence the recruitment of Microprocessor to plant pri-miRNAs. Therefore, the lack of MTA inmtamutant plants disturbs pri-miRNA processing and leads to the decrease in miRNA accumulation. Furthermore, our findings reveal that reduced miR393b levels likely contributes to the impaired auxin response phenotypes ofmtamutant plants.

NTP-driven translocation and regulation of downstream template opening by multi-subunit RNA polymerases

2005 ◽  
Vol 83 (4) ◽  
pp. 486-496 ◽  
Author(s):  
Zachary F Burton ◽  
Michael Feig ◽  
Xue Q Gong ◽  
Chunfen Zhang ◽  
Yuri A Nedialkov ◽  
...  
Keyword(s):  
Rna Polymerase ◽  
Rna Polymerase Ii ◽  
Active Site ◽  
Binding Sites ◽  
Rna Synthesis ◽  
Rna Polymerases ◽  
Phosphoryl Transfer ◽  
Elongation Complex ◽  
Detailed Model ◽  
Loop 2

Multi-subunit RNA polymerases bind nucleotide triphosphate (NTP) substrates in the pretranslocated state and carry the dNMP–NTP base pair into the active site for phosphoryl transfer. NTP-driven translocation requires that NTP substrates enter the main-enzyme channel before loading into the active site. Based on this model, a new view of fidelity and efficiency of RNA synthesis is proposed. The model predicts that, during processive elongation, NTP-driven translocation is coupled to a protein conformational change that allows pyrophosphate release: coupling the end of one bond-addition cycle to substrate loading and translocation for the next. We present a detailed model of the RNA polymerase II elongation complex based on 2 low-affinity NTP binding sites located in the main-enzyme channel. This model posits that NTP substrates, elongation factors, and the conserved Rpb2 subunit fork loop 2 cooperate to regulate opening of the downstream transcription bubble.Key words: RNA polymerase, NTP-driven translocation, transcriptional fidelity, transcriptional efficiency, α-amanitin.

scholarly journals Transient Reversal of RNA Polymerase II Active Site Closing Controls Fidelity of Transcription Elongation

2008 ◽  
Vol 30 (5) ◽  
pp. 557-566 ◽  
Author(s):  
Maria L. Kireeva ◽  
Yuri A. Nedialkov ◽  
Gina H. Cremona ◽  
Yuri A. Purtov ◽  
Lucyna Lubkowska ◽  
...  
Keyword(s):  
Rna Polymerase ◽  
Rna Polymerase Ii ◽  
Active Site ◽  
Transcription Elongation

scholarly journals Translocation after Synthesis of a Four-Nucleotide RNA Commits RNA Polymerase II to Promoter Escape

2002 ◽  
Vol 22 (3) ◽  
pp. 762-773 ◽  
Author(s):  
Jennifer F. Kugel ◽  
James A. Goodrich
Keyword(s):  
Rna Polymerase ◽  
Rna Polymerase Ii ◽  
Active Site ◽  
Molecular Mechanisms ◽  
Sequence Specificity ◽  
Promoter Escape ◽  
General Transcription Factors ◽  
Dna Elements ◽  
Promoter Dna ◽  
Transcription Complexes

ABSTRACT Transcription is a complex process, the regulation of which is crucial for cellular and organismic growth and development. Deciphering the molecular mechanisms that define transcription is essential to understanding the regulation of RNA synthesis. Here we describe the molecular mechanism of escape commitment, a critical step in early RNA polymerase II transcription. During escape commitment ternary transcribing complexes become stable and committed to proceeding forward through promoter escape and the remainder of the transcription reaction. We found that the point in the transcription reaction at which escape commitment occurs depends on the length of the transcript RNA (4 nucleotides [nt]) as opposed to the position of the active site of the polymerase with respect to promoter DNA elements. We found that single-stranded nucleic acids can inhibit escape commitment, and we identified oligonucleotides that are potent inhibitors of this specific step. These inhibitors bind RNA polymerase II with low nanomolar affinity and sequence specificity, and they block both promoter-dependent and promoter-independent transcription, the latter occurring in the absence of general transcription factors. We demonstrate that escape commitment involves translocation of the RNA polymerase II active site between synthesis of the third and fourth phosphodiester bonds. We propose that a conformational change in ternary transcription complexes occurs during translocation after synthesis of a 4-nt RNA to render complexes escape committed.

scholarly journals Mechanism of RNA polymerase II bypass of oxidative cyclopurine DNA lesions

2015 ◽  
Vol 112 (5) ◽  
pp. E410-E419 ◽  
Author(s):  
Celine Walmacq ◽  
Lanfeng Wang ◽  
Jenny Chong ◽  
Kathleen Scibelli ◽  
Lucyna Lubkowska ◽  
...  
Keyword(s):  
Rna Polymerase ◽  
Rna Polymerase Ii ◽  
Active Site ◽  
Dna Lesions ◽  
Abasic Site ◽  
Pol Ii ◽  
Dna Lesion ◽  
Trigger Loop ◽  
Pyrimidine Dimers ◽  
Bulky Dna Lesions

In human cells, the oxidative DNA lesion 8,5′-cyclo-2'-deoxyadenosine (CydA) induces prolonged stalling of RNA polymerase II (Pol II) followed by transcriptional bypass, generating both error-free and mutant transcripts with AMP misincorporated immediately downstream from the lesion. Here, we present biochemical and crystallographic evidence for the mechanism of CydA recognition. Pol II stalling results from impaired loading of the template base (5′) next to CydA into the active site, leading to preferential AMP misincorporation. Such predominant AMP insertion, which also occurs at an abasic site, is unaffected by the identity of the 5′-templating base, indicating that it derives from nontemplated synthesis according to an A rule known for DNA polymerases and recently identified for Pol II bypass of pyrimidine dimers. Subsequent to AMP misincorporation, Pol II encounters a major translocation block that is slowly overcome. Thus, the translocation block combined with the poor extension of the dA.rA mispair reduce transcriptional mutagenesis. Moreover, increasing the active-site flexibility by mutation in the trigger loop, which increases the ability of Pol II to accommodate the bulky lesion, and addition of transacting factor TFIIF facilitate CydA bypass. Thus, blocking lesion entry to the active site, translesion A rule synthesis, and translocation block are common features of transcription across different bulky DNA lesions.

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