scholarly journals Single-Stranded Breaks in DNA but Not Oxidative DNA Base Damages Block Transcriptional Elongation by RNA Polymerase II in HeLa Cell Nuclear Extracts

2004 ◽  
Vol 279 (18) ◽  
pp. 18511-18520 ◽  
Author(s):  
Scott D. Kathe ◽  
Guang-Ping Shen ◽  
Susan S. Wallace
Keyword(s):  
Rna Polymerase ◽  
Hela Cell ◽  
Rna Polymerase Ii ◽  
Transcriptional Elongation ◽  
Nuclear Extracts ◽  
Dna Base

scholarly journals Transcription of Hepatitis Delta Virus RNA by RNA Polymerase II

2007 ◽  
Vol 82 (3) ◽  
pp. 1118-1127 ◽  
Author(s):  
Jinhong Chang ◽  
Xingcao Nie ◽  
Ho Eun Chang ◽  
Ziying Han ◽  
John Taylor
Keyword(s):  
Rna Polymerase ◽  
Rna Polymerase Ii ◽  
Hepatitis Delta Virus ◽  
Cell System ◽  
Hepatitis Delta ◽  
Pol Ii ◽  
Delta Antigen ◽  
Nuclear Extracts ◽  
Virus Rna ◽  
Delta Virus

ABSTRACT Previous studies have indicated that the replication of the RNA genome of hepatitis delta virus (HDV) involves redirection of RNA polymerase II (Pol II), a host enzyme that normally uses DNA as a template. However, there has been some controversy about whether in one part of this HDV RNA transcription, a polymerase other than Pol II is involved. The present study applied a recently described cell system (293-HDV) of tetracycline-inducible HDV RNA replication to provide new data regarding the involvement of host polymerases in HDV transcription. The data generated with a nuclear run-on assay demonstrated that synthesis not only of genomic RNA but also of its complement, the antigenome, could be inhibited by low concentrations of amanitin specific for Pol II transcription. Subsequent studies used immunoprecipitation and rate-zonal sedimentation of nuclear extracts together with double immunostaining of 293-HDV cells, in order to examine the associations between Pol II and HDV RNAs, as well as the small delta antigen, an HDV-encoded protein known to be essential for replication. Findings include evidence that HDV replication is somehow able to direct the available delta antigen to sites in the nucleoplasm, almost exclusively colocalized with Pol II in what others have described as transcription factories.

Identification of rpo30, a vaccinia virus RNA polymerase gene with structural similarity to a eucaryotic transcription elongation factor

1990 ◽  
Vol 10 (10) ◽  
pp. 5433-5441
Author(s):  
B Y Ahn ◽  
P D Gershon ◽  
E V Jones ◽  
B Moss
Keyword(s):  
Rna Polymerase ◽  
Vaccinia Virus ◽  
Rna Polymerase Ii ◽  
Elongation Factor ◽  
Viral Rna ◽  
In Vitro Translation ◽  
Virus Protein ◽  
Transcriptional Elongation

Eucaryotic transcription factors that stimulate RNA polymerase II by increasing the efficiency of elongation of specifically or randomly initiated RNA chains have been isolated and characterized. We have identified a 30-kilodalton (kDa) vaccinia virus-encoded protein with apparent homology to SII, a 34-kDa mammalian transcriptional elongation factor. In addition to amino acid sequence similarities, both proteins contain C-terminal putative zinc finger domains. Identification of the gene, rpo30, encoding the vaccinia virus protein was achieved by using antibody to the purified viral RNA polymerase for immunoprecipitation of the in vitro translation products of in vivo-synthesized early mRNA selected by hybridization to cloned DNA fragments of the viral genome. Western immunoblot analysis using antiserum made to the vaccinia rpo30 protein expressed in bacteria indicated that the 30-kDa protein remains associated with highly purified viral RNA polymerase. Thus, the vaccinia virus protein, unlike its eucaryotic homolog, is an integral RNA polymerase subunit rather than a readily separable transcription factor. Further studies showed that the expression of rpo30 is regulated by dual early and later promoters.

Localization of active promoters for eucaryotic RNA polymerase II in the long terminal repeat of avian sarcoma virus DNA

1983 ◽  
Vol 3 (5) ◽  
pp. 811-818
Author(s):  
S A Mitsialis ◽  
J L Manley ◽  
R V Guntaka
Keyword(s):  
Rna Polymerase ◽  
Hela Cell ◽  
Long Terminal Repeat ◽  
Rna Polymerase Ii ◽  
Terminal Repeat ◽  
Avian Sarcoma Virus ◽  
Cell Extracts ◽  
Sarcoma Virus ◽  
Avian Sarcoma

The nucleotide sequences in the long terminal repeat of avian sarcoma virus that are recognized in vitro by HeLa cell RNA polymerase II have been identified. For this purpose, various 5' and 3' deletions were introduced into a cloned long terminal repeat fragment. The effects of these deletions on transcription initiation in HeLa whole-cell extracts were then studied. Three specific transcripts have been identified. The major transcript is initiated at nucleotide +1 (relative to the cap site). Deletion of the upstream sequence between -299 and -55 has no effect on the level of transcription from this start site, whereas deletion of the sequence downstream of -14 drastically reduces the levels of transcription. In contrast, deletion of the sequence downstream from the TATA box has no effect on the initiation or efficiency of synthesis of the two minor RNA species, which are initiated at around nucleotides -260 and -105. The transcription of these RNA products, however, is abolished by an upstream deletion between -299 and -55. These results suggest that HeLa cell RNA polymerase II recognizes in vitro more than one promoter site present in the long terminal repeat of the avian sarcoma virus genome and defines the sequences required for initiation of the major transcript.

scholarly journals Mechanism and regulation of transcriptional elongation by RNA polymerase II

1999 ◽  
Vol 11 (3) ◽  
pp. 342-346 ◽  
Author(s):  
Daniel Reines ◽  
Ronald C Conaway ◽  
Joan Weliky Conaway
Keyword(s):  
Rna Polymerase ◽  
Rna Polymerase Ii ◽  
Transcriptional Elongation

scholarly journals GSK-3 is an RNA polymerase II phospho-CTD kinase

2020 ◽  
Vol 48 (11) ◽  
pp. 6068-6080 ◽  
Author(s):  
Nicolás Nieto Moreno ◽  
Florencia Villafañez ◽  
Luciana E Giono ◽  
Carmen Cuenca ◽  
Gastón Soria ◽  
...  
Keyword(s):  
Dna Damage ◽  
Rna Polymerase ◽  
Rna Polymerase Ii ◽  
Glycogen Synthase ◽  
Dominant Negative ◽  
Dominant Negative Mutant ◽  
Transcriptional Elongation ◽  
Induced Apoptosis ◽  
Carboxy Terminal

Abstract We have previously found that UV-induced DNA damage causes hyperphosphorylation of the carboxy terminal domain (CTD) of RNA polymerase II (RNAPII), inhibition of transcriptional elongation and changes in alternative splicing (AS) due to kinetic coupling between transcription and splicing. In an unbiased search for protein kinases involved in the AS response to DNA damage, we have identified glycogen synthase kinase 3 (GSK-3) as an unforeseen participant. Unlike Cdk9 inhibition, GSK-3 inhibition only prevents CTD hyperphosphorylation triggered by UV but not basal phosphorylation. This effect is not due to differential degradation of the phospho-CTD isoforms and can be reproduced, at the AS level, by overexpression of a kinase-dead GSK-3 dominant negative mutant. GSK-3 inhibition abrogates both the reduction in RNAPII elongation and changes in AS elicited by UV. We show that GSK-3 phosphorylates the CTD in vitro, but preferentially when the substrate is previously phosphorylated, consistently with the requirement of a priming phosphorylation reported for GSK-3 efficacy. In line with a role for GSK-3 in the response to DNA damage, GSK-3 inhibition prevents UV-induced apoptosis. In summary, we uncover a novel role for a widely studied kinase in key steps of eukaryotic transcription and pre-mRNA processing.

scholarly journals Localization of active promoters for eucaryotic RNA polymerase II in the long terminal repeat of avian sarcoma virus DNA.

1983 ◽  
Vol 3 (5) ◽  
pp. 811-818 ◽  
Author(s):  
S A Mitsialis ◽  
J L Manley ◽  
R V Guntaka
Keyword(s):  
Rna Polymerase ◽  
Hela Cell ◽  
Long Terminal Repeat ◽  
Rna Polymerase Ii ◽  
Terminal Repeat ◽  
Avian Sarcoma Virus ◽  
Cell Extracts ◽  
Sarcoma Virus ◽  
Avian Sarcoma

The nucleotide sequences in the long terminal repeat of avian sarcoma virus that are recognized in vitro by HeLa cell RNA polymerase II have been identified. For this purpose, various 5' and 3' deletions were introduced into a cloned long terminal repeat fragment. The effects of these deletions on transcription initiation in HeLa whole-cell extracts were then studied. Three specific transcripts have been identified. The major transcript is initiated at nucleotide +1 (relative to the cap site). Deletion of the upstream sequence between -299 and -55 has no effect on the level of transcription from this start site, whereas deletion of the sequence downstream of -14 drastically reduces the levels of transcription. In contrast, deletion of the sequence downstream from the TATA box has no effect on the initiation or efficiency of synthesis of the two minor RNA species, which are initiated at around nucleotides -260 and -105. The transcription of these RNA products, however, is abolished by an upstream deletion between -299 and -55. These results suggest that HeLa cell RNA polymerase II recognizes in vitro more than one promoter site present in the long terminal repeat of the avian sarcoma virus genome and defines the sequences required for initiation of the major transcript.

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