scholarly journals Modification of Ser 7 of the RNA Polymerase II C‐terminal domain (CTD) regulates CTD phosphorylation patterns and transcription

2019 ◽  
Vol 33 (S1) ◽  
Author(s):  
Nathaniel Tate Burkholder ◽  
Sarah Sipe ◽  
Mukesh Kumar ◽  
Edwin Escobar ◽  
Seema Irani ◽  
...  
Keyword(s):  
Rna Polymerase ◽  
Rna Polymerase Ii ◽  
Terminal Domain ◽  
Ctd Phosphorylation

scholarly journals Sub1 Globally Regulates RNA Polymerase II C-Terminal Domain Phosphorylation

2010 ◽  
Vol 30 (21) ◽  
pp. 5180-5193 ◽  
Author(s):  
Alicia García ◽  
Emanuel Rosonina ◽  
James L. Manley ◽  
Olga Calvo
Keyword(s):  
Rna Polymerase ◽  
Rna Polymerase Ii ◽  
Large Subunit ◽  
Terminal Domain ◽  
Mrna Metabolism ◽  
Genes Encoding ◽  
The One ◽  
Ctd Phosphorylation ◽  
Transcription Cycle

ABSTRACT The transcriptional coactivator Sub1 has been implicated in several aspects of mRNA metabolism in yeast, such as activation of transcription, termination, and 3′-end formation. Here, we present evidence that Sub1 plays a significant role in controlling phosphorylation of the RNA polymerase II large subunit C-terminal domain (CTD). We show that SUB1 genetically interacts with the genes encoding all four known CTD kinases, SRB10, KIN28, BUR1, and CTK1, suggesting that Sub1 acts to influence CTD phosphorylation at more than one step of the transcription cycle. To address this directly, we first used in vitro kinase assays, and we show that, on the one hand, SUB1 deletion increased CTD phosphorylation by Kin28, Bur1, and Ctk1 but, on the other, it decreased CTD phosphorylation by Srb10. Second, chromatin immunoprecipitation assays revealed that SUB1 deletion decreased Srb10 chromatin association on the inducible GAL1 gene but increased Kin28 and Ctk1 chromatin association on actively transcribed genes. Taken together, our data point to multiple roles for Sub1 in the regulation of CTD phosphorylation throughout the transcription cycle.

scholarly journals RNA Polymerase II Carboxy-Terminal Domain Phosphorylation Is Required for Cotranscriptional Pre-mRNA Splicing and 3′-End Formation

2004 ◽  
Vol 24 (20) ◽  
pp. 8963-8969 ◽  
Author(s):  
Gregory Bird ◽  
Diego A. R. Zorio ◽  
David L. Bentley
Keyword(s):  
Rna Polymerase ◽  
Rna Polymerase Ii ◽  
Kinase Inhibitors ◽  
Mrna Splicing ◽  
Pol Ii ◽  
Terminal Domain ◽  
Carboxy Terminal Domain ◽  
Carboxy Terminal ◽  
Ctd Phosphorylation

ABSTRACT We investigated the role of RNA polymerase II (pol II) carboxy-terminal domain (CTD) phosphorylation in pre-mRNA processing coupled and uncoupled from transcription in Xenopus oocytes. Inhibition of CTD phosphorylation by the kinase inhibitors 5,6-dichloro-1β-d-ribofuranosyl-benzimidazole and H8 blocked transcription-coupled splicing and poly(A) site cleavage. These experiments suggest that pol II CTD phosphorylation is required for efficient pre-mRNA splicing and 3′-end formation in vivo. In contrast, processing of injected pre-mRNA was unaffected by either kinase inhibitors or α-amanitin-induced depletion of pol II. pol II therefore does not appear to participate directly in posttranscriptional processing, at least in frog oocytes. Together these experiments show that the influence of the phosphorylated CTD on pre-mRNA splicing and 3′-end processing is mediated by transcriptional coupling.

scholarly journals The Myc Transactivation Domain Promotes Global Phosphorylation of the RNA Polymerase II Carboxy-Terminal Domain Independently of Direct DNA Binding

2007 ◽  
Vol 27 (6) ◽  
pp. 2059-2073 ◽  
Author(s):  
Victoria H. Cowling ◽  
Michael D. Cole
Keyword(s):  
Dna Binding ◽  
Rna Polymerase ◽  
Rna Polymerase Ii ◽  
Transcription Initiation ◽  
Target Genes ◽  
Dependent Manner ◽  
Transactivation Domain ◽  
Pol Ii ◽  
Terminal Domain ◽  
Ctd Phosphorylation

ABSTRACT Myc is a transcription factor which is dependent on its DNA binding domain for transcriptional regulation of target genes. Here, we report the surprising finding that Myc mutants devoid of direct DNA binding activity and Myc target gene regulation can rescue a substantial fraction of the growth defect in myc −/− fibroblasts. Expression of the Myc transactivation domain alone induces a transcription-independent elevation of the RNA polymerase II (Pol II) C-terminal domain (CTD) kinases cyclin-dependent kinase 7 (CDK7) and CDK9 and a global increase in CTD phosphorylation. The Myc transactivation domain binds to the transcription initiation sites of these promoters and stimulates TFIIH binding in an MBII-dependent manner. Expression of the Myc transactivation domain increases CDK mRNA cap methylation, polysome loading, and the rate of translation. We find that some traditional Myc transcriptional target genes are also regulated by this Myc-driven translation mechanism. We propose that Myc transactivation domain-driven RNA Pol II CTD phosphorylation has broad effects on both transcription and mRNA metabolism.

scholarly journals Tyr1 phosphorylation promotes the phosphorylation of Ser2 on the C-terminal domain of RNA polymerase II by P-TEFb

2019 ◽  
Author(s):  
Joshua E. Mayfield ◽  
Seema Irani ◽  
Edwin E. Escobar ◽  
Zhao Zhang ◽  
Nathanial T. Burkholder ◽  
...  
Keyword(s):  
Rna Polymerase ◽  
Rna Polymerase Ii ◽  
Transcription Initiation ◽  
Elongation Factor ◽  
Mass Spectrometry Analysis ◽  
Spectrometry Analysis ◽  
Terminal Domain ◽  
Ctd Phosphorylation

SummaryThe Positive Transcription Elongation Factor b (P-TEFb) phosphorylates Ser2 residues of RNA polymerase II’s C-terminal domain (CTD) and is essential for the transition from transcription initiation to elongation in vivo. Surprisingly, P-TEFb exhibits Ser5 phosphorylation activity in vitro. The mechanism garnering Ser2 specificity to P-TEFb remains elusive and hinders understanding of the transition from transcription initiation to elongation. Through in vitro reconstruction of CTD phosphorylation, mass spectrometry analysis, and chromatin immunoprecipitation sequencing (ChIP-seq) analysis we uncover a mechanism by which Tyr1 phosphorylation directs the kinase activity of P-TEFb and alters its specificity from Ser5 to Ser2. The loss of Tyr1 phosphorylation causes a reduction of phosphorylated Ser2 and accumulation of RNA polymerase II in the promoter region as detected by ChIP-seq. We demonstrate the ability of Tyr1 phosphorylation to generate a heterogeneous CTD modification landscape that expands the CTD’s coding potential. These findings provide direct experimental evidence for a combinatorial CTD phosphorylation code wherein previously installed modifications direct the identity and abundance of subsequent coding events by influencing the behavior of downstream enzymes.

scholarly journals BRCA1 Can Modulate RNA Polymerase II Carboxy-Terminal Domain Phosphorylation Levels

2004 ◽  
Vol 24 (16) ◽  
pp. 6947-6956 ◽  
Author(s):  
Annie Moisan ◽  
Chantal Larochelle ◽  
Benoît Guillemette ◽  
Luc Gaudreau
Keyword(s):  
Rna Polymerase ◽  
Rna Polymerase Ii ◽  
Terminal Region ◽  
Terminal Domain ◽  
Carboxy Terminal Domain ◽  
Hcc1937 Cell ◽  
Carboxy Terminal ◽  
Cycle Regulation ◽  
Ctd Phosphorylation

ABSTRACT A high incidence of breast and ovarian cancers has been linked to mutations in the BRCA1 gene. BRCA1 has been shown to be involved in both positive and negative regulation of gene activity as well as in numerous other processes such as DNA repair and cell cycle regulation. Since modulation of the RNA polymerase II carboxy-terminal domain (CTD) phosphorylation levels could constitute an interface to all these functions, we wanted to directly test the possibility that BRCA1 might regulate the phosphorylation state of the CTD. We have shown that the BRCA1 C-terminal region can negatively modulate phosphorylation levels of the RNA polymerase II CTD by the Cdk-activating kinase (CAK) in vitro. Interestingly, the BRCA1 C-terminal region can directly interact with CAK and inhibit CAK activity by competing with ATP. Finally, we demonstrated that full-length BRCA1 can inhibit CTD phosphorylation when introduced in the BRCA1−/− HCC1937 cell line. Our results suggest that BRCA1 could play its ascribed roles, at least in part, by modulating CTD kinase components.

scholarly journals Tyr1 phosphorylation promotes phosphorylation of Ser2 on the C-terminal domain of eukaryotic RNA polymerase II by P-TEFb

eLife ◽  
2019 ◽  
Vol 8 ◽  
Author(s):  
Joshua E Mayfield ◽  
Seema Irani ◽  
Edwin E Escobar ◽  
Zhao Zhang ◽  
Nathaniel T Burkholder ◽  
...  
Keyword(s):  
Rna Polymerase ◽  
Rna Polymerase Ii ◽  
Transcription Initiation ◽  
Elongation Factor ◽  
Mass Spectrometry Analysis ◽  
Spectrometry Analysis ◽  
Terminal Domain ◽  
Ctd Phosphorylation

The Positive Transcription Elongation Factor b (P-TEFb) phosphorylates Ser2 residues of the C-terminal domain (CTD) of the largest subunit (RPB1) of RNA polymerase II and is essential for the transition from transcription initiation to elongation in vivo. Surprisingly, P-TEFb exhibits Ser5 phosphorylation activity in vitro. The mechanism garnering Ser2 specificity to P-TEFb remains elusive and hinders understanding of the transition from transcription initiation to elongation. Through in vitro reconstruction of CTD phosphorylation, mass spectrometry analysis, and chromatin immunoprecipitation sequencing (ChIP-seq) analysis, we uncover a mechanism by which Tyr1 phosphorylation directs the kinase activity of P-TEFb and alters its specificity from Ser5 to Ser2. The loss of Tyr1 phosphorylation causes an accumulation of RNA polymerase II in the promoter region as detected by ChIP-seq. We demonstrate the ability of Tyr1 phosphorylation to generate a heterogeneous CTD modification landscape that expands the CTD’s coding potential. These findings provide direct experimental evidence for a combinatorial CTD phosphorylation code wherein previously installed modifications direct the identity and abundance of subsequent coding events by influencing the behavior of downstream enzymes.

Regulated phosphorylation of the RNA polymerase II C-terminal domain (CTD)

1999 ◽  
Vol 77 (4) ◽  
pp. 249-255 ◽  
Author(s):  
Olivier Bensaude ◽  
François Bonnet ◽  
Céline Cassé ◽  
Marie-Françoise Dubois ◽  
Van Trung Nguyen ◽  
...  
Keyword(s):  
Rna Polymerase ◽  
Rna Polymerase Ii ◽  
Actinomycin D ◽  
Developmental Stages ◽  
Mrna Synthesis ◽  
Cyclin Dependent Kinases ◽  
Terminal Domain ◽  
Transcription Inhibitors ◽  
Carboxyl Terminal ◽  
Ctd Phosphorylation

The largest subunit of RNA polymerase II has an intriguing feature in its carboxyl-terminal domain (CTD) that consists of multiple repeats of an evolutionary conserved motif of seven amino acids. CTD phosphorylation plays a pivotal role in controlling mRNA synthesis and maturation. In exponentially growing cells, the phosphate turnover on the CTD is fast; it is blocked by common inhibitors of transcription, such as 5,6-dichloro-1-beta-D-ribofuranosylbenzimidazole and actinomycin D. Transcription-independent changes in CTD phosphorylation are observed at critical developmental stages, such as meiosis and early development.Key words: RNA polymerase II, phosphorylation, transcription inhibitors, cyclin-dependent kinases, development.

scholarly journals hnRNP U Inhibits Carboxy-Terminal Domain Phosphorylation by TFIIH and Represses RNA Polymerase II Elongation

1999 ◽  
Vol 19 (10) ◽  
pp. 6833-6844 ◽  
Author(s):  
Myung K. Kim ◽  
Vera M. Nikodem
Keyword(s):  
Rna Polymerase ◽  
Rna Polymerase Ii ◽  
Early Stage ◽  
Pol Ii ◽  
Terminal Domain ◽  
Carboxy Terminal Domain ◽  
Hnrnp U ◽  
Carboxy Terminal ◽  
Ctd Phosphorylation

ABSTRACT This study describes a potential new function of hnRNP U as an RNA polymerase (Pol II) elongation inhibitor. We demonstrated that a subfraction of human hnRNP U is associated with the Pol II holoenzyme in vivo and as such recruited to the promoter as part of the preinitiation complex. hnRNP U, however, appears to dissociate from the Pol II complex at the early stage of transcription and is therefore absent from the elongating Pol II complex. When tested in the human immunodeficiency virus type 1 transcription system, hnRNP U inhibits elongation rather than initiation of transcription by Pol II. This inhibition requires the carboxy-terminal domain (CTD) of Pol II. We showed that hnRNP U can bind TFIIH in vivo under certain conditions and inhibit TFIIH-mediated CTD phosphorylation in vitro. We find that the middle domain of hnRNP U is sufficient to mediate its Pol II association and its inhibition of TFIIH-mediated phosphorylation and Pol II elongation. The abilities of hnRNP U to inhibit TFIIH-mediated CTD phosphorylation and its Pol II association are necessary for hnRNP U to mediate the repression of Pol II elongation. Based on these observations, we suggest that a subfraction of hnRNP U, as a component of the Pol II holoenzyme, may downregulate TFIIH-mediated CTD phosphorylation in the basal transcription machinery and repress Pol II elongation. With such functions, hnRNP U might provide one of the mechanisms by which the CTD is maintained in an unphosphorylated state in the Pol II holoenzyme.

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