The nuclear matrix protein p255 is a highly phosphorylated form of RNA polymerase II largest subunit which associates with spliceosomes

Vincent, Michel; Lauriault, Pascal; Dubois, Marie-Françoise; Lavoie, Sébastien B.; Bensaude, Olivier; Chabot, Benoît

doi:10.1093/nar/24.23.4649

Cited by 84 publications

(74 citation statements)

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“…The splicing components that interact with the RNA Pol II-containing complex include SR proteins and snRNPs, factors that also associate with hyperphosphorylated RNA Pol II (20,22,23). In addition, we document for the first time the interaction of U2AF 65 with transcription complexes.…”

Section: Discussionmentioning

confidence: 83%

“…Truncation of the CTD leads to inefficient capping, polyadenylation, and splicing in cultured mammalian cells (16,19). Both the overexpression of phosphorylated CTD peptides (26) and the use of antibodies directed against the CTD have been shown to inhibit splicing (20,23,24). These results suggest that the phosphorylated CTD of RNA Pol II provides an interface for the recruitment of splicing factors.…”

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confidence: 99%

“…Whereas a phosphorylated CTD is essential for the interaction with capping enzymes, the polyadenylation factor CPSF is brought to the transcription complex through an interaction with TFIID and transferred to the phosphorylated CTD after transcription initiation (19,25). The phosphorylated RNA Pol IIO isoform has been found to associate with splicing factors and has been identified in active spliceosomes (21)(22)(23). Truncation of the CTD leads to inefficient capping, polyadenylation, and splicing in cultured mammalian cells (16,19).…”

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confidence: 99%

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A Human RNA Polymerase II-containing Complex Associated with Factors Necessary for Spliceosome Assembly

Robert¹,

Blanchette²,

Maës³

et al. 2002

Journal of Biological Chemistry

111

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Transcription and splicing are coordinated processes in mammalian cells. We have used affinity chromatography with immobilized transcription elongation factor SII to purify a protein complex that contains core RNA polymerase II (RNA Pol II), the general transcription initiation factors, and several splicing factors, including the U1, U2, and U4 small nuclear RNPs, the U2AF 65 , and serine/arginine-rich proteins. The splicing factors and the transcription machinery co-purify through a gel filtration column and co-immunoprecipitate in experiments using an anti-U2AF 65 antibody, indicating that they are part of a unique complex. Although the RNA Pol II-containing complex does not possess splicing activity, it can complement small nuclear RNP-inactivated extracts and can promote the formation of a pre-spliceosome complex. Because interactions between components of the splicing and transcription machineries occur in the context of a complex containing a hypophosphorylated RNA Pol II capable of initiating transcription, our results suggest that the coupling between transcription and splicing begins before transcription initiation.The splicing of pre-mRNA in mammalian cells is the nuclear process during which introns are removed from primary transcripts synthesized by RNA polymerase II (RNA Pol II).1 Splicing is achieved by the spliceosome, a large macromolecular complex, composed of small nuclear ribonucleoprotein (snRNP) particles and additional proteins including members of the serine/arginine-rich (SR) protein family (reviewed in Refs. 1 and 2). Cytological studies have revealed that splicing can occur in a co-transcriptional manner (see Refs. 3 and 4 for examples) and that factors necessary for splicing are recruited to sites of transcription (5-7). A growing body of data indicates that coupling between transcription and a variety of RNA maturation events may be achieved through the C-terminal domain (CTD) of the RPB1 subunit of RNA Pol II (reviewed in Refs. 8 -11). The CTD is a seven-amino acid motif tandemly repeated 52 times in humans and 26 -27 times in yeast. It is highly conserved among eukaryotic organisms and is subject to reversible phosphorylation during the transcription cycle (reviewed in Refs. 12 and 13). Two isoforms of RNA Pol II exist in vivo, namely RNA Pol IIA and IIO. RNA Pol IIA possesses a hypophosphorylated CTD and preferentially enters the preinitiation complex, whereas RNA Pol IIO harbors an extensively phosphorylated CTD and is found in the elongation complex (13,14). Biochemical studies indicate that RNA Pol II physically interacts with components involved in capping (15-18), polyadenylation (19), and splicing (20 -24). Whereas a phosphorylated CTD is essential for the interaction with capping enzymes, the polyadenylation factor CPSF is brought to the transcription complex through an interaction with TFIID and transferred to the phosphorylated CTD after transcription initiation (19,25). The phosphorylated RNA Pol IIO isoform has been found to associate with splicing factors and has been i...

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Section: Discussionmentioning

confidence: 83%

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confidence: 99%

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confidence: 99%

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A Human RNA Polymerase II-containing Complex Associated with Factors Necessary for Spliceosome Assembly

Robert¹,

Blanchette²,

Maës³

et al. 2002

Journal of Biological Chemistry

111

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“…The phosphorylated CTD binds the capping enzymes (Cho et al 1997;McCracken et al 1997a). CTD overexpression and CTD peptide addition to extracts (Yuryev et al 1996) influence splicing in HeLa cells, which, together with observations that the hyperphosphorylated polymerase II binds snRNPs and SR proteins (Chabot et al 1995;Mortarillo et al 1996;Vincent et al 1996;Kim et al 1997), indicates that spliceosomal components are also associated with RNA polymerase II.…”

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confidence: 83%

Transcriptional termination in the Balbiani ring 1 gene is closely coupled to 3′-end formation and excision of the 3′-terminal intron

Baurén

Беликов²,

Wieslander³

1998

Genes Dev.

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We have analyzed transcription termination, 3 -end formation, and excision of the 3 -terminal intron in vivo in the Balbiani ring 1 (BR1) gene and its pre-mRNA. We show that full-length RNA transcripts are evenly spaced on the gene from a position 300 bp upstream to a region 500-700 bp downstream of the polyadenylation sequence. Very few full-length transcripts and no short, cleaved, nascent transcripts could be observed downstream of this region. Pre-mRNA with 10-20 adenylate residues accumulates at the active gene and then rapidly leaves from the gene locus. Only polyadenylated pre-mRNAs could be detected in the nucleoplasm. Our results are consistent with the hypothesis that transcription termination occurs in a narrow region for the majority of transcripts, simultaneous with 3 -end formation. Excision of the 3 -terminal intron occurs before 3 -end formation in about 5% of the nascent transcripts. When transcription terminates, 3 cleavage takes place and 10-20 adenylate residues are added, the 3 -terminal intron is excised from additionally about 75% of the pre-mRNA at the gene locus. Our data support a close temporal and spatial coupling of transcription termination and the cleavage and initial polyadenylation of 3 -end formation. Excision of the 3 -terminal intron is highly stimulated as the cleavage/polyadenylation complex assembles and 3 -end formation is initiated.

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“…A variety of evidence strongly suggests that the phospho-CTD is involved in splicing. The hyperphosphorylated form of RNA polymerase II colocalizes with splicing factors when transcriptionally active (15,16) and in the nuclear matrix (17)(18)(19). Several phospho-CTD binding proteins have been identified that contain SR and RRM domains as found in many splicing factors (20,21).…”

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confidence: 99%

The Splicing Factor, Prp40, Binds the Phosphorylated Carboxyl-terminal Domain of RNA Polymerase II

Morris

Greenleaf

2000

Journal of Biological Chemistry

164

165

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We showed previously that the WW domain of the prolyl isomerase, Ess1, can bind the phosphorylated carboxyl-terminal domain (phospho-CTD) of the largest subunit of RNA Polymerase II. Analysis of phospho-CTD binding by four other WW domain-containing Saccharomyces cerevisiae proteins indicates the splicing factor, Prp40, and the RNA polymerase II ubiquitin ligase, Rsp5, can also bind the phospho-CTD. The identification of Prp40 as a phospho-CTD binding protein represents the first demonstration of direct interaction between a documented splicing factor and the phospho-CTD. Domain dissection studies reveal that phospho-CTD binding occurs at multiple locations in Prp40, including sites in both the WW and FF domain regions. Because the conserved repeats of the CTD make it an ideal ligand for multi-site binding events, the implications of multi-site binding are discussed. Our data suggest a mechanism by which the phospho-CTD of elongating RNA polymerase II facilitates commitment complex formation by juxtaposing the 5 and 3 splice sites.The carboxyl-terminal domain (CTD) 1 of the largest subunit of RNA polymerase II (1) plays a central role in mRNA synthesis. The CTD is composed of 26 -52 heptad repeats with the consensus sequence, YSPTSPS. These repeats are extensively phosphorylated in RNA polymerase II actively engaged in transcript elongation (2-5).In the past several years evidence has accumulated indicating that the phosphorylated form of the CTD acts to coordinate pre-mRNA processing events (for review see Refs. 6 -10). The 5Ј capping complex is not only localized near the pre-mRNA by direct association with the phospho-CTD (11, 12) but is also allosterically activated by phospho-CTD binding (13,14). A variety of evidence strongly suggests that the phospho-CTD is involved in splicing. The hyperphosphorylated form of RNA polymerase II colocalizes with splicing factors when transcriptionally active (15, 16) and in the nuclear matrix (17-19). Several phospho-CTD binding proteins have been identified that contain SR and RRM domains as found in many splicing factors (20,21). Fusion proteins with a hyperphosphorylated CTD can inhibit splicing in vivo (22). In addition, transcriptionally unengaged but phosphorylated RNA polymerase II is able to stimulate splicing in vitro (23).Recently, we demonstrated that the prolyl isomerase, Ess1, can bind the phosphorylated form of the CTD and that this binding is mediated by its WW domain (24). Because the prolyl isomerase activity of Ess1 preferentially acts on phospho-SerPro peptide bonds, as are found in abundance in the phospho-CTD, our findings provided a plausible explanation for earlier results implicating Ess1 in pre-mRNA 3Ј end formation (25,26).WW domains, named for two highly conserved tryptophan residues, are small independently folding protein domains consisting of slightly more than 30 amino acids arranged in three anti-parallel ␤ sheets (27, 28). These domains have been shown to bind proline-rich sequences containing several different motifs (29). Of five protei...

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The nuclear matrix protein p255 is a highly phosphorylated form of RNA polymerase II largest subunit which associates with spliceosomes

Cited by 84 publications

References 34 publications

A Human RNA Polymerase II-containing Complex Associated with Factors Necessary for Spliceosome Assembly

A Human RNA Polymerase II-containing Complex Associated with Factors Necessary for Spliceosome Assembly

Transcriptional termination in the Balbiani ring 1 gene is closely coupled to 3′-end formation and excision of the 3′-terminal intron

The Splicing Factor, Prp40, Binds the Phosphorylated Carboxyl-terminal Domain of RNA Polymerase II

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