Interactions between fission yeast Cdk9, its cyclin partner Pch1, and mRNA capping enzyme Pct1 suggest an elongation checkpoint for mRNA quality control Journal Article
| Authors: | Pei, Y.; Schwer, B.; Shuman, S. |
| Article Title: | Interactions between fission yeast Cdk9, its cyclin partner Pch1, and mRNA capping enzyme Pct1 suggest an elongation checkpoint for mRNA quality control |
| Abstract: | RNA polymerase II (pol II) is subject to an early elongation delay induced by negative factors Spt5/Spt4 and NELF, which is overcome by the positive factor P-TEFb (Cdk9/cyclin T), a protein kinase that phosphorylates the pol II C-terminal domain (CTD) and the transcription elongation factor Spt5. Although the rationale for this arrest and restart is unclear, recent studies suggest a connection to mRNA capping, which is coupled to transcription elongation via physical and functional interactions between the cap-forming enzymes, the CTD-PO4, and Spt5. Here we identify a novel interaction between fission yeast RNA triphosphatase Pct1, the enzyme that initiates cap formation, and Schizosaccharomyces pombe Cdk9. The C-terminal segment of SpCdk9 comprises a Pct1-binding domain distinct from the N-terminal Cdk domain. We show that the Cdk domain interac6ts with S. pombe Pch1, a homolog of cyclin T, and that the purified recombinant SpCdk9/Pch1 heterodimer can phosphorylate both the pol II CTD and the C-terminal domain of S. pombe Spt5. We provide genetic evidence that SpCdk9 and Pch1 are functional orthologs of the Saccharomyces cerevisiae CTD kinase Bur1/Bur2, a putative yeast P-TEFb. Mutations of the kinase active site and the regulatory T-loop of SpCdk9 abolish its activity in vivo. Deleting the C-terminal domain of SpCdk9 causes a severe growth defect. We suggest a model whereby Spt5-induced arrest of early elongation ensures a temporal window for recruitment of the capping enzymes, which in turn attract Cdk9 to alleviate the arrest. This elongation checkpoint may avoid wasteful rounds of transcription of uncapped pre-mRNAs. |
| Keywords: | unclassified drug; dna binding protein; promoter region; gene deletion; genetics; mutation; dna-binding proteins; nonhuman; molecular genetics; protein domain; quality control; nonhistone protein; metabolism; chromosomal proteins, non-histone; cell division; complex formation; biological model; models, biological; cyclin dependent kinase 9; carboxy terminal sequence; protein binding; genetic transcription; transcription, genetic; rna; chemistry; transcription regulation; amino acid sequence; molecular sequence data; sequence homology, amino acid; hybrid protein; recombinant fusion proteins; messenger rna; saccharomyces cerevisiae; rna, messenger; recombinant proteins; recombinant protein; transcription; plasmid; plasmids; binding site; protein structure, tertiary; binding sites; molecular interaction; yeast; cycline; sequence homology; cyclin-dependent kinases; cyclins; cyclin dependent kinase; two hybrid system; biochemistry; rna polymerase ii; hydrolysis; protein tertiary structure; enzymes; rna transcription; schizosaccharomyces; schizosaccharomyces pombe proteins; schizosaccharomyces pombe protein; schizosaccharomyces pombe; transcriptional elongation factors; two-hybrid system techniques; elongation factor; cyclin-dependent kinase 9; cells; rna capping; affinity chromatography; chromatography, affinity; transcription elongation factor; baculovirus; saccharomyces; baculoviridae; messenger rna synthesis; priority journal; article; elongation factor spt5; mcs1 protein, s pombe; pch1 protein, s pombe; spt5 transcriptional elongation factor |
| Journal Title: | Journal of Biological Chemistry |
| Volume: | 278 |
| Issue: | 9 |
| ISSN: | 0021-9258 |
| Publisher: | American Society for Biochemistry and Molecular Biology |
| Date Published: | 2003-02-28 |
| Start Page: | 7180 |
| End Page: | 7188 |
| Language: | English |
| DOI: | 10.1074/jbc.M211713200 |
| PUBMED: | 12475973 |
| PROVIDER: | scopus |
| DOI/URL: | |
| Notes: | Export Date: 12 September 2014 -- Source: Scopus |
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