Modulation of RNA polymerase II elongation efficiency by C-terminal heptapeptide repeat domain kinase I.
Hyperphosphorylation of the C-terminal heptapeptide repeat domain (CTD) of the RNA polymerase II largest subunit has been suggested to play a key role in regulating transcription initiation and elongation. To facilitate investigating functional consequences of CTD phosphorylation we developed new templates, the double G-less cassettes, which make it possible to assay simultaneously the level of initiation and the efficiency of elongation. Using these templates, we examined the effects of yeast CTD kinase I or CTD kinase inhibitors on transcription and CTD phosphorylation in HeLa nuclear extracts. Our results showed that polymerase II elongation efficiency and CTD phosphorylation are greatly reduced by CTD kinase inhibitors, whereas both are greatly increased by CTD kinase I; in contrast, transcription initiation is much less affected. These results demonstrate that CTD kinase I modulates the elongation efficiency of RNA polymerase II and are consistent with the idea that one function of CTD phosphorylation is to promote effective production of long transcripts by stimulating the elongation efficiency of RNA polymerase II.
Lee
J M
JM
Department of Biochemistry, Duke University Medical Center, Durham, North Carolina 27710, USA.
Greenleaf
A L
AL
D002467:Cell Nucleus / Q000378:metabolism*; D004004:Dichlororibofuranosylbenzimidazole / Q000494:pharmacology; D006367:HeLa Cells; D006801:Humans; D007546:Isoquinolines / Q000494:pharmacology; D019384:Nucleic Acid Synthesis Inhibitors / Q000494:pharmacology; D011494:Protein Kinases / Q000378:metabolism*; D012319:RNA Polymerase II / Q000378:metabolism*; D013347:Subcellular Fractions / Q000378:metabolism; D014158:Transcription, Genetic / Q000187:drug effects
D007546:Isoquinolines; D019384:Nucleic Acid Synthesis Inhibitors; D004004:Dichlororibofuranosylbenzimidazole; C053177:N-(2-(methylamino)ethyl)-5-isoquinolinesulfonamide; D011494:Protein Kinases; C060137:carboxy-terminal domain kinase; D012319:RNA Polymerase II