The sensitivity of RNA polymerase II in elongation complexes to C-terminal domain phosphatase.
The phosphorylation state of the carboxyl-terminal domain (CTD) of the largest RNA polymerase (RNAP) II subunit plays an important role in the regulation of transcript elongation. This report examines the sensitivity of RNAP II to dephosphorylation by CTD phosphatase (CTDP) and addresses factors that regulate its sensitivity. The CTDP sensitivity of RNAP IIO in paused elongation complexes on a dC-tailed template does not significantly differ from that of free RNAP IIO. RNAP IIO contained in elongation complexes that initiate transcription from the adenovirus-2 major late promoter in the presence of a nuclear extract is relatively resistant to dephosphorylation. Complexes treated with 1% Sarkosyl remain elongation-competent but demonstrate a 5-fold increase in CTDP sensitivity. Furthermore, the sensitivity of RNAP IIO in both control and Sarkosyl-treated elongation complexes is dependent on their position relative to the start site of transcription. Elongation complexes 11-24 nucleotides downstream are more sensitive to dephosphorylation than complexes 50-150 nucleotides downstream. The incubation of Sarkosyl-treated elongation complexes with nuclear extract restores the original resistance to dephosphorylation. These results suggest that a conformational change occurs in RNAP II as it clears the promoter, which results in an increased resistance to dephosphorylation. Furthermore, the sensitivity to dephosphorylation can be modulated by a factor(s) present in the nuclear extract.
Lehman
A L
AL
Section of Molecular and Cellular Biology, Division of Biological Sciences, University of California, Davis, California 95616, USA.
Dahmus
M E
ME
D000260:Adenoviruses, Human / Q000235:genetics*; D000818:Animals; D047390:Casein Kinase II; D002417:Cattle; D002467:Cell Nucleus / Q000378:metabolism; D003902:Detergents / Q000494:pharmacology; D007700:Kinetics; D046911:Macromolecular Substances; D010749:Phosphoprotein Phosphatases / Q000378:metabolism*; D010766:Phosphorylation; D011401:Promoter Regions, Genetic / Q000187:drug effects; D017346:Protein Serine-Threonine Kinases / Q000378:metabolism; D012319:RNA Polymerase II / Q000737:chemistry* / Q000378:metabolism*; D012521:Sarcosine / Q000031:analogs & derivatives / Q000494:pharmacology; D013698:Templates, Genetic; D013950:Thymus Gland / Q000201:enzymology; D014158:Transcription, Genetic / Q000187:drug effects
D003902:Detergents; D046911:Macromolecular Substances; C025231:sarkosyl; D047390:Casein Kinase II; D017346:Protein Serine-Threonine Kinases; D012319:RNA Polymerase II; D010749:Phosphoprotein Phosphatases; C116022:carboxy-terminal domain phosphatase; D012521:Sarcosine