Abstract
In humans, 5' m(7)G cap addition is accomplished cotranscriptionally by the sequential action of the capping enzyme (Hce1) and the cap methyltransferase (Hcm1). We found that guanylylation and methylation occur efficiently during transcription with t(1/2)'s of less than 15 and 70 s, respectively. A two to four order of magnitude increase was found in the rate of guanylylation of RNA in transcription complexes compared to free RNA. This stimulation required only the RNA polymerase II elongation complex and Hce1. Capping activity was weakly associated with elongation but not preinitiation complexes. The CTD was not required for functional coupling but stimulated the rate of capping 4-fold. Inhibition of Cdk7 but not Cdk9 similarly slowed the rate of capping.
Publication types
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Research Support, U.S. Gov't, P.H.S.
MeSH terms
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Antibodies / metabolism
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Cell Fractionation
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Guanosine / analogs & derivatives*
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Guanosine / immunology
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Guanosine / metabolism
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HeLa Cells
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Humans
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Macromolecular Substances
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Methyltransferases / genetics
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Methyltransferases / metabolism*
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Nuclear Proteins / genetics
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Nuclear Proteins / metabolism
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Nucleotidyltransferases / genetics
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Nucleotidyltransferases / metabolism*
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Protein Binding
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RNA Caps / metabolism*
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RNA Polymerase II / metabolism
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RNA Processing, Post-Transcriptional*
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RNA, Messenger / genetics
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RNA, Messenger / metabolism*
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Recombinant Fusion Proteins / genetics
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Recombinant Fusion Proteins / metabolism
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Time Factors
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Transcription, Genetic*
Substances
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Antibodies
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Macromolecular Substances
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Nuclear Proteins
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RNA Caps
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RNA, Messenger
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Recombinant Fusion Proteins
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Guanosine
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N(2),N(2),7-trimethylguanosine
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Methyltransferases
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mRNA (guanine(N7))-methyltransferase
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Nucleotidyltransferases
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RNA Polymerase II
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mRNA guanylyltransferase