Evidence of the Involvement of O-GlcNAc-modified Human RNA Polymerase II CTD in Transcription in Vitro and in Vivo*
- From the ‡Metabolism Branch, Center for Cancer Research, NCI, National Institutes of Health, Bethesda, Maryland 20892,
- the §Laboratory of Cell Biochemistry and Biology, NIDDK, National Institutes of Health, Bethesda, Maryland 20892, and
- the ¶Department of Biological Chemistry, Johns Hopkins University School of Medicine, Baltimore, Maryland 21205
- ↵2 To whom correspondence should be addressed: Transcriptional Regulation and Biochemistry Unit, Metabolism Branch, Ctr. for Cancer Research, NCI, National Institutes of Health, Bldg. 10, Rm. 6B05, 9000 Rockville Pike, Bethesda, MD 20892. Tel.: 301-435-8323; E-mail: lewisbri{at}mail.nih.gov.
Abstract
The RNA polymerase II C-terminal domain (CTD), which serves as a scaffold to recruit machinery involved in transcription, is modified post-translationally. Although the O-GlcNAc modification of RNA polymerase II CTD was documented in 1993, its functional significance remained obscure. We show that O-GlcNAc transferase (OGT) modified CTD serine residues 5 and 7. Drug inhibition of OGT and OGA (N-acetylglucosaminidase) blocked transcription during preinitiation complex assembly. Polymerase II and OGT co-immunoprecipitated, and OGT is a component of the preinitiation complex. OGT shRNA experiments showed that reduction of OGT causes a reduction in transcription and RNA polymerase II occupancy at several B-cell promoters. These data suggest that the cycling of O-GlcNAc on and off of polymerase II occurs during assembly of the preinitiation complex. Our results define unexpected roles for both the CTD and O-GlcNAc in the regulation of transcription initiation in higher eukaryotes.
Footnotes
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↵1 Recipient of royalties from the sale of the CTD110.6 O-GlcNAc antibody under a licensing agreement among The Johns Hopkins University, Covance Research Products, Sigma-Aldrich, and Santa Cruz Biotechnology. The terms of this agreement are managed by The Johns Hopkins University.
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↵* This work was supported, in whole or in part, by National Institutes of Health Intramural Research Program grants from NCI, Center for Cancer Research (to B. A. L.) and NIDDK (to J. A. H.) and by National Institutes of Health grants NIH R01-CA42486 and R01-DK61671 (to G. W. H.).
- Received December 5, 2011.
- Revision received May 10, 2012.
- © 2012 by The American Society for Biochemistry and Molecular Biology, Inc.
