Growth-related Changes in Phosphorylation of Yeast RNA Polymerase II*
- Meera Patturajan‡,
- Roberta J. Schulte§,
- Bartholomew M. Sefton§,
- Ronald Berezney¶,
- Michel Vincent‖,
- Olivier Bensaude**,
- Stephen L. Warren‡ and
- Jeffry L. Cordenत
- From the ‡Department of Molecular Biology and Genetics, The Johns Hopkins University School of Medicine, Baltimore, Maryland 21205, The §Salk Institute for Biological Studies, San Diego, California 92186,‡Nexstar Pharmaceuticals, Boulder, Colorado 80301, the ¶Department of Biological Sciences, State University of New York, Buffalo, New York 14260, the‖Recherche en Sciences de la Vie et de la Sante and Department de Medicine, Universite Laval, Ste-Foy, Quebec, Canada G1K 7P4;**Genetique Moleculaire, Ecole Normale Superieure, 75230 Paris Cedex 05, France
Abstract
The largest subunit of RNA polymerase II contains a unique C-terminal domain (CTD) consisting of tandem repeats of the consensus heptapeptide sequence Tyr1-Ser2-Pro3-Thr4-Ser5-Pro6-Ser7. Two forms of the largest subunit can be separated by SDS-polyacrylamide gel electrophoresis. The faster migrating form termed IIA contains little or no phosphate on the CTD, whereas the slower migrating II0 form is multiply phosphorylated. CTD kinases with different phosphoryl acceptor specificities are able to convert IIA to II0 in vitro, and different phosphoisomers have been identified in vivo. In this paper we report the binding specificities of a set of monoclonal antibodies that recognize different phosphoepitopes on the CTD. Monoclonal antibodies like H5 recognize phosphoserine in position 2, whereas monoclonal antibodies like H14 recognize phosphoserine in position 5. The relative abundance of these phosphoepitopes changes when growing yeast enter stationary phase or are heat-shocked. These results indicate that phosphorylation of different CTD phosphoacceptor sites are independently regulated in response to environmental signals.
Footnotes
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↵* This work was supported by Grant 82371 from the American Cancer Society (to J. L. C.).The costs of publication of this article were defrayed in part by the payment of page charges. The article must therefore be hereby marked “advertisement” in accordance with 18 U.S.C. Section 1734 solely to indicate this fact.
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↵§§ To whom reprint requests should be addressed: Dept. of Molecular Biology and Genetics, The Johns Hopkins University School of Medicine, 725 N. Wolfe St., Baltimore, MD 21205-2185. Tel.: 410-955-4719; Fax: 410-502-6718; E-mail: jeff_corden{at}qmail.bs.jhu.edu.
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↵1 The abbreviations used are: pol, polymerase; CTD, C-terminal domain; TF, transcription factor; mAb, monoclonal antibody; GST, glutathione S-transferase; PAGE, polyacrylamide gel electrophoresis; WT, wild type.
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↵2 R. J. Schulte and B. M. Sefton, submitted for publication.
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↵3 M. Patturajan and J. L. Corden, unpublished observations.
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- Received October 17, 1997.
- Revision received November 21, 1997.
- The American Society for Biochemistry and Molecular Biology, Inc.
