Fcp1 directly recognizes the C-terminal domain (CTD) and interacts with a site on RNA polymerase II distinct from the CTD

doi:10.1073/pnas.0507987102

Fcp1 directly recognizes the C-terminal domain (CTD) and interacts with a site on RNA polymerase II distinct from the CTD

^*Department of Molecular Biology and Genetics, Cornell University, Biotechnology Building, Ithaca, NY 14853; ^‡Molecular Biology Program, Sloan–Kettering Institute, New York, NY 10021; and ^§Department of Pharmacology and Experimental Therapeutics, University of Maryland School of Medicine, Baltimore, MD 21021

Communicated by Roger D. Kornberg, Stanford University School of Medicine, Stanford, CA, October 9, 2005 (received for review June 28, 2005)

Abstract

Fcp1 is an essential protein phosphatase that hydrolyzes phosphoserines within the C-terminal domain (CTD) of the largest subunit of RNA polymerase II (Pol II). Fcp1 plays a major role in the regulation of CTD phosphorylation and, hence, critically influences the function of Pol II throughout the transcription cycle. The basic understanding of Fcp1–CTD interaction has remained ambiguous because two different modes have been proposed: the “dockingsite” model versus the “distributive” mechanism. Here we demonstrate biochemically that Fcp1 recognizes and dephosphorylates the CTD directly, independent of the globular non-CTD part of the Pol II structure. We point out that the recognition of CTD by the phosphatase is based on random access and is not driven by Pol II conformation. Results from three different types of experiments reveal that the overall interaction between Fcp1 and Pol II is not stable but dynamic. In addition, we show that Fcp1 also interacts with a region on the polymerase distinct from the CTD. We emphasize that this non-CTD site is functionally distinct from the docking site invoked previously as essential for the CTD phosphatase activity of Fcp1. We speculate that Fcp1 interaction with the non-CTD site may mediate its stimulatory effect on transcription elongation reported previously.

Footnotes

↵ ¶ To whom correspondence should be addressed at: Department of Molecular Biology and Genetics, 221 Biotechnology Building, Campus Road, Cornell University, Ithaca, NY 14853. E-mail: jf227{at}cornell.edu.
↵ † M.-H.S. and P.Y. contributed equally to this work.
Conflict of interest statement: No conflicts declared.
Abbreviations: CTD, C-terminal domain; CTDo, hyperphosphorylated CTD; CTDa, hypophosphorylated CTD; Pol II, RNA polymerase II; BRCT, breast cancer 1 C terminus; SEC, size-exclusion chromatography.