Dephosphorylation of the Carboxyl-Terminal Domain of RNA Polymerase II

Abstract

This chapter reviews the carboxyl-terminal domain (CTD) of the largest RNA polymerase (RNAP) II subunit which is composed of tandem repeats of the consensus sequence Tyr 1 Ser 2 Pro 3 Thr 4 Ser 5 Pro 6 Ser 7 . Reversible phosphorylation of the CTD, especially at Ser positions 2 and 5, plays an important role in the regulation of gene expression. RNAP IIA, which contains an unmodified CTD, and RNAP IIO, which contains a hyperphosphorylated CTD, has distinct functions in the transcription cycle. RNAP IIA is recruited actively to the promoter as part of the preinitiation complex, whereas RNAP IIO is responsible for transcript elongation. It discusses the phosphorylated CTD that mediates promoter escape, and the recruitment of factors necessary for processing of pre-mRNA. CTD kinases and phosphatases modulate the state of CTD phosphorylation, and accordingly play a critical role in regulating the synthesis and processing of the primary transcript. The chapter describes procedures for various assays involving CTD dephosphorylation. These procedures provide the tools for the identification and characterization of enzymes that dephosphorylate the CTD of RNAP II.