Characterization of the residues phosphorylated in vitro by different C-terminal domain kinases.
The C-terminal part of the largest subunit of eukaryotic RNA polymerase II is composed solely of the highly repeated consensus sequence Tyr1-Ser2-Pro3-Thr4-Ser5-Pro6-Ser7. This domain, called the C-terminal domain (CTD), is phosphorylated mostly at serine residues during transcription initiation, but the precise role of this phosphorylation remains controversial. Several protein kinases are able to phosphorylate this sequence in vitro. The aim of this work was to define the positions of the amino acids phosphorylated by four of these CTD kinases (transcription factor (TF) IIH-kinase, DNA-dependent protein kinase, and the mitogen-activated protein kinases ERK1 and ERK2) and to compare the specificity of these different protein kinases. We show that TFIIH kinase and the mitogen-activated protein kinases phosphorylate only serine 5 of the CTD sequence, whereas DNA-dependent protein kinase phosphorylates serines 2 and 7. Among the different CTD kinases, only TFIIH kinase is appreciably more active on two repeats of the consensus sequence than on one motif. These in vitro results can provide some clues to the nature of the protein kinases responsible for the in vivo phosphorylation of the RNA polymerase CTD. In particular, the ratio of phosphorylated serine to threonine observed in vivo cannot be explained if TFIIH kinase is the only protein kinase involved in the phosphorylation of the CTD.
Trigon
S
S
Ecole Normale Superieure, Unité de Génétique Moléculaire, 46, rue d'Ulm, 75230 Paris Cedex 05, France.
Serizawa
H
H
Conaway
J W
JW
Conaway
R C
RC
Jackson
S P
SP
Morange
M
M
D000595:Amino Acid Sequence; D016384:Consensus Sequence; D004790:Enzyme Induction; D018869:Heat-Shock Response; D008969:Molecular Sequence Data; D018384:Oxidative Stress; D010766:Phosphorylation; D011494:Protein Kinases / Q000096:biosynthesis / Q000737:chemistry / Q000378:metabolism*; D012694:Serine / Q000378:metabolism; D013379:Substrate Specificity
D012694:Serine; D011494:Protein Kinases; C060137:carboxy-terminal domain kinase