title: T-loop phosphorylation stabilizes the CDK7-cyclin H-MAT1 complex in vivo and regulates its CTD kinase activity.
abstract: Cyclin-dependent kinase (CDK)7-cyclin H, the CDK-activating kinase (CAK) and TFIIH-associated kinase in metazoans can be activated in vitro through T-loop phosphorylation or binding to the RING finger protein MAT1. Although the two mechanisms can operate independently, we show that in a physiological setting, MAT1 binding and T-loop phosphorylation cooperate to stabilize the CAK complex of Drosophila. CDK7 forms a stable complex with cyclin H and MAT1 in vivo only when phosphorylated on either one of two residues (Ser164 or Thr170) in its T-loop. Mutation of both phosphorylation sites causes temperature-dependent dissociation of CDK7 complexes and lethality. Furthermore, phosphorylation of Thr170 greatly stimulates the activity of the CDK7- cyclin H-MAT1 complex towards the C-terminal domain of RNA polymerase II without significantly affecting activity towards CDK2. Remarkably, the substrate-specific increase in activity caused by T-loop phosphorylation is due entirely to accelerated enzyme turnover. Thus phosphorylation on Thr170 could provide a mechanism to augment CTD phosphorylation by TFIIH-associated CDK7, and thereby regulate transcription.
authors.affiliation: Cell Biology Program, Memorial Sloan-Kettering Cancer Center, 1275 York Avenue, New York, NY 10021, USA.
authors.forename: S
authors.initials: S
authors.lastname: Larochelle
authors.forename: J
authors.initials: J
authors.lastname: Chen
authors.forename: R
authors.initials: R
authors.lastname: Knights
authors.forename: J
authors.initials: J
authors.lastname: Pandur
authors.forename: P
authors.initials: P
authors.lastname: Morcillo
authors.forename: H
authors.initials: H
authors.lastname: Erdjument-Bromage
authors.forename: P
authors.initials: P
authors.lastname: Tempst
authors.forename: B
authors.initials: B
authors.lastname: Suter
authors.forename: R P
authors.initials: RP
authors.lastname: Fisher
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D001704:Biopolymers; D056748:Cyclin H; D018844:Cyclin-Dependent Kinases; D016213:Cyclins / Q000037:antagonists & inhibitors / Q000378:metabolism*; D004330:Drosophila; D029721:Drosophila Proteins; D007700:Kinetics; D008969:Molecular Sequence Data; D010766:Phosphorylation; D011494:Protein Kinases / Q000378:metabolism*; D017346:Protein Serine-Threonine Kinases / Q000037:antagonists & inhibitors / Q000378:metabolism*; D013379:Substrate Specificity; D000095043:Cyclin-Dependent Kinase-Activating Kinase
chemical_list: D001704:Biopolymers; C534754:CycH protein, Drosophila; D056748:Cyclin H; D016213:Cyclins; D029721:Drosophila Proteins; D011494:Protein Kinases; C060137:carboxy-terminal domain kinase; D017346:Protein Serine-Threonine Kinases; D018844:Cyclin-Dependent Kinases; D000095043:Cyclin-Dependent Kinase-Activating Kinase