View the Current Issue

Article

  • The EMBO Journal (2001) 20, 3749 - 3759
  • doi:10.1093/emboj/20.14.3749

T-loop phosphorylation stabilizes the CDK7–cyclin H–MAT1 complex in vivo and regulates its CTD kinase activity

Stéphane Larochelle1,2, Jian Chen2, Ronald Knights1, Judit Pandur2, Patrick Morcillo1, Hediye Erdjument-Bromage3, Paul Tempst3, Beat Suter2 and Robert P. Fisher1

  1. Cell Biology Program, Memorial Sloan-Kettering Cancer Center, 1275 York Avenue, New York, NY 10021, USA
  2. Department of Biology, McGill University, 1205 Dr Penfield Avenue, Montreal, PQ, Canada H3A 1B1
  3. Molecular Biology Program, Memorial Sloan-Kettering Cancer Center, 1275 York Avenue, New York, NY 10021, USA

Correspondence to:

Robert P. Fisher, E-mail: r-fisher@ski.mskcc.org

Received 1 December 2000; Accepted 25 May 2001; Revised 30 April 2001

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.

  • Keywords:

    • CAK,
    • cell cycle,
    • cyclin-dependent kinase,
    • Drosophila,
    • TFIIH