Chemical inhibition of the TFIIH-associated kinase Cdk7/Kin28 does not impair global mRNA synthesis

*Department of Biochemistry and
^‖Genome Center of Wisconsin, University of Wisconsin, Madison, WI 53706;
^†Department of Cellular and Molecular Pharmacology, University of California, San Francisco, CA 94143;
^‡School of Pharmacy, University of Wisconsin, Madison, WI 53705;
^§Memorial Sloan–Kettering Cancer Center, New York, NY 10021; and
^¶Fred Hutchinson Cancer Research Center, Seattle, WA 98109

Communicated by Mark Ptashne, Memorial Sloan–Kettering Cancer Center, New York, NY, December 26, 2006 (received for review November 27, 2006)

Abstract

The process of gene transcription requires the recruitment of a hypophosphorylated form of RNA polymerase II (Pol II) to a gene promoter. The TFIIH-associated kinase Cdk7/Kin28 hyperphosphorylates the promoter-bound polymerase; this event is thought to play a crucial role in transcription initiation and promoter clearance. Studies using temperature-sensitive mutants of Kin28 have provided the most compelling evidence for an essential role of its kinase activity in global mRNA synthesis. In contrast, using a small molecule inhibitor that specifically inhibits Kin28 in vivo, we find that the kinase activity is not essential for global transcription. Unlike the temperature-sensitive alleles, the small-molecule inhibitor does not perturb protein–protein interactions nor does it provoke the disassociation of TFIIH from gene promoters. These results lead us to conclude that other functions of TFIIH, rather than the kinase activity, are critical for global gene transcription.

Footnotes

**To whom correspondence should be addressed at:
Department of Biochemistry and The Genome Center of Wisconsin, University of Wisconsin, 433 Babcock Drive, Madison, WI 53706.
E-mail: ansari{at}biochem.wisc.edu
Author contributions: E.I.K. and A.Z.A. designed research; E.I.K., R.T.K., and A.V. performed research; E.I.K., C.K., S.H., and K.M.S. contributed new reagents/analytic tools; E.I.K., M.S., S.H., K.M.S., and A.Z.A. analyzed data; and E.I.K. and A.Z.A. wrote the paper.
The authors declare no conflict of interest.
This article contains supporting information online at www.pnas.org/cgi/content/full/0611505104/DC1.
Abbreviations:

Pol II,

RNA polymerase II;

CTD,

C-terminal domain of the largest subunit of Pol II;

PIC,

preinitiation complex;

Cdk7/Kin28,

yeast cyclin-dependent kinase;

Kin28as,

analog-sensitive allele of Kin28;

Kin28ts,

temperature-sensitive allele(s) of Kin28.