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Molecular and Cellular Biology, September 2009, p. 4852-4863, Vol. 29, No. 17
0270-7306/09/$08.00+0     doi:10.1128/MCB.00609-09
Copyright © 2009, American Society for Microbiology. All Rights Reserved.

Phosphorylation of the Transcription Elongation Factor Spt5 by Yeast Bur1 Kinase Stimulates Recruitment of the PAF Complex{triangledown} ,{dagger}

Ying Liu,1,{ddagger} Linda Warfield,1 Chao Zhang,2 Jie Luo,3 Jasmina Allen,2 Walter H. Lang,4 Jeffrey Ranish,3 Kevan M. Shokat,2 and Steven Hahn1*

Fred Hutchinson Cancer Research Center, Division of Basic Sciences, 1100 Fairview Ave N., MS A1-162, Seattle, Washington 98109,1 University of California—San Francisco, Department of Cellular and Molecular Pharmacology, 600 16th Street, MC 2280, San Francisco, California 94158-2280,2 Institute for Systems Biology, 1441 North 34th Street, Seattle, Washington 98103-8904,3 Life Sciences Division, Lawrence Berkeley National Laboratory, 1 Cyclotron Road, MS 83R0101, Berkeley, California 947204

Received 9 May 2009/ Returned for modification 10 June 2009/ Accepted 26 June 2009

The Saccharomyces cerevisiae kinase Bur1 is involved in coupling transcription elongation to chromatin modification, but not all important Bur1 targets in the elongation complex are known. Using a chemical genetics strategy wherein Bur1 kinase was engineered to be regulated by a specific inhibitor, we found that Bur1 phosphorylates the Spt5 C-terminal repeat domain (CTD) both in vivo and in isolated elongation complexes in vitro. Deletion of the Spt5 CTD or mutation of the Spt5 serines targeted by Bur1 reduces recruitment of the PAF complex, which functions to recruit factors involved in chromatin modification and mRNA maturation to elongating polymerase II (Pol II). Deletion of the Spt5 CTD showed the same defect in PAF recruitment as rapid inhibition of Bur1 kinase activity, and this Spt5 mutation led to a decrease in histone H3K4 trimethylation. Brief inhibition of Bur1 kinase activity in vivo also led to a significant decrease in phosphorylation of the Pol II CTD at Ser-2, showing that Bur1 also contributes to Pol II Ser-2 phosphorylation. Genetic results suggest that Bur1 is essential for growth because it targets multiple factors that play distinct roles in transcription.

* Corresponding author. Mailing address: Fred Hutchinson Cancer Research Center, Division of Basic Sciences, 1100 Fairview Ave. N., MS A1-162, Seattle, WA 98109. Phone: (206) 667 5261. Fax: (206) 667 6497. E-mail: shahn{at}fhcrc.org

{triangledown} Published ahead of print on 6 July 2009.

{dagger} Supplemental material for this article may be found at http://mcb.asm.org/.

{ddagger} Present address: Center for Computational and Integrative Biology, Massachusetts General Hospital, 185 Cambridge St., Boston, MA 02114.

Molecular and Cellular Biology, September 2009, p. 4852-4863, Vol. 29, No. 17
0270-7306/09/$08.00+0     doi:10.1128/MCB.00609-09
Copyright © 2009, American Society for Microbiology. All Rights Reserved.





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