Mutual targeting of mediator and the TFIIH kinase Kin28 Journal Article
| Authors: | Guidi, B. W.; Bjornsdottir, G.; Hopkins, D. C.; Lacomis, L.; Erdjument-Bromage, H.; Tempst, P.; Myers, L. C. |
| Article Title: | Mutual targeting of mediator and the TFIIH kinase Kin28 |
| Abstract: | In Saccharomyces cerevisiae, Kin28 is a member of the cyclin-dependent kinase family. Kin28 is a subunit of the basal transcription factor holo-TFIIH and its trimeric sub-complex TFIIK. Kin28 is the primary kinase that phosphorylates the RNA polymerase II (RNA pol II) C-terminal domain (CTD) within a transcription initiation complex. Mediator, a global transcriptional co-activator, dramatically enhances the phosphorylation of the CTD of RNA pol II by holo-TFIIH in vitro. Using purified proteins we have determined that the subunits of TFIIK are sufficient for Mediator to enhance Kin28 CTD kinase activity and that Mediator enhances phosphorylation of a glutathione S-transferase-CTD fusion protein, despite the absence of multiple Mediator and/or TFIIH interactions with polymerase. Mediator does not stimulate the activity of several other CTD kinases, suggesting that the specific enhancement of TFIIH kinase activity results in Kin28 being the primary CTD kinase at initiation. In addition, we have found that Kin28 phosphorylates Mediator subunit Med4 in an assay, including purified holo-TFIIH, and either Mediator or recombinant Med4 alone. Furthermore, Kin28 appears to be, at least in part, responsible for the phosphorylation of Med4 in vivo. We have identified Thr-237 as the site of phosphorylation of Med4 by Kin28 in vitro. The mutation of Thr-237 to Ala has no effect on the growth of a yeast strain under normal conditions but confirms that Thr-237 is also the site of Med4 phosphorylation in vivo. |
| Keywords: | controlled study; protein phosphorylation; unclassified drug; nonhuman; protein domain; complex formation; protein kinases; carboxy terminal sequence; protein targeting; transcription initiation; transcription factor; transcription, genetic; in vitro study; phosphorylation; rna; enzyme phosphorylation; transcription regulation; hybrid protein; recombinant fusion proteins; protein purification; saccharomyces cerevisiae; bioassay; recombinant protein; glutathione transferase; threonine; trans-activators; yeast; saccharomyces cerevisiae proteins; protein subunit; protein subunits; cyclin-dependent kinases; cyclin dependent kinase; enzyme subunit; biochemistry; kinase activity; rna polymerase ii; fungi; enzymes; fungus growth; gene expression regulation, fungal; transcription initiation site; transcription factor tfiih; saccharomyces; mediator release; transcription factors, tfii; priority journal; article; phosphorylates; protein kin28; protein med4; transcription factor 2h kinase; transcription factor 2k; casein kinases |
| Journal Title: | Journal of Biological Chemistry |
| Volume: | 279 |
| Issue: | 28 |
| ISSN: | 0021-9258 |
| Publisher: | American Society for Biochemistry and Molecular Biology |
| Date Published: | 2004-07-09 |
| Start Page: | 29114 |
| End Page: | 29120 |
| Language: | English |
| DOI: | 10.1074/jbc.M404426200 |
| PROVIDER: | scopus |
| PUBMED: | 15126497 |
| DOI/URL: | |
| Notes: | J. Biol. Chem. -- Cited By (since 1996):24 -- Export Date: 16 June 2014 -- CODEN: JBCHA -- Source: Scopus |
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