Article

The human CDK8 subcomplex is a histone kinase that requires Med12 for activity and can function independently of mediator.

Department of Chemistry and Biochemistry, University of Colorado, Boulder, Colorado 80309, USA.
Molecular and cellular biology (impact factor: 6.06). 01/2009; 29(3):650-61. DOI:10.1128/MCB.00993-08 pp.650-61
Source: PubMed

ABSTRACT The four proteins CDK8, cyclin C, Med12, and Med13 can associate with Mediator and are presumed to form a stable "CDK8 subcomplex" in cells. We describe here the isolation and enzymatic activity of the 600-kDa CDK8 subcomplex purified directly from human cells and also via recombinant expression in insect cells. Biochemical analysis of the recombinant CDK8 subcomplex identifies predicted (TFIIH and RNA polymerase II C-terminal domain [Pol II CTD]) and novel (histone H3, Med13, and CDK8 itself) substrates for the CDK8 kinase. Notably, these novel substrates appear to be metazoan-specific. Such diverse targets imply strict regulation of CDK8 kinase activity. Along these lines, we observe that Mediator itself enables CDK8 kinase activity on chromatin, and we identify Med12--but not Med13--to be essential for activating the CDK8 kinase. Moreover, mass spectrometry analysis of the endogenous CDK8 subcomplex reveals several associated factors, including GCN1L1 and the TRiC chaperonin, that may help control its biological function. In support of this, electron microscopy analysis suggests TRiC sequesters the CDK8 subcomplex and kinase assays reveal the endogenous CDK8 subcomplex--unlike the recombinant submodule--is unable to phosphorylate the Pol II CTD.

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  • Article: TFIIH is negatively regulated by cdk8-containing mediator complexes.
    S Akoulitchev, S Chuikov, D Reinberg
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    ABSTRACT: The mammalian cyclin-dependent kinase 8 (cdk8) gene has been linked with a subset of acute lymphoblastic leukaemias, and its corresponding protein has been functionally implicated in regulation of transcription. Mammalian cdk8 and cyclin C, and their respective yeast homologues, Srb10 and Srb11, are components of the RNA polymerase II holoenzyme complex where they function as a protein kinase that phosphorylates the carboxy-terminal domain (CTD) of the largest subunit of RNA polymerase II (ref. 7). The yeast SRB10 and SRB11 genes have been implicated in the negative regulation of transcription. The cdk8/cyclin C protein complex is also found in a number of mammalian Mediator-like protein complexes, which repress activated transcription independently of the CTD in vitro. Here we show that cdk8/cyclin C can regulate transcription by targeting the cdk7/cyclin H subunits of the general transcription initiation factor IIH (TFIIH). cdk8 phosphorylates mammalian cyclin H in the vicinity of its functionally unique amino-terminal and carboxy-terminal alpha-helical domains. This phosphorylation represses both the ability of TFIIH to activate transcription and its CTD kinase activity. In addition, mimicking cdk8 phosphorylation of cyclin H in vivo has a dominant-negative effect on cell growth. Our results link the Mediator complex and the basal transcription machinery by a regulatory pathway involving two cyclin-dependent kinases. This pathway appears to be unique to higher organisms.
    Nature 10/2000; 407(6800):102-6. · 36.28 Impact Factor
Data provided are for informational purposes only. Although carefully collected, accuracy cannot be guaranteed. The impact factor represents a rough estimation of the journal's impact factor and does not reflect the actual current impact factor. Publisher conditions are provided by RoMEO. Differing provisions from the publisher's actual policy or licence agreement may be applicable.

Keywords

600-kDa CDK8 subcomplex purified
 
associated factors
 
Biochemical analysis
 
CDK8
 
CDK8 kinase
 
CDK8 kinase activity
 
CDK8 subcomplex
 
cyclin C
 
diverse targets
 
electron microscopy analysis
 
endogenous CDK8 subcomplex
 
four proteins CDK8
 
human cells
 
mass spectrometry analysis
 
novel substrates
 
Pol II CTD
 
recombinant CDK8 subcomplex
 
recombinant expression
 
RNA polymerase II C-terminal domain [Pol II CTD]
 
stable