Walleye dermal sarcoma virus cyclin interacts with components of the mediator complex and the RNA polymerase II holoenzyme.
Walleye dermal sarcoma virus (WDSV) encodes an accessory protein, OrfA, with sequence homology to cyclins (retrovirus cyclin). In cells transfected with an expression construct, OrfA was localized to the nucleus and was concentrated in interchromatin granule clusters (IGCs), sites where splicing factors are concentrated. Other proteins identified in IGCs include transcription factors, the large subunit of RNA polymerase II (Pol II), and cyclin-dependent kinase 8 (cdk8). cdk8 is the kinase partner of cyclin C and a component of the mediator complex, associated with the Pol II holoenzyme. cdk8 and cyclin C can regulate transcription via phosphorylation of cyclin H and the carboxy-terminal domain of Pol II. OrfA in transfected HeLa cells was found to colocalize and copurify with hyperphosphorylated forms of Pol II (Pol IIO) in IGCs, and OrfA was coimmunoprecipitated from lysates of transfected cells with an antibody against Pol IIO. Likewise, Pol IIO could be coprecipitated with an antibody against OrfA. A survey with antibodies against several different cdks resulted in coimmunoprecipitation of OrfA with anti-cdk8, and antiserum against OrfA was able to coprecipitate cdk8 from lysates of cells that express OrfA. Coprecipitation of OrfA with anti-cyclin C demonstrated that it was included in complexes with OrfA and cdk8. OrfA has sequence and structural similarities to cyclin C, and, functionally, OrfA appears to have the capacity to both enhance and inhibit the activity of promoters in a cell-specific manner, similar to functions of the mediator complex. These data suggest that WDSV OrfA functions through its interactions with these large, transcription complexes. Further investigations will clarify the role of the retrovirus cyclin in control of virus expression and transformation.
Department of Molecular Biosciences, The University of Kansas, Lawrence 66045, USA.
Joel
J
Rovnak
Sandra L
SL
Quackenbush
D016475:3T3 Cells; D000818:Animals; D056850:Cyclin-Dependent Kinase 8; D018844:Cyclin-Dependent Kinases; D016213:Cyclins / Q000737:chemistry / Q000235:genetics* / Q000302:isolation & purification / Q000378:metabolism*; D030283:Epsilonretrovirus / Q000235:genetics* / Q000378:metabolism*; D005814:Genes, Viral; D006367:HeLa Cells; D020035:Holoenzymes / Q000378:metabolism; D006801:Humans; D051379:Mice; D009154:Mutation; D010766:Phosphorylation; D011233:Precipitin Tests; D011401:Promoter Regions, Genetic; D017346:Protein Serine-Threonine Kinases / Q000378:metabolism; D017433:Protein Structure, Secondary; D012319:RNA Polymerase II / Q000302:isolation & purification / Q000378:metabolism*; D012326:RNA Splicing; D011994:Recombinant Proteins / Q000235:genetics / Q000378:metabolism; D014157:Transcription Factors; D014162:Transfection
D016213:Cyclins; D020035:Holoenzymes; D011994:Recombinant Proteins; D014157:Transcription Factors; D017346:Protein Serine-Threonine Kinases; C095686:CDK8 protein, human; D056850:Cyclin-Dependent Kinase 8; D018844:Cyclin-Dependent Kinases; D012319:RNA Polymerase II