A role for SKIP in EBNA2 activation of CBF1-repressed promoters.
EBNA2 is essential for Epstein-Barr virus (EBV) immortalization of B lymphocytes. EBNA2 functions as a transcriptional activator and targets responsive promoters through interaction with the cellular DNA binding protein CBF1. We have examined the mechanism whereby EBNA2 overcomes CBF1-mediated transcriptional repression. A yeast two-hybrid screen performed using CBF1 as the bait identified a protein, SKIP, which had not previously been recognized as a CBF1-associated protein. Protein-protein interaction assays demonstrated contacts between SKIP and the SMRT, CIR, Sin3A, and HDAC2 proteins of the CBF1 corepressor complex. Interestingly, EBNA2 also interacted with SKIP in glutathione S-transferase affinity and mammalian two-hybrid assays and colocalized with SKIP in immunofluorescence assays. Interaction with SKIP was not affected by mutation of EBNA2 conserved region 6, the CBF1 interaction region, but was abolished by mutation of conserved region 5. Mutation of conserved region 5 also severely impaired EBNA2 activation of a reporter containing CBF1 binding sites. Thus, interaction with both CBF1 and SKIP is necessary for efficient promoter activation by EBNA2. A model is presented in which EBNA2 competes with the SMRT-corepressor complex for contacts on SKIP and CBF1.
Zhou
S
S
Department of Pharmacology and Molecular Sciences, Johns Hopkins School of Medicine, Baltimore, Maryland 21205, USA.
Fujimuro
M
M
Hsieh
J J
JJ
Chen
L
L
Hayward
S D
SD
D000595:Amino Acid Sequence; D001665:Binding Sites; D002461:Cell Line, Transformed; D004268:DNA-Binding Proteins / Q000235:genetics / Q000378:metabolism*; D019309:Epstein-Barr Virus Nuclear Antigens; D005786:Gene Expression Regulation; D006367:HeLa Cells; D056464:Histone Deacetylase 2; D006655:Histone Deacetylases / Q000378:metabolism; D006801:Humans; D051819:Immunoglobulin J Recombination Signal Sequence-Binding Protein; D008969:Molecular Sequence Data; D009687:Nuclear Proteins / Q000235:genetics / Q000378:metabolism / Q000502:physiology*; D056985:Nuclear Receptor Co-Repressor 2; D056918:Nuclear Receptor Coactivators; D011401:Promoter Regions, Genetic; D011993:Recombinant Fusion Proteins / Q000235:genetics / Q000378:metabolism; D012097:Repressor Proteins / Q000378:metabolism; D056264:Sin3 Histone Deacetylase and Corepressor Complex; D014157:Transcription Factors; D014162:Transfection; D014764:Viral Proteins / Q000235:genetics / Q000378:metabolism*
C117385:CIR1 protein, human; D004268:DNA-Binding Proteins; C402641:EBNA-2 protein, Human herpesvirus 4; D019309:Epstein-Barr Virus Nuclear Antigens; D051819:Immunoglobulin J Recombination Signal Sequence-Binding Protein; C539130:NCOR2 protein, human; D009687:Nuclear Proteins; D056985:Nuclear Receptor Co-Repressor 2; D056918:Nuclear Receptor Coactivators; C497570:RBPJ protein, human; D011993:Recombinant Fusion Proteins; D012097:Repressor Proteins; C099683:SIN3A transcription factor; C111791:SNW1 protein, human; D014157:Transcription Factors; D014764:Viral Proteins; D056464:Histone Deacetylase 2; D006655:Histone Deacetylases; D056264:Sin3 Histone Deacetylase and Corepressor Complex