Epstein-Barr virus nuclear antigen 3C is a powerful repressor of transcription when tethered to DNA.
The expression of Epstein-Barr virus (EBV) nuclear antigen 3C (EBNA3C) is essential for the activation and immortalization of human B lymphocytes by EBV. EBNA3C consists of 992 amino acids and includes a potential bZIP motif and regions rich in acidic, proline, and glutamine residues. Thus, EBNA3C resembles several trans regulators of gene expression. It has recently been shown that a fragment of EBNA3C can activate reporter gene expression when fused to the DNA-binding domain of GAL4 (D. Marshall and C. Sample, J. Virol. 69:3624-3630,1995). Although EBNA3C binds DNA, a specific site for EBNA3C binding has not been identified; to test the ability of full-length EBNA3C to regulate transcription, EBNA3C (amino acids 11 to 992) was fused to the DNA-binding domain of GAL4. We show that this fusion protein does not transactivate but rather is a potent repressor of reporter gene expression. Repression is dependent on the dose of GAL4-EBNA3C and on the presence of GAL4-binding sites within reporter plasmids. Repression is not restricted to B cells nor is it species or promoter specific. Repression is independent of the location of the GAL4-binding sites relative to the transcription start site. A fragment of EBNA3C (amino acids 280 to 525) which represses expression in a manner which is nearly identical to that of the full-length protein has been identified; this fragment is rich in acidic and proline residues. A second, less potent repressor region located C terminal to amino acids 280 to 525 has also been identified; this domain is rich in proline and glutamine residues. We also show binding of EBNA3C, in vitro, to the TATA-binding protein component of TFIID, and this suggests a mechanism by which EBNA3C may communicate with the basal transcription complex.
Bain
M
M
Ludwig Institute for Cancer Research, Imperial College School of Medicine at St. Mary's, London, United Kingdom.
Watson
R J
RJ
Farrell
P J
PJ
Allday
M J
MJ
D016475:3T3 Cells; D000818:Animals; D000956:Antigens, Viral / Q000235:genetics / Q000502:physiology*; D001402:B-Lymphocytes / Q000821:virology; D001665:Binding Sites; D002472:Cell Transformation, Viral; D004247:DNA / Q000378:metabolism*; D004268:DNA-Binding Proteins / Q000235:genetics / Q000502:physiology*; D019309:Epstein-Barr Virus Nuclear Antigens; D005656:Fungal Proteins / Q000235:genetics; D015967:Gene Expression Regulation, Viral; D004854:Herpesvirus 4, Human / Q000235:genetics / Q000502:physiology*; D006801:Humans; D051379:Mice; D011401:Promoter Regions, Genetic; D011485:Protein Binding; D011993:Recombinant Fusion Proteins / Q000235:genetics; D012097:Repressor Proteins / Q000502:physiology*; D029701:Saccharomyces cerevisiae Proteins; D014157:Transcription Factors; D014158:Transcription, Genetic; D014407:Tumor Cells, Cultured
D000956:Antigens, Viral; D004268:DNA-Binding Proteins; D019309:Epstein-Barr Virus Nuclear Antigens; D005656:Fungal Proteins; C052130:GAL4 protein, S cerevisiae; D011993:Recombinant Fusion Proteins; D012097:Repressor Proteins; D029701:Saccharomyces cerevisiae Proteins; D014157:Transcription Factors; D004247:DNA