Epstein-Barr virus nuclear antigen 3C putative repression domain mediates coactivation of the LMP1 promoter with EBNA-2.
The Epstein-Barr virus (EBV) nuclear antigen 3C (EBNA-3C) regulates virus and cell genes and is essential for EBV-mediated transformation of primary B lymphocytes. EBNA-3C associates with the cellular DNA sequence-specific transcription factors RBP-Jkappa and PU.1 and coactivates the EBV LMP1 promoter with EBNA-2 in BL2 and Raji cells under conditions of restrictive growth. We now find that EBNA-3C is similar to EBNA-LP in coactivating the LMP1 promoter with EBNA-2 in non-EBV-infected Burkitt lymphoma cells under conditions of maximal cell growth, whereas the EBV Cp promoter is repressed under the same conditions. EBNA-3A and EBNA-3B coactivation are at most 40% that of EBNA-3C. The RBP-Jkappa binding sites of EBNA-2 and the LMP1 promoter are not required for EBNA-3C coactivation, whereas the PU.1 site in the LMP1 promoter is required for EBNA-2-mediated activation and EBNA-3C coactivation. EBNA-3C amino acids (aa) 365 to 545, including most of the previously identified repression domain (M. Bain, R. J. Watson, P. J. Farrell, and M. J. Allday, J. Virol. 70:2481-2489, 1996), are necessary and sufficient for coactivation with wild-type EBNA-2. EBNA-3C can also coactivate with the EBNA-2 acidic activating domain; this activation does not require aa 343 to 545. These data indicate that there are at least two mechanisms by which EBNA-3C coactivates the LMP1 promoter with EBNA-2. Of the proteins that interact with EBNA-3C in a yeast two-hybrid screen, only the ubiquitin-like proteins SUMO-1 and SUMO-3/hSMT3B map to aa 365 to 545, implicating these molecules in EBNA-3C coactivation. In addition, SUMO-1 associates at a high level with EBNA-3C in lymphoblasts. Promoter coactivation by EBNA-3C is likely to be important in ensuring adequate levels of LMP1, while inhibition of the EBNA-Cp promoter under the same conditions prevents uncontrolled up-regulation of EBNA expression from a positive-feedback loop.
Lin
Jeffrey
J
Virology Program and Department of Medicine, Channing Laboratory, Brigham and Women's Hospital and Harvard Medical School, Boston, Massachusetts 02115, USA.
Johannsen
Eric
E
Robertson
Erle
E
Kieff
Elliott
E
D000956:Antigens, Viral / Q000235:genetics / Q000378:metabolism* / Q000502:physiology*; D001402:B-Lymphocytes / Q000821:virology; D002460:Cell Line; D002472:Cell Transformation, Viral; D019309:Epstein-Barr Virus Nuclear Antigens / Q000378:metabolism*; D006801:Humans; D008213:Lymphocyte Activation; D011401:Promoter Regions, Genetic; D012097:Repressor Proteins / Q000235:genetics / Q000502:physiology*; D014158:Transcription, Genetic; D014763:Viral Matrix Proteins / Q000235:genetics / Q000378:metabolism*; D014764:Viral Proteins
D000956:Antigens, Viral; C402641:EBNA-2 protein, Human herpesvirus 4; C498774:EBNA-3C, epstein-barr virus; C037662:EBV-associated membrane antigen, Epstein-Barr virus; D019309:Epstein-Barr Virus Nuclear Antigens; D012097:Repressor Proteins; D014763:Viral Matrix Proteins; D014764:Viral Proteins