Epstein-Barr Virus (EBV) is a human herpes virus which infects and transforms B lymphocytes. It is the causative agent of infectious mononucleosis and is strongly associated with two human tumors, Burkitt's lymphoma and nasopharyngeal carcinoma. Details of the molecular biology of this virus are still poorly understood; such studies have been severely hampered by the lack of an in vitro system which is completely permissive for viral replication. Although the entire DNA sequence of EBV was recently determined, very few gene products have been assigned to specific open reading frames. We propose to identify and map the genes for EBV nucleocapsid components and to initiate studies on the regulation of EBV gene expression by studying the control of EA (early antigen) synthesis. We have developed a method for the purification of biologically active EB virions. We now propose to use these virions to develop monoclonal antibodies specific for individual virion components; these antibodies will be used to screen for expression of the protein in cells transfected with various regions of the EBV genome inserted into expression vectors. Once the general region coding for a particular component has been identified, the precise boundaries of the gene will be delineated by S1 nuclease mapping. The results of these experiments will identify the various polypeptides which comprise the EB nucleocapsid and determine which segment of the EBV genome codes for them. In addition, as the first step in understanding the control of viral replication in B lymphocytes, we will investigate the mechanism by which the synthesis of early antigen is controlled. EA appears to be one of the first gene products expressed at the start of the replication cycle. The putative genes for EA will be placed in an SV40 expression vector to verify that these genes do, in fact, code for the EAs previously identified in EBV positive cells using specific antibodies. The upstream regulatory sequences of these EAs will be studied for their role in the chemically induced expression of EA using site-specific mutagenesis. The results of these experiments will greatly enhance our knowledge of EBV replication and of the control of EBV gene expression. The information gained concerning the regulation of EA expression is likely to reflect a general mechanism of eukaryotic gene control.

Agency
National Institute of Health (NIH)
Institute
National Cancer Institute (NCI)
Type
Research Project (R01)
Project #
1R01CA043532-01
Application #
3185720
Study Section
Virology Study Section (VR)
Project Start
1986-04-01
Project End
1989-03-31
Budget Start
1986-04-01
Budget End
1987-03-31
Support Year
1
Fiscal Year
1986
Total Cost
Indirect Cost
Name
Johns Hopkins University
Department
Type
Schools of Medicine
DUNS #
045911138
City
Baltimore
State
MD
Country
United States
Zip Code
21218
Murray, P G; Swinnen, L J; Constandinou, C M et al. (1996) BCL-2 but not its Epstein-Barr virus-encoded homologue, BHRF1, is commonly expressed in posttransplantation lymphoproliferative disorders. Blood 87:706-11
Esolen, L M; Park, S W; Hardwick, J M et al. (1995) Apoptosis as a cause of death in measles virus-infected cells. J Virol 69:3955-8
Carruth, L M; Hardwick, J M; Morse, B A et al. (1994) Visna virus Tat protein: a potent transcription factor with both activator and suppressor domains. J Virol 68:6137-46
Hardwick, J M; Tse, L; Applegren, N et al. (1992) The Epstein-Barr virus R transactivator (Rta) contains a complex, potent activation domain with properties different from those of VP16. J Virol 66:5500-8
Cox, M A; Leahy, J; Hardwick, J M (1990) An enhancer within the divergent promoter of Epstein-Barr virus responds synergistically to the R and Z transactivators. J Virol 64:313-21
Lieberman, P M; Hardwick, J M; Sample, J et al. (1990) The zta transactivator involved in induction of lytic cycle gene expression in Epstein-Barr virus-infected lymphocytes binds to both AP-1 and ZRE sites in target promoter and enhancer regions. J Virol 64:1143-55
Lieberman, P M; Hardwick, J M; Hayward, S D (1989) Responsiveness of the Epstein-Barr virus NotI repeat promoter to the Z transactivator is mediated in a cell-type-specific manner by two independent signal regions. J Virol 63:3040-50
Hardwick, J M; Lieberman, P M; Hayward, S D (1988) A new Epstein-Barr virus transactivator, R, induces expression of a cytoplasmic early antigen. J Virol 62:2274-84