Characterization of a second Epstein-Barr virus-determined nuclear antigen associated with the BamHI WYH region of EBV DNA

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Abstract
The Epstein-Barr virus-determined nuclear antigen (EBNA) is the only known virally-determined component that is regularly associated with EBV-transformed cells. A main component of EBNA, herein designated EBNA-I, has been conclusively localized to the BamHI K fragment of the viral genome. EBNA-I is present in all EBV-carrying cell lines so far studied. Our current study deals with a 2nd component. The EBNA reaction detected by anticomplement immunofluorescence (ACIF) in Burkitt lymphoma lines Daudi, jijoye, and P3HR-I could be completely removed by preabsorption of sera with any 1 of these 3 lines, when tested against any other of them. The same absorbed sera still gave a brilliant nuclear staining against other EBV-carrying lines, e.g. Raji or B95-8. The 3 lines in the 1st category carry EBV genomes that have deletions in the BamHI WYH region of the EBV genome. This region is intact in the 2nd group of lines. This result is interpreted as showing the existence of 2 different ACIF-stainable EBV-determined nuclear antigens, 1 of which is associated with the BamHI WYH region. This antigen was designated as EBNA-2. The 2 different EBNAs are different with regard to their association with metaphase chromosomes. In lines positive for both EBNA subtypes, metaphase chromosomes gave brilliant EBNA-I staining, but could not be stained for EBNA-2, indicating differences in chromatin association of the 2 EBNAs. An 86 kd polypeptide was identified by immunoblotting of DNA-binding proteins from EBV-transformed lymphoid cell lines. EBV-specificity of the polypeptide was demonstrated by the presence of antibodies against this polypeptide in antisera from a population of EBV-seropositive donors, but not from seronegative donors, by the presence of the polypeptide itself in EBV-carrying but not in EBV-negative cell lines and by the appearance of antibodies against this polypeptide during the course of infectious mononucleosis (IM). The polypeptide was absent from the EBV-carrying P3HR-I, Daudi and Jijoye cell lines, which suggested that it may be encoded by the BamHI WYH region that is deleted from the viral substrains carried by these lines.