Evidence for two classes of chromatin-associated Epstein-Barr virus-determined nuclear antigen

Abstract

A new class of Epstein-Barr virus nuclear antigen (EBNA) was identified by the complement fixation assay. This new species of EBNA is more tightly bound to chromatin and was termed class II EBNA, as opposed to the more weakly associated species, class I EBNA. Preparations of this new antigen(s) specifically reduced absorption with the titer of anti-EBNA antibodies as determined by the anticomplement immunofluorescence assay. Therefore, the complement fixation antigens (class II EBNA) appear to be related to the classical EBNA (class I EBNA). The class I EBNA was found to focus at the same pH (4.6) as the soluble antigen found in the cytosol. The class II EBNA differed from the class I EBNA with regard to its overall charge, molecular size, antigenicity, and affinity for chromatin. The class II EBNA appeared to be a basic protein, based on its apparent pl of 9.2 and its binding to cation-exchange resins. It differed from histones with regard to its molecular size (molecular weight between 60,000 and 70,000) and its elution from hydroxylapatite chromatography. Steps were taken to prevent proteolysis and artifacts in the immunological assays and in the overall charge estimation of the new antigen by nonspecific basic histone protein-acidic protein interactions. Both class I and class II EBNA were identified by radioimmunoelectrophoresis on two-dimensional polyacrylamide gels with pI values of 5.0 and 8.5, respectively, and a molecular weight range of 60,000 to 70,000 for both. A lower-molecular-weight antigen identified by molecular sieve chromatography appeared to be due to interference by histones in the immunoassays since it was not observed by the two-dimensional gel electrophoresis. Further characterization of this class II EBNA is in progress.

Identification and characterization of a cellular protein that cross-reacts with the Epstein-Barr virus nuclear antigen

“…The 72K EBNA protein is heterogeneous in isoelectric point and chromatin binding (16,17;Luka et al., submitted for publication) and is encoded by an EBV DNA fragment which contains the IR3 repetitive element (2, 3, 5-7). The correct reading frame for translation of this EBV gene is known from the nucleotide sequence of a fusion gene between EBV IR3 and lac Z since the fusion gene codes for a protein with epitopes of EBNA (8).…”

Induction of Epstein-Barr virus nuclear antigens

“…More recent evidence has demonstrated the existence of multiple EBV-specific antigens in EBV-transformed cells. The most prominent antigen (EBNA 1) is a polypeptide partially encoded by the BamHI K fragment, and the second (EBNA 2) is a polypeptide encoded by the BamHI WYH region (3,4,6,8,(19)(20)(21).…”

Identification of multiple Epstein-Barr virus-induced nuclear antigens with sera from patients with rheumatoid arthritis

“…Strnad et al (24), using fluoroimmunoelectrophoresis to detect EBNA, noted the presence of an additional 81,000molecular-weight (81K) antigen in B95-8 and Raji cells. Spelsberg et al (23) presented evidence for the existence of two distinct components of the EBV-induced nuclear antigen in NC37 cells. Hennessy and Kieff (7), employing protein immunoblots, observed that some EBV-immune human antisera recognized a second intracellular antigen that was specific for latently infected cells.…”