Non-immortalizing P3J-HR-1 Epstein-Barr virus: a deletion mutant of its transforming parent, Jijoye

Abstract

The P3J-HR-1 strain of Epstein-Barr virus (EBV) fails to immortalize human lymphocytes. We wished to understand the nature of the genomic alterations which correlated with the loss of this ability. As a first step, the heterogeneity of DNA molecules in the P3J-HR-1 line was eliminated by cell cloning. Then a physical map was prepared of virion DNA from one cell clone, designated FF452-3. By comparison with the genomes of two EBVs, B95-8 and FF41, which are competent to immortalize lymphocytes, we identified a total of eight modifications of BamHI and EcoRI restriction endonuclease fragments of EBV (FF452-3) DNA consisting of insertions, deletions, or loss of a restriction endonuclease recognition site. To determine which of these alterations might be responsible for the loss of transforming phenotype, we examined homologous DNA fragments of the Jijoye strain of EBV, the progenitor of the HR-1 strain which still retains the ability to immortalize lymphocytes. We also studied viral DNA in lymphocytes transformed in vitro by Jijoye virus. Six of the eight alterations were found both in Jijoye and in clonal HR-1 DNA and were presumably genomic traits characteristic of this lineage of EBV. A small deletion in the BamHI-K fragment of HR-1 DNA was not found in Jijoye virion DNA, but this deletion was present in intracellular Jijoye DNA. Thus only one major genomic lesion in HR-1 DNA, a deletion of at least 2.4 x 106 molecular weight of DNA from a fused BamHI-H-Y fragment, consistently distinguished Jijoye DNA from its non-immortalizing P3J-HR-1 derivative. This deletion is likely to affect EBV genes which are directly or indirectly involved in immortalizing lymphocytes.

Epstein-Barr virus-mediated B-cell proliferation is dependent upon latent membrane protein 1,which simulates an activated CD40 receptor

“…All cell lines were grown in RPMI 1640 culture medium supplemented with 10% fetal calf serum, 2 mM L-glutamine, 1 mM pyruvate, 100 U/ ml penicillin and 100 μg/ml streptomycin. HH514 is a single cell clone of the Burkitt's lymphoma cell line P3HR1 (Rabson et al, 1982). WI38, a human fibroblast cell line, was obtained from the American Type Culture Collection, ATCC.…”

Epstein-Barr virus latent membrane protein (LMP1) is not sufficient to maintain proliferation of B cells but both it and activated CD40 can prolong their survival.

“…The cell line was diluted 1:2 twice a week in standard RPMI 1640 medium supplemented with 1 ,uM estrogen. HH514 is a single cell clone of the Burkitt's lymphoma cell line P3HR1 (Rabson et al, 1982). MRC5, a human embryonic lung fibroblast cell line, was obtained from the American Type Culture Collection.…”

Epstein-Barr virus with heterogeneous DNA disrupts latency

“…Presently available stocks of P3J-HR-1 do not immortalize lymphocytes (15,16). This deficiency is correlated with a deletion of about 6,800 base pairs by comparison with its transformation-competent parent Jijoye (2,13,19) and other immortalizing EBVs (1,6,7,19).…”