When Epstein-Barr virus persistently infects B-cell lines, it frequently integrates.
In this study we used Gardella gel analysis of intact DNA, Southern blotting of digested DNA, and fluorescence in situ hybridization to provide complementary and unequivocal information on the state of the Epstein-Barr virus (EBV) genome in persistently infected cells. The fluorescence in situ hybridization technique allowed us to directly visualize both integrated and episomal EBV DNA at the single-cell level. We show here that circularization of the EBV genome is rarely detected upon infecting activated normal B cells. The virus can persist upon infection of a different proliferating B-cell target, EBV-negative Burkitt's lymphoma tumor cell lines. Analysis of 16 such lines reveal again, that the virus infrequently persists as covalently closed episomes; rather, the virus preferentially persists by integrating into the host DNA (10 of 16 clones). The integrated virus is linear and usually intact, although 3 of 10 isolates have deletions from the left-hand end including the latent origin of replication. At the level of our analysis, no obvious relationship was seen between the integration sites. These studies provide, for the first time, a reproducible in vitro model system to study integration by EBV.
Hurley
E A
EA
Department of Pathology, Tufts University School of Medicine, Boston, Massachusetts 02111.
Agger
S
S
McNeil
J A
JA
Lawrence
J B
JB
Calendar
A
A
Lenoir
G
G
Thorley-Lawson
D A
DA
D001402:B-Lymphocytes; D015139:Blotting, Southern; D002051:Burkitt Lymphoma; D002460:Cell Line; D002472:Cell Transformation, Viral; D002872:Chromosome Deletion; D004279:DNA, Viral / Q000235:genetics / Q000302:isolation & purification; D005814:Genes, Viral; D004854:Herpesvirus 4, Human / Q000235:genetics*; D006801:Humans; D008242:Lysogeny; D009693:Nucleic Acid Hybridization; D015183:Restriction Mapping
D004279:DNA, Viral