Episomal Viral DNA in a Herpesvirus saimiri -Transformed Lymphoid Cell Line

Abstract

The lymphoid cell line #1670 was derived from the infiltrated spleen of a tumor-bearing marmoset monkey infected with Herpesvirus [H.] saimiri. The cells contain both types of H. saimiri DNA, unique light (L-) DNA (36% cytosine plus guanine) and repetitive heavy (H-) DNA (71% cytosine plus guanine), without producing infectious virus. Viral DNA persisted in these cells as nonintegrated circular DNA molecules. Closed circular superhelical viral DNA molecules were isolated by 3 subsequent centrifugation steps: isopycnic centrifugation in CsCl, sedimentation through glycerol gradients and equilibrium centrifugation in CsCl-ethidium bromide. The iolated circles had a MW of 131.5 .times. 106. This is significantly higher than the MW of linear DNA molecules isolated from purified H. saimiri virions (about 100 .times. 106). Partial denaturation mapping of circular molecules from #1670 lymphoid cells showed uniform arrangement of H- and L-DNA sequences in all circles. All denatured molecules contained 2 L-DNA regions (MW of 54.0 .+-. 1.8 .times. 106 and 31.5 .+-. 1.3 .times. 106) and 2 H-DNA regions (MW of 25.6 .+-. 1.9 .times. 106 and 20.0 .+-. 0.8 .times. 106) of constant length. Maps of both L-regions suggested that the sequences of the shorter L-DNA region were a subset of those of the longer region. The sequences of both L-regions had the same orientation. Circular molecules from H. saimiri-transformed lymphoid cell line #1670 appeared to represent defective genomes, containing only 75% of the genetic information present in L-DNA of H. saimiri virions.