Rearrangements of host and viral DNA in mouse cells transformed by simian virus 40.
We have determined the structure of host DNA and viral DNA at the site of integration of Simian virus 40 (SV40) in a line of transformed Balb/c-3T3 cells (SVB400) isolated by single cell cloning after virus infection. Recombinant phage containing integrated viral DNA and flanking host DNA were purified from a genomic library and, in conjunction with restriction endonuclease cleavage analysis of the transformed cell DNA, were used to determine the organization of the integrated viral sequences. There is heterogeneity in the arrangement of the viral sequences resulting from tandem duplications of all or part of the SV40 genome with preservation of the viral-host junctions. The predominant arrangement is the result of tandem duplication of 41% of the SV40 genome from 0.64 to 0.23. Analysis of the structure of integrated viral DNA in SVB400 at different passage numbers and in single cell clones derived from the 20th passage indicated that rearrangements of viral DNA occur after the integration event and continue with passage of the cells. The organization of host sequences before and after the integration of SV40 was determined by restriction endonuclease cleavage analysis of parental 3T3 DNA and SVB400 DNA, and by analysis of recombinant phage isolated from genomic libraries. A deletion of at least 15 X 10(3) bases of host DNA occurred at the site of integration, which indicates that viral integration was not a result of a simple insertion of SV40. Nucleotide sequence analysis of the virus-host junctions showed that retained SV40 sequences were colinear with the viral genome, and that the junctions with SV40 DNA occurred at nucleotide numbers 1377 and 3610. There was no evidence of duplications of viral or host sequences at the junctions, and a comparison of the flanking mouse sequences with the deleted SV40 sequences revealed no significant homology at the point of joining of the two genomes.
D000818:Animals; D001483:Base Sequence; D002471:Cell Transformation, Neoplastic; D002472:Cell Transformation, Viral; D002478:Cells, Cultured; D004247:DNA / Q000235:genetics*; D004262:DNA Restriction Enzymes; D004274:DNA, Recombinant; D004279:DNA, Viral / Q000235:genetics*; D004587:Electrophoresis, Agar Gel; D051379:Mice; D008807:Mice, Inbred BALB C; D008854:Microscopy, Electron; D009692:Nucleic Acid Heteroduplexes; D009693:Nucleic Acid Hybridization; D011995:Recombination, Genetic; D013539:Simian virus 40 / Q000235:genetics*
D004274:DNA, Recombinant; D004279:DNA, Viral; D009692:Nucleic Acid Heteroduplexes; D004247:DNA; D004262:DNA Restriction Enzymes