Transforming DNA sequences in rat cells transformed by DNA fragments of highly oncogenic human adenovirus type 12

Abstract

Rat cell lines transformed by viral DNA fragments, EcoRI-C and HindIII-G, of adenovirus type 12 DNA were analyzed for the viral transforming DNA sequences present in cell DNAs. Cell lines transformed by the EcoRI-C fragment of adenovirus type 12 DNA (leftmost 16.5% of the viral genome) contain most of the HindIII-G sequences (leftmost 6.8% of the viral genome; the smallest DNA fragment with complete transforming ability) at two to five copies per haploid cell DNA. Cell lines transformed by the HindIII-G fragment contain nearly all of the DNA sequences of the HindIII-G fragment, but at a different frequency depending on the portions of the fragment. The sequence of the AccI-H fragment of adenovirus type 12 DNA (the left part of the HindIII-G; leftmost 4.5% of the viral genome) was detected dominantly in cells transformed by the HindIII-G fragment. Southern blot analysis showed that viral DNA sequences are present at multiple integration sites in high-molecular-weight cell DNA from cells transformed by the EcoRI-C or HindIII-G fragment of adenovirus type 12 DNA. These results suggest that most of the HindIII-G sequences in cells transformed by the HindIII-G fragment are present as fragmented forms.

Mapping of adenovirus 12 mRNA's transcribed from the transforming region

“…2B). These data reflect the complicated integration of multiple copies of viral DNA in GYl-l cells (21).…”

“…The early region 1 sequences (0.9 to 11.2 map units [see below]) of Adl2 DNA are fully included in the EcoRI-C fragment (0 to 16.5 map units). CYl-1 cells contain the EcoRI-C fragment of Adl2 DNA at 5 to 6 copies per haploid cell (21). Four major and three minor virus-specific mRNA species were detected in CYl-1 cytoplasm, and all seven mRNA's had spliced structures (Fig.…”

“…DNA fragments were separated by 1.4% agarose gel electrophoresis and purified as described previously (23). Highly radioactive DNA fragments (5 x 107 to 2 x 108 cpm/4ug) were prepared with [a-32P]_ dATP and [a-32P]dCTP by the nick translation procedure as previously described (21).…”

Structure of viral DNA in a rat cell line transformed by the clonedEcoRI-Cfragment of adenovirus 12

“…According to the results of saturation hybridization and reassociation ©) I R L Press Limited, Oxford, England. kinetics, however, fragmented HindIII-G sequences occurred at high frequencies in GY1 cells(70 to 470 copies per haploid genome), whereas the viral copies estimated for CY1 cells were 2 to 5 per haploid genome (6). The results strongly suggest that the processes involved in the establishment of transformation were different with the HindIII-G and EcoRI-C fragments.…”