Transformation of a continuous rat embryo fibroblast cell line requires three separate domains of simian virus 40 large T antigen.
Mouse C3H 10T1/2 cells and the established rat embryo fibroblast cell line REF-52 are two cell lines widely used in studies of viral transformation. Studies have shown that transformation of 10T1/2 cells requires only the amino-terminal 121 amino acids of simian virus 40 (SV40) large T antigen, while transformation of REF-52 cells requires considerably more of large T antigen, extending from near the N terminus to beyond residue 600. The ability of a large set of linker insertion, small deletion, and point mutants of SV40 T antigen to transform these two cell lines and to bind p105Rb was determined. Transformation of 10T1/2 cells was greatly reduced by mutations within the first exon of the gene for large T antigen but was only modestly affected by mutations affecting the p105Rb binding site or the p53 binding region. All mutants defective for transformation of 10T1/2 cells were also defective for transformation of REF-52 cells. In addition, mutants whose T antigens had alterations in the Rb binding site showed a substantial reduction in transformation of REF-52 cells, and the degree of this reduction could be correlated with the ability of the mutant T antigens to bind p105Rb. There was a tight correlation between the ability of mutants to transform REF-52 cells and the ability of their T antigens to bind p53. These results demonstrate that multiple regions of large T antigen are required for full transformation by SV40.
Zhu
J
J
Department of Biochemistry, Dartmouth Medical School, Hanover, New Hampshire 03755.
Rice
P W
PW
Gorsch
L
L
Abate
M
M
Cole
C N
CN
D000818:Animals; D000956:Antigens, Viral / Q000235:genetics* / Q000378:metabolism; D001483:Base Sequence; D002460:Cell Line; D002472:Cell Transformation, Viral / Q000235:genetics*; D004252:DNA Mutational Analysis; D004622:Embryo, Mammalian / Q000166:cytology; D005347:Fibroblasts / Q000473:pathology*; D008969:Molecular Sequence Data; D051381:Rats; D016160:Retinoblastoma Protein / Q000378:metabolism; D013539:Simian virus 40 / Q000235:genetics*; D014412:Tumor Virus Infections / Q000235:genetics*
D000956:Antigens, Viral; D016160:Retinoblastoma Protein