Transformation by purified early genes of simian virus 40

Abstract

A rapid soft-agar assay using baby hamster kidney (BHK21 cl.13) cells has been developed to establish the functional roles for the large T and small t antigens of SV40 in transformation. Plasmids expressing either large T or small t antigens of SV40 have also been constructed and these plasmids have been used separately or in combination for transformation. A large T clone, pD3-05, containing a deletion in the small t-specific coding region [0.584-0.54 map units (mu)], transformed a low-background subclone of baby hamster kidney (BHK21 cl.13) cell line and F111 rat fibroblasts to anchorage independence at a low level (10-20 and 1%, respectively, of an early region clone from wild type [WT], pW2). A WT-derived small t clone, pW2-t, containing a deletion in the large T-specific coding region (0.373-0.169 mu), did not transform F111 cells, but transformed BHK21 cells at a very low level (about 2% of pW2). Another WT-derived small t clone, pW2-t/B1, containing a larger deletion in the large T-specific coding region (0.512-0.169 mu), did not transform either BHK21 or F111 cells. However, cotransformation with pD3-05 clone and pW2-t or pW2-t/B1 clone increased the frequency of transformation to about the same level as that of pW2. The ability of the small t clones to enhance the transformation efficiency of the large T clone was not due to recombination between the two plasmids, since cotransformation with pD3-05 and a small t clone without the polyadenylation [poly(A)] signal sequence from WT, pW-t8, did not increase the frequency of transformation. When the frequency of transformation was determined by the focus assay using F111 cells, pD3-05 transformed as well as pW2. Also, cotransformation with pD3-05 and pW2-t/B1 did not increase the frequency of focus formation. Therefore, the small t antigen was not required for this morphological transformation.