Revertants of an ASV-transformed rat cell line have lost the complete provius or sustained mutations in src

We have isolated and characterized 12 revertants of a clonal line (B31) of avian sarcoma virus (ASV)-transformed rat-1 cells. The B31 cells contain a single normal ASV provirus, display the classical features of virally transformed cells, and revert to normal phenotype at low frequency. Revertants isolated after selective killing of transformed cells resemble uninfected rat-1 cells morphologically, fail to grow in suspension, and are at least 100-fold less tumorigenic than B31 cells. Two mechanisms of reversion have been identified in these cells. (i) Three of the revertant lines have lost the entire provirus, including both copies of the sequences repeated at the ends of the provirus; the manner in which the provirus is lost is not known. (ii) The other nine revertants retain a provirus of normal size and unaltered flanking cellular DNA: contain the same species of viral RNA at the same concentrations as in the parental line, B31; are susceptible to retransformation by wild-type ASV; and yield transformation-defective (td) virus after fusion with chicken cells. In one case, the rescued virus transforms chicken cells, but produces fusiform rather than normal foci and does not retransform rat cells morphologically. Hence these revertants arise as a consequence of nonconditional mutations (base substitutions or small deletions) in the viral transforming gene,src. In several cases, the revertant cells retransform spontaneously, or transforming virus appears in stocks of rescued td virus after passage through chicken cells, indicating back mutations to wild-type. Several of the rescued td viruses can also recombine to restore a wild-type phenotype. Analysis of the structure and enzymatic activity of products ofsrc confirms that the revertant cells bear various mutations insrc.