Temperature-sensitive transformation by Rous sarcoma virus and temperature-sensitive protein kinase activity

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Abstract
The transforming protein of Rous sarcoma virus, p60src, has associated with it a protein kinase activity. The correlation between the cellular concentration of enzymatically active p60src and the degree to which chick cells are transformed by mutants of Rous sarcoma virus which are temperature-sensitive for transformation was examined. Such a correlation does exist, but cells infected with some mutants contained, at the nonpermissive temperature, an amount of protein kinase activity equal to 30-40% of that in a wild-type transformed cell. The amount of virus-induced protein kinase activity was quantified by precipitation of p60src with an excess of antitumor antiserum. Initial measurements of activity were serious underestimates due to the lability of the protein kinase activity associated with p60src of at least 4 temperature-sensitive mutants. No activity at all was associated with p60src of tsLA90 when immunonoprecipitation was performed by standard means. When immunoprecipitation was performed with procedures which minimize inactivation, cells transformed by tsLA90 contained protein kinase activity and cells infected with NY68 or BK5 contained at the nonpermissive temperature, 1/3-1/2 as much activity as wild-type transformed cells. This level of activity was much more than that arising from p60src in uninfected cells. Uninfected cells had 3-5% as much kinase activity as that in a virally transformed cell. The lability of the protein kinase activity of each of these mutants is a further demonstration that this activity is essential for the transformation of cells by Rous sarcoma virus. To explain the high protein kinase levels in cells infected with NY68 and BK5 at the nonpermissive temperature, the idea that transformation may be a response to a small quantitative change in the total activity of p60src and the possibility that there may be more than 1 viral function which is essential for transformation are discussed.