Guanine nucleotide-binding activity as an assay for src protein of rat-derived murine sarcoma viruses

Abstract

A 21,000 dalton protein, p21 coded for by Kirsten or Harvey murine sarcoma virus, was recently identified. On the basis of the results obtained with the p21 of a mutant of Kirsten sarcoma virus, temperature sensitive for the maintenance of transformation, it was concluded that the p21 was required for the maintenance of transformation induced by either virus. When extracts from cells transformed by Kirsten or Harvey sarcoma virus are incubated with [3H]GDP or [a-32P]GTP, picomole quantities of guanine nucleotide can be immunoprecipitated with antisera that contain antibodies to the p21. Previously it was shown that the temperature-sensitive mutant of Kirsten, sarcoma virus is thermolabile. The binding of guanine nucleotide is also thermolabile in extracts of cells transformed by the same mutant. The immunoprecipitability of the [35S] methionine-labeled p21 in such extracts of the temperature-sensitive mutant can be preserved if the extracts containing labeled p21 are incubated with added GDP or GTP prior to heating. The results suggest an interaction between p21 and certain guanine nucleotides, and the possible roles of guanine nucleotides and p21 in the maintenance of transformation are discussed. [The cells used were mouse 3T3 fibroblasts, rat NRK cells, canine MDCK kidney cells, mink CCL64 lung cells. The other viruses were Moloney murine leukemia virus, Friend type C virus, rat sarcoma virus, Moloney murine sarcoma virus, Snyder-Theilin strain of feline sarcoma virus and 3 strains of Rous sarcoma virus.].