Helper-independent transformation by unintegrated Harvey sarcoma virus DNA

Abstract

The unintegrated infectious DNA of Harvey sarcoma virus (Ha-SV) and Moloney leukemia virus (Mo-MuLV) was studied. The source of infectious viral DNA was the Hirt supernatant fraction from [mouse fibroblast] cells acutely infected with Ha-SV and Mo-MuLV. To obtain a direct quantitative assay for infectious viral DNA, recipient mouse cells were first exposed to calcium phosphate-precipitated viral DNA and then treated with dimethyl sulfoxide. Infectivity was monitored by focus formation for Ha-SV and [rat sarcoma] XC [cell] plaque formation for Mo-MuLV. The viral DNA titration pattern followed single-hit kinetics for both foci and plaques, indicating that a single molecule carried information for each function. Focus-forming and plaque-forming activity were present in different molecules, since these 2 biological activities could be separated from each other by agarose gel electrophoresis. The focus-forming molecule was linear DNA with a MW of about 4 .times. 106 daltons. The focus-forming activity of the viral DNA was sensitive to EcoRI and resistant to XhoI restriction endonucleases, but the plaque-forming activity was resistant to EcoRI and sensitive to XhoI. The generation of helper-independent foci indicates that Ha-SV DNA can transform mouse cells in the absence of helper virus or its proteins.