Identification and characterization of c-raf phosphoproteins in transformed murine cells.

Protein products of the c-raf gene were detected and characterized in two transformed murine cells lines by immunoprecipitation analysis with raf-specific sera. Both proteins reacted with an antiserum directed to the carboxyl terminus of the c-raf coding region. The p48raf (Mr48,000) phosphoprotein was found in a cell line transformed by an LTR-activated c-raf gene (Mueller & Mueller, 1984). It was found to be phosphorylated on serine and threonine residues and became phosphorylated in immunoprecipitates when supplied with [gamma-32]ATP. In contrast, P74raf, which was detected into a spontaneously transformed 3T3 (R+/Cl 3) cell line and may represent the full-length gene product of c-raf, appeared to incorporate [32P]PO4 less efficiently in vivo and exhibited a barely detectable associated kinase activity in only half of the experiments. The p48raf is a transforming protein which, like P75gag-raf, lacks the amino-terminal region of the c-raf coding region. P74raf, which retains the amino-terminal region, differs from p48raf in its phosphorylation characteristics and may not be a transforming protein. These data are consistent with a model in which lack of amino-terminal sequences in the c-raf gene product p48raf may, in and of itself, suffice to make it a transforming protein.