Detection of phosphorylated retTPC oncogene product in cytoplasm

The product of the retTPC oncogene, an activated form of the ret proto-oncogene found in human papillary thyroid carcinomas, was identified as a 57-kDa protein (p57retTPC) by Western blotting with a polyclonal antibody raised by an oligopeptide corresponding to the carboxy-terminal region of the ret proto-oncogene product. Subcellular fractionation experiments using NIH3T3 cell transformants induced by the retTPC cDNA showed that p57retTPC was localized in a soluble cytoplasmic fraction, whereas the ret proto-oncogene products expressed in a neuroblastoma cell line were present in a membrane fraction. Immunostaining also demonstrated that p57retTPC is localized in the cytoplasm. Immunoprecipitation with anti-phosphotyrosine antibody followed by Western blotting revealed that p57retTPC is constitutively phosphorylated, whereas the ret proto-oncogene products are not. These findings suggest that p57retTPC has an aberrant tyrosine kinase activity resulting in autophosphorylation associated with change in its location.