A mutation at tyrosine 1062 in MEN2A-Ret and MEN2B-Ret impairs their transforming activity and association with shc adaptor proteins.
Germ line mutations of the ret proto-oncogene are associated with the development of three dominantly inherited neoplastic disorders, multiple endocrine neoplasia (MEN) 2A, MEN 2B, and familial medullary thyroid carcinoma. It has been demonstrated that the mutations result in constitutive activation of the Ret protein, leading to transformation of NIH 3T3 cells. In the present study we investigated the role of tyrosine residues present in the carboxyl-terminal sequence for the transforming activity of Ret with the MEN 2A or MEN 2B mutation (MEN2A-Ret or MEN2B-Ret). Substitution of phenylalanine for tyrosine 1062 (designated Y1062F) markedly impaired the transforming activity of both MEN2A-Ret and MEN2B-Ret, whereas substitution or deletion for four other tyrosines (codons 981, 1015, 1090, and 1096) did not affect their activity. The Shc adaptor proteins bound to the MEN2A-Ret and MEN2B-Ret proteins and were phosphorylated on tyrosine in the transfectants. The binding of Shc to the Y1062F mutant proteins was reduced by approximately 80%, indicating that tyrosine 1062 is a major binding site for Shc. In addition, phosphopeptide analysis of MEN2A-Ret demonstrated that tyrosine 1062 represents an autophosphorylation site of the mutant Ret proteins.
Asai
N
N
Department of Pathology, Nagoya University School of Medicine, 65 Tsurumai-cho, Showa-ku, Nagoya 466, Japan.
Murakami
H
H
Iwashita
T
T
Takahashi
M
M
D048868:Adaptor Proteins, Signal Transducing; D033942:Adaptor Proteins, Vesicular Transport; D000595:Amino Acid Sequence; D000818:Animals; D002471:Cell Transformation, Neoplastic / Q000235:genetics*; D029721:Drosophila Proteins; D006801:Humans; D051379:Mice; D008969:Molecular Sequence Data; D009377:Multiple Endocrine Neoplasia / Q000235:genetics*; D018813:Multiple Endocrine Neoplasia Type 2a / Q000235:genetics; D018814:Multiple Endocrine Neoplasia Type 2b / Q000235:genetics; D009154:Mutation; D011506:Proteins / Q000378:metabolism*; D000090063:Proto-Oncogene Mas; D011518:Proto-Oncogene Proteins / Q000235:genetics*; D051096:Proto-Oncogene Proteins c-ret; D020794:Receptor Protein-Tyrosine Kinases / Q000235:genetics*; D055765:Shc Signaling Adaptor Proteins; D015398:Signal Transduction; D000071425:Src Homology 2 Domain-Containing, Transforming Protein 1; D014443:Tyrosine / Q000235:genetics
D048868:Adaptor Proteins, Signal Transducing; D033942:Adaptor Proteins, Vesicular Transport; D029721:Drosophila Proteins; C000715470:MAS1 protein, human; D011506:Proteins; D000090063:Proto-Oncogene Mas; D011518:Proto-Oncogene Proteins; C528024:SHC1 protein, human; D055765:Shc Signaling Adaptor Proteins; C528034:Shc1 protein, mouse; D000071425:Src Homology 2 Domain-Containing, Transforming Protein 1; D014443:Tyrosine; D051096:Proto-Oncogene Proteins c-ret; D020794:Receptor Protein-Tyrosine Kinases; C416649:Ret protein, Drosophila; C494999:Ret protein, mouse