Biological and biochemical properties of Ret with kinase domain mutations identified in multiple endocrine neoplasia type 2B and familial medullary thyroid carcinoma

Several mutations were identified in the kinase domain of the RET proto-onc ogene in patients with multiple endocrine neoplasia (MEN) 2B, familial medu llary thyroid carcinoma (FMTC) or sporadic medullary thyroid carcinoma. We introduced seven mutations (glutamic acid 768-->aspartic acid (E768D), vali ne 804-->leucine (V804L), alanine 883-->phenylalanine (A1883F), serine 891- ->alanine (S891A), methionine 918 -->threonine (M918T), alanine 919-->proli ne (A919P) and E768D/A919P) into the short and long isoforms of RET cDNA an d transfected the mutant cDNAs into NIH3T3 cells, The transforming activity of the long isoform of Ret with each mutation was much higher that that of its short isoform, Based on the levels of the transforming activity, these mutant RET genes were classified into two groups; a group with high transf orming activity (A883F, M918T and E768D/A919P) and a group with low transfo rming activity (E768D, V804L, S891A and A919P) (designated high group and l ow group). Interestingly, the level of transforming activity correlated wit h clinical phenotypes; high group Bet with the A883F or M918T mutation and low group Ret with the E768D, V804L or S891A mutation were associated with the development of MEN 2B and FMTC, respectively. in addition, we found tha t substitution of phenylalanine for tyrosine 905 present in the kinase doma in abolished both transforming and autophosphorylation activities of low gr oup Ret whereas it did not affect the activities of high group Ret.