RET(MEN 2B) is active in the endoplasmic reticulum before reaching the cell surface

MEN 2B ( multiple endocrine neoplasia type 2B) is an autosomal dominant cancer syndrome caused by an oncogenic form of the receptor tyrosine kinase REarranged during transfection (RET). The MEN 2B syndrome is associated with an abnormal autophosphorylation of the mutated receptor even without ligand-stimulation. Here, we characterize the activation of a RETMEN2B variant carrying the point mutation Met918Thr, and show that the 150 kDa precursor of RETMEN2B becomes phosphorylated already during synthesis in the endoplasmic reticulum (ER). At least three different tyrosine residues (Tyr905, Tyr1062, Tyr1096) of the RETMEN2B precursor are phosphorylated before the oncogenic receptor reaches the cell surface. We also demonstrate that the precursor of RETMEN2B interacts with both growth factor receptor-bound protein and Src homology 2 domain-containing already in the ER, and that this interaction is dependent on the kinase activity of RET. With the aid of two RET mutants (RETMEN2B/S32L and RETMEN2B/ F393L), which accumulate in the ER, we show that the oncogenic precursor of the receptor has the capacity to activate AKT, extracellular signal-regulated kinase and signal transducer and activator of transcription 3 from the ER. Taken together, our data demonstrate that the oncogenic precursor of RETMEN2B is phosphorylated, interacts with adapter proteins and induces downstream signalling from the ER.