Alternative splicing of transcripts encoding the RET kinase receptor leads to isoforms differing in their cytoplasmic tail. Although in vitro studies have demonstrated a higher transforming activity of the long RET isoform (RET51), only the short isoform (RET9) can rescue the effects of a RET null mutation in the enteric nervous system and kidney development. The molecular basis underlying the distinct functions of the two RET isoforms is not understood. Here we demonstrated that activated RET51 associated more strongly with the ubiquitin ligase Cbl than did RET9, leading to increased ubiquitylation and faster turnover of RET51. The association of Cbl with RET was indirect and was mediated through Grb2. A constitutive complex of Grb2 and Cbl could be recruited to both receptor isoforms via docking of Shc to phosphorylated Tyr-1062 in RET. A mutant Shc protein unable to recruit the Grb2⅐Cbl complex decreased the turnover and prolonged the half-life of RET9, thus ascribing a previously unknown negative role to the Shc adaptor molecule. In addition, phosphorylation of Tyr-1096, which is present in RET51 but absent in RET9, endowed the longer isoform with a second route to recruit the Grb2⅐Cbl complex. These findings establish a mechanism for the differential down-regulation of RET9 and RET51 signaling that could explain the apparently paradoxical activities of these two RET isoforms. More generally, these results illustrate how alternative splicing can regulate the half-life and function of a growth factor receptor.The RET proto-oncogene encodes a transmembrane tyrosine kinase receptor that plays a crucial role in the development of the neural crest and the excretory system (1, 2). The RET protein is activated by engaging members of a family of structurally related ligands consisting of glial cell line-derived neurotrophic factor (GDNF), 1 neurturin, artemin, and persephin, in the presence of specific auxiliary receptors called GFR␣1-4 (GDNF family receptor alphas). Targeted deletion of the RET gene leads to severely impaired development or complete loss of kidneys (44,45). RET is expressed in the ureteric bud and mediates the growth and branching morphogenesis of this structure. The role of RET in neural crest development is highlighted in the establishment of the enteric nervous system. Neural crest progenitors that migrate to the foregut give rise to enteric neurons and glial cells. In the absence of RET, the rostrocaudal migration of enteric neural progenitors fails, leading to a lack of innervation of the intestinal tract posterior to the stomach (3). Survival and proliferative effects of RET signaling on progenitor cells of the enteric nervous system have also been proposed (4). Characteristic aganglionosis of the colon in Hirschsprung disease has also been linked to inactivating mutations of RET (5). The alternative splicing of the 3Ј exons of the RET gene can give rise to three distinct protein products (6, 7). The isoforms RET9, RET43, and RET51 have different sizes because of distinct amino acid sequences...
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