Differential subcellular localization regulates c-Cbl E3 ligase activity upon Notch3 protein in T-cell leukemia.

Notch3 and pTalpha signaling events are essential for T-cell leukemogenesis and characterize murine and human T-cell acute lymphoblastic leukemia. Genetic ablation of pTalpha expression in Notch3 transgenic mice abrogates tumor development, indicating that pTalpha signaling is crucial to the Notch3-mediated leukemogenesis. Here we report a novel direct interaction between Notch3 and pTalpha. This interaction leads to the recruitment and persistence of the E3 ligase protein c-Cbl to the lipid rafts in Notch3-IC transgenic thymocytes. Conversely, deletion of pTalpha in Notch3 transgenic mice leads to cytoplasmic retention of c-Cbl that targets Notch3 protein to the proteasomal-degradative pathway. It appears that protein kinase C theta (PKCtheta), by regulating tyrosine and serine phosphorylation of Cbl, is able to control its function. We report here that the increased Notch3-IC degradation correlates with higher levels of c-Cbl tyrosine phosphorylation in Notch3-IC/pTalpha(-/-) double-mutant thymocytes, which also display a decreased PKCtheta activity. Our data indicate that pTalpha/pre-T-cell receptor is able to regulate the different subcellular localization of c-Cbl and, by regulating PKCtheta activity, is also able to influence its ubiquitin ligase activity upon Notch3 protein.