Functional interaction between SEL-10, an F-box protein, and the nuclear form of activated Notch1 receptor

The Notch signaling pathway is essential in many cell fate decisions in inv ertebrates as well as in vertebrates. After ligand binding, a two-step prot eolytic cleavage releases the intracellular part of the receptor which tran slocates to the nucleus and acts as a transcriptional activator. Although N otch-induced transcription of genes has been reported extensively, its endo genous nuclear form has been seldom visualized. We report that the nuclear intracellular domain of Notch1 is stabilized by proteasome inhibitors and i s a substrate for polyubiquitination in vitro. SEL-10, an F-box protein of the Cdc4 family, was isolated in a genetic screen for Lin12/Notch-negative regulators in Caenorhabditis elegans. We isolated human and murine counterp arts of SEL-10 and investigated the role of a dominant-negative form of thi s protein, deleted of the F-box, on Notch1 stability and activity. This mol ecule could stabilize intracellular Notch1 and enhance its transcriptional activity but had no effect on inactive membrane-anchored forms of the recep tor. We then demonstrated that SEL-10 specifically interacts with nuclear f orms of Notch1 and that this interaction requires a phosphorylation event. Taken together, these data suggest that SEL-10 is involved in shutting off Notch signaling by ubiquitin-proteasome-mediated degradation of the Active transcriptional factor after a nuclear phosphorylation event.