Binding of Delta1, Jagged1, and Jagged2 to Notch2 rapidly induces cleavage, nuclear translocation, and hyperphosphorylation of Notch2

Delta1, Jagged1, and Jagged2, commonly designated Delta/Serrate/LAG-2 (DSL) proteins, are known to be ligands for Notch1. However, it has been less un derstood whether they are ligands for Notch receptors other than Notch1. Me anwhile, ligand-induced cleavage and nuclear translocation of the Notch pro tein are considered to be fundamental for Notch signaling, yet direct obser vation of the behavior of the Notch molecule after ligand binding, includin g cleavage and nuclear translocation, has been lacking. In this report, we investigated these issues for Notch2. All of the three DSL proteins bound t o endogenous Notch2 on the surface of BaF3 cells, although characteristics of Jagged2 for binding to Notch2 apparently differed from that of Delta1 an d Jagged1. After binding, the three DSL proteins induced cleavage of the me mbrane-spanning subunit of Notch2 (Notch2(TM)), which occurred within 15 mi n. In a simultaneous time course, the cleaved fragment of Notch2(TM) was tr anslocated into the nucleus. Interestingly, the cleaved Notch2 fragment was hyperphosphorylated also in a time-dependent manner. Finally, binding of D SL proteins to Notch2 also activated the transcription of reporter genes dr iven by the RBP-J kappa-responsive promoter. Together, these data indicate that all of these DSL proteins function as ligands for Notch2. Moreover, th e findings of rapid cleavage, nuclear translocation, and phosphorylation of Notch2 after ligand binding facilitate the understanding of the Notch sign aling.