Delta-induced notch signaling mediated by RBP-J inhibits MyoD expression and myogenesis

Signaling induced by interaction between the receptor Notch and its ligand Delta plays an important role in cell fate determination in vertebrates as well as invertebrates. Vertebrate Notch signaling has been investigated usi ng its constitutively active form, i.e. the truncated intracellular region which is believed to mimic Notch-Delta signaling by interaction with a DNA- binding protein RBP-J. However, the molecular mechanism for Notch signaling triggered by ligand binding, which leads to inhibition of differentiation, is not clear. We have established a myeloma cell line expressing mouse Del tal on its cell surface which can block muscle differentiation by co-cultur e with C2C12 muscle progenitor cells. We showed that Delta-induced Notch si gnaling stimulated transcriptional activation of RBP-J binding motif, conta ining promoters including the HES1 promoter. Furthermore, ligand-induced No tch signaling up-regulated HES1 mRNA expression within 1 h and subsequently reduced expression of MyoD mRNA, Since cycloheximide treatment did not inh ibit induction of HES1 mRNA, the HES1 promoter appears to be a primary targ et of activated Notch. In addition, a transcriptionally active form of RBP- J, i.e. VP16-RBP-J, inhibited muscle differentiation of C2C12 cells by bloc king the expression of MyoD protein. These results suggest that HES1 induct ion by the Deltal/Notch signaling is mediated by RBP-J and blocks myogenic differentiation of C2C12 cells by subsequent inhibition of MyoD expression.