TRUNCATED MAMMALIAN NOTCH1 ACTIVATES CBF1 RBPJK-REPRESSED GENES BY A MECHANISM RESEMBLING THAT OF EPSTEIN-BARR-VIRUS EBNA2/

The Notch/Lin-12/Glp-1 receptor family participates in cell-cell signa ling events that influence cell fate decisions. Although several Notch homologs and receptor ligands have been identified, the nuclear event s involved in this pathway remain incompletely understood. A truncated form of Notch, consisting only of the intracellular domain (NotchIC), localizes to the nucleus and functions as an activated receptor. Usin g both an in vitro binding assay and a cotransfection assay based on t he two-hybrid principle, we show that mammalian NotchIC interacts with the transcriptional repressor CBF1, which is the human homolog of Dro sophila Suppressor of Hairless. Cotransfection assays using segments o f mouse NotchIC and CBF1 demonstrated that the N-terminal 114-amino-ac id region of mouse NotchIC contains the CBF1 interactive domain and th at the cdc10/ankyrin repeats are not essential for this interaction. T his result was confirmed in immunoprecipitation assays in which the N- terminal 114-amino-aeid segment of NotchIC, but not the ankyrin repeat region, coprecipitated with CBF1. Mouse NotchIC itself is targeted to the transcriptional repression domain (aa179 to 361) of CBF1. Further more, transfection assays in which mouse NotchIC was targeted through Gal4-CBF1 or through endogenous cellular CBF1 indicated that NotchIC t ransactivates gene expression via CBF1 tethering to DNA. Transactivati on by NotchIC occurs partially through abolition of CBF1-mediated repr ession. This same mechanism is used by Epstein-Barr virus EBNA2. Thus, mimicry of Notch signal transduction is involved in Epstein-Barr viru s-driven immortalization.