Fig. 1. Synergistic effect of p65-NFB and Notch-IC in the transactivation of the Hes1 promoter. (A) N1-IC (0.5 µg) was cotransfected along with the Hes1-luc reporter (0.75 µg) in NIH-3T3 cells and increasing amounts of p65 wt. Lower panels show N1-IC and p65 protein levels. Protein loading for each lane was normalized according to ß-galactosidase expression. (B,C) p65-NFB can synergize with different Notch-IC homologs. 0.5 µg of N2-IC (B) or N3-IC (C) were cotransfected with p65 wt (0.5 µg) along with the Hes1-luc reporter plasmid (1µg) in NIH-3T3 cells. (D) Synergistic effect of p65-NFB and Notch-IC is not modified by ectopic expression of p50-NFB. NIH-3T3 cells were cotransfected with different combinations of equivalent amounts (0.5 µg) of p50 subunit, N1-IC and p65 wt expression vectors. Luciferase activity is represented as the fold induction relative to the basal level measured in cells transfected with empty vector. The average and standard deviation of duplicates from a representative experiment are presented.