FIGURE 3.

Effect of second generation GSMs on Notch, EphB2, and EphA4 processing. A, none of the AZ GSMs (square, triangle, and ×) or E2012 (○) affect Notch processing, whereas the GSIs L685458, DBZ, MRK-560, and Semagacestat do, as measured by quantifying the amount of nuclearly translocated NICD in HEK293 cells stably transfected with ΔENotch using immunocytochemistry. % NICD translocation is set relative to 0.5% DMSO (100%) and 0.5 μm L685458 (0%) controls (mean ± S.E., n = 2). B, HEK293 cells, transiently transfected with Myc-tagged EphB2, EphA4, and ΔENotch constructs, were exposed to 100 nm DBZ or 1 μm 4800 for 15 h prior to Western blot analysis. DBZ, but not AZ4800, results in accumulation of respective protein, as explored with an anti-Myc antibody (left panel). Each Western blot was probed with an anti-GAPDH antibody to control for loaded protein (right panel). The figure shows one representative blot out of at least three independent experiments.