NBEA deficiency perturbs NOTCH proteins expression and cell localization. (A) Representative images of OPM2 cell lines expressing different NBEA-shRNAs (1 and 2) and control-shRNA. Transduced OPM2 cells for each shRNA were sorted based on mCherry⁺ expression. (B) Cell growth curves for OPM2 transduced cells. (C) Annexin V analysis in control- and NBEA-shRNA OPM2 cells. Percentage of annexin V⁺ cells is shown. (D-G) NOTCH1 and NOTCH2 cell surface and total expression in OPM2 transduced cells. (D) Experimental schematic. (E) Flow cytometry histograms showing NOTCH1 and NOTCH2 protein levels in the cell surface and total expression for each OPM2 cell line. (F) Quantification of NOTCH1 and NOTCH2 protein levels normalized to control-shRNA. (G) Ratio of cell surface and total levels of NOTCH1 and NOTCH2. (H) qRT-PCR analysis of NOTCH-regulated genes in OPM2 cell lines. Data are represented as mean ± SEM. ∗P < .05; ∗∗P < .005; ∗∗∗P < .001, relative to control-shRNA. ∗ refers to NBEA-shRNA 1; # refers to NBEA-shRNA 2. Ab, antibody.