Figure 5.

Inhibition of NOTCH signaling in PCT by DNMAML1. A, PCR analysis (left) of expression of DNMAML1; the NOTCH targets Dtx1, Hey1, Dll3, and Myc; and β-actin in the ABPC8298 PCT cell line infected with viruses expressing EGFP alone or human DNMAML1 and EGFP. The MAML1 primers are specific for the human transcript. Right, quantitative analyses of NOTCH target gene expression in ABPC8298 expressing DNMAML1. Data are normalized to level of expression in the PCT cell lines expressing only EGFP. Columns, mean compared with the expression in cells expressing EGFP only for three independent experiments; bars, SE. *, significantly different from expression in EGFP-only cells (P < 0.05). B, analyses of cell proliferation. Left, proliferation assay in PCT cell lines expressing DNMAML or EGFP only; middle, percent of cells incorporating BrdUrd after 30 min in culture (*, P < 0.05); right, Western blot analysis of CDK4, CDK6, pRb, and α-tubulin expression. C, analyses of cell death in cultured ABPC8298 cells: percent of cells staining with Annexin V. Left, columns, mean of three independent experiments; bars, SE. *, P < 0.05. Middle, capase-3 activity determined in the cell lysates (black columns) or cell lysates incubated with the caspase-3 inhibitor. **, P < 0.005. Right, Western blot analyses of caspase-8, cleaved caspase-3, and α-tubulin levels.