Figure 4. UCH L1 Suppression Induces G0/G1 Arrest and Apoptosis.

Ingenuity pathway analysis was performed on genes in common between UCH L1 siRNA-expressing 293T and KR4 cell lines, after which tox list was generated for genes affecting apoptosis and cell-cycle (A). Cell-cycle analysis was performed on UCH L1 siRNA-expressing 293T (left panel) and KR4 (right panel) cells (B) after culturing the cells under sub-optimal reduced serum (1%) conditions for 24 h. For the analysis, 10⁶ cells were collected, fixed and stained with propidium iodide and then analyzed with flow cytometry. Representative histograms are shown in which cell count vs DNA content was plotted. Cell-cycle statistics were generated with ModFit software. To measure apoptosis, 293T and KR4 cells (10⁴) were treated with 3 and 6 µM concentrations of camptothecin for 10 h in (1%) serum. The number of viable cells was measured with a Cell Death ELISA assay, which measures the production of histone-associated DNA fragments at 405 nm wavelength. Each reaction was performed in duplicate. Graphical representation of cell death rate in 293T (left panel) and KR4 (right panel) cells (C).