FIGURE 3.

The endonuclease activity of DFF40/CAD is necessary for apoptotic nuclear collapse and disassembly. IMR-5 cells stably transfected with empty pcDNA3 (Neo), pcDNA3-mCAD wild-type (mCAD WT), pcDNA3-mCADH242A (mCAD H242A), pcDNA3-mCADH263A (mCAD H263A), or pcDNA3-mCADH313A (mCAD H313A) plasmids were treated with 1 μm STP for 24 h or left untreated. A, total protein extracts from the different cell lines generated were analyzed by Western blot analysis to confirm the overexpression of the different mCAD variants. The membrane was reprobed with an antibody against actin as a loading control. B, genomic DNA analysis of the different stably transfected cell lines treated with STP (+) or left untreated (−). C, morphological aspect of the nuclei of the different cell lines stained with Hoechst 33258 after STP treatment or no treatment (Control). In each condition, the right panels are magnifications of the insets in the left panels. Scale bar = 40 μm.