CASPASE-3 IS REQUIRED FOR DNA FRAGMENTATION AND MORPHOLOGICAL-CHANGESASSOCIATED WITH APOPTOSIS

Interleukin 1 beta-converting enzyme-like proteases (caspases) are cru cial components of cell death pathways. Among the caspases identified, caspase-3 stands out because it is commonly activated by numerous dea th signals and cleaves a variety of important cellular proteins. Studi es in caspase-3 knock-out mice have shown that this protease is essent ial for brain development. To investigate the requirement for caspase- 3 in apoptosis, we took advantage of the MCF-7 breast carcinoma cell l ine, which we show here has lost caspase-3 owing to a 47-base pair del etion within exon 3 of the CASP-3 gene. This deletion results in the s kipping of exon 3 during pre-mRNA splicing, thereby abrogating transla tion of the CASP-3 mRNA. Although MCF-7 cells were still. sensitive to tumor necrosis factor (TNF)- or staurosporine-induced apoptosis, no D NA fragmentation was observed. In addition, MCF-7 cells undergoing cel l death did not display some of the distinct morphological features ty pical of apoptotic cells such as shrinkage and blebbing. Introduction of the CASP-3 gene into MCF-7 cells resulted in DNA fragmentation and cellular blebbing following TNF treatment. These results indicate that although caspase-3 is not essential for TNF- or staurosporine-induced apoptosis, it is required for DNA fragmentation and some of the typic al morphological changes of cells undergoing apoptosis.