Caspase-3 is required for DNA fragmentation and morphological changes associated with apoptosis.
Interleukin 1beta-converting enzyme-like proteases (caspases) are crucial components of cell death pathways. Among the caspases identified, caspase-3 stands out because it is commonly activated by numerous death signals and cleaves a variety of important cellular proteins. Studies in caspase-3 knock-out mice have shown that this protease is essential for brain development. To investigate the requirement for caspase-3 in apoptosis, we took advantage of the MCF-7 breast carcinoma cell line, which we show here has lost caspase-3 owing to a 47-base pair deletion within exon 3 of the CASP-3 gene. This deletion results in the skipping of exon 3 during pre-mRNA splicing, thereby abrogating translation of the CASP-3 mRNA. Although MCF-7 cells were still sensitive to tumor necrosis factor (TNF)- or staurosporine-induced apoptosis, no DNA fragmentation was observed. In addition, MCF-7 cells undergoing cell death did not display some of the distinct morphological features typical 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 typical morphological changes of cells undergoing apoptosis.
Jänicke
R U
RU
Institute of Molecular and Cell Biology, The National University of Singapore, 30 Medical Drive, Singapore 117609, Republic of Singapore. mcbrj@imcb.nus.edu.sg
Sprengart
M L
ML
Wati
M R
MR
Porter
A G
AG
D000818:Animals; D017209:Apoptosis / Q000187:drug effects / Q000502:physiology*; D001943:Breast Neoplasms; D053148:Caspase 3; D020169:Caspases; D003546:Cysteine Endopeptidases / Q000096:biosynthesis / Q000235:genetics / Q000378:metabolism*; D053938:DNA Fragmentation; D005091:Exons; D005260:Female; D006801:Humans; D051379:Mice; D018345:Mice, Knockout; D009447:Neuroblastoma; D016133:Polymerase Chain Reaction; D014176:Protein Biosynthesis; D012322:RNA Precursors / Q000378:metabolism; D012326:RNA Splicing; D012333:RNA, Messenger / Q000096:biosynthesis; D011994:Recombinant Proteins / Q000096:biosynthesis / Q000378:metabolism; D017384:Sequence Deletion; D019311:Staurosporine / Q000494:pharmacology; D014162:Transfection; D014407:Tumor Cells, Cultured; D014409:Tumor Necrosis Factor-alpha / Q000494:pharmacology
D012322:RNA Precursors; D012333:RNA, Messenger; D011994:Recombinant Proteins; D014409:Tumor Necrosis Factor-alpha; C505407:CASP3 protein, human; C505409:Casp3 protein, mouse; D053148:Caspase 3; D020169:Caspases; D003546:Cysteine Endopeptidases; D019311:Staurosporine