Cleavage of DFF-45/ICAD by multiple caspases is essential for its function during apoptosis.
Apoptosis involves the proteolysis of specific cellular proteins by a group of cysteine proteases known as caspases. Many of these cellular targets are either functionally inactivated (e.g. poly(ADP-ribose) polymerase) or activated (e.g. other caspases, gelsolin) by such processing, thereby facilitating the cell death process. Caspase 3 is involved in the processing of many of these proteins. Recently, however, it was reported that caspase 3 is dispensable for the cleavage of a large number of cellular caspase substrates during apoptosis. Among these substrates is DFF-45/ICAD, a subunit of the heterodimeric DNA fragmentation factor (DFF), otherwise known as caspase-activated DNase (CAD), that mediates genomic DNA degradation during apoptosis. Conversely, others have reported that caspase 3 is essential for the cleavage and activation of DFF-45/ICAD. To resolve this controversy we examined DFF-45/ICAD processing during apoptosis in MCF-7 breast carcinoma cells that lack functional caspase 3 and in MCF-7 cells expressing caspase 3. We found that DFF-45/ICAD is cleaved by two distinct caspases, one of which is caspase 3. Furthermore, cleavage of the carboxyl-terminal region of DFF-45/ICAD, which is necessary for activation of the enzyme, requires functional caspase 3. In the absence of caspase 3 cleavage of the amino-terminal region of DFF-45/ICAD by another caspase occurs, but the DFF-45 enzyme remains inactive.
Tang
D
D
Department of Tumor Cell Biology, St. Jude Children's Research Hospital, Memphis, Tennessee 38101, USA.
Kidd
V J
VJ
D017209:Apoptosis; D051017:Apoptosis Regulatory Proteins; D053148:Caspase 3; D020169:Caspases / Q000378:metabolism*; D003513:Cycloheximide / Q000494:pharmacology; D018260:Gelsolin / Q000378:metabolism; D006801:Humans; D006868:Hydrolysis; D011506:Proteins / Q000378:metabolism*; D019311:Staurosporine / Q000494:pharmacology; D014407:Tumor Cells, Cultured; D014409:Tumor Necrosis Factor-alpha / Q000494:pharmacology
D051017:Apoptosis Regulatory Proteins; D018260:Gelsolin; D011506:Proteins; D014409:Tumor Necrosis Factor-alpha; C109664:caspase-activated DNase inhibitor; D003513:Cycloheximide; C505407:CASP3 protein, human; D053148:Caspase 3; D020169:Caspases; D019311:Staurosporine