Cleavage of DFF-45/ICAD by Multiple Caspases Is Essential for Its Function during Apoptosis*
- From the Department of Tumor Cell Biology, St. Jude Children’s Research Hospital, Memphis, Tennessee 38101
Abstract
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.
Footnotes
-
↵* This research was supported by National Institutes of Health Grant GM 44088 (to V. J. K.), by National Institutes of Health Cancer Center Core Grant CA 21765 (awarded to SJCRH), and by support from the American Lebanese Syrian Associated Charities.The costs of publication of this article were defrayed in part by the payment of page charges. The article must therefore be hereby marked “advertisement” in accordance with 18 U.S.C. Section 1734 solely to indicate this fact.
-
↵‡ To whom correspondence should be addressed: Dept. of Tumor Cell Biology, St. Jude Children’s Research Hospital, 332 N. Lauderdale, Memphis, TN 38101. Tel.: 901-495-3597; Fax: 901-495-2381; E-mail:vincent.kidd{at}stjude.org.
- Abbreviations:
- DFF-45
-
DNA fragmentation factor 45-kDa subunit
- TNF
-
tumor necrosis factor
- CHX
-
cycloheximide
- STS
-
stuarosporine
- PBS
-
phosphate-buffered saline.
-
- Received August 17, 1998.
- The American Society for Biochemistry and Molecular Biology, Inc.
