Activation of Caspase-2 in Apoptosis

Abstract

Members of the CED-3/interleukin-1␤-converting enzyme (ICE) protease (caspase) family are synthesized as proforms, which are proteolytically cleaved and activated during apoptosis. We report here that caspase-2 (ICH-1/NEDD-2), a member of the ICE family, is activated during apoptosis by another ICE member, a caspase-3 (CPP32)-like protease(s). When cells are induced to undergo apoptosis, endogenous caspase-2 is first cleaved into three fragments of 32-33 kDa and 14 kDa, which are then further processed into 18-and 12-kDa active subunits. Up to 50 M N-acetyl-Asp-Glu-ValAsp-aldehyde (DEVD-CHO), a caspase-3-preferred peptide inhibitor, inhibits caspase-2 activation and DNA fragmentation in vivo, but does not prevent loss of mitochondrial function, while higher concentrations of DEVD-CHO (>50 M) inhibit both. In comparison, although the activity of caspase-3 is very sensitive to the inhibition of DEVD-CHO (<50 nM), inhibition of caspase-3 activation as marked by processing of the proform requires more than 100 M DEVD-CHO. Our results suggest that the first cleavage of caspase-2 is accomplished by a caspase-3-like activity, and other ICE-like proteases less sensitive to DEVD-CHO may be responsible for activation of caspase-3 and loss of mitochondrial function. Interleukin-1␤-converting enzyme (ICE)1 caspase-1 (1, 2) was identified as the first mammalian homolog of the Caenorhabditis elegans cell death gene product CED-3 (3, 4). Subsequently, a growing number of ICE-like cysteine proteases have been isolated and characterized, including caspase-2 (NEDD-2/ICH-1) (5, 6), caspase-3 (CPP32/YAMA/Apopain) (7,8,39), caspase-6 (Mch-2) (9), caspase-4 (TX/Ich-2/ICE rel II) (10 -12), caspase-5 (ICE rel III) (12), caspase-7 (Mch-3/CMH-1/ICE-LAP3) (13-15), caspase-8 (FLICE/MACH/Mch-5) (16 -18), caspase-10 (Mch-4) (18), and caspase-9 (ICE-LAP6/Mch-6) (19,20). Increasing evidence suggests that caspases play critical roles in the control of programmed cell death (for review, see Refs. 21-23). Microinjection of an expression vector encoding CrmA, a serpin encoded by cowpox virus, inhibits the death of dorsal root ganglia neurons induced by nerve growth factor deprivation (24). Viral inhibitors of caspases, p35 and CrmA, inhibit serum withdrawal-, tumor necrosis factor-, and Fas-induced apoptosis, as well as cytotoxic T lymphocyte (CTL)-mediated apoptosis (6,(25)(26)(27)(28)(29). Ice Ϫ/Ϫ thymocytes undergo apoptosis normally when treated with dexamethasome and ␥-irradiation but are partially resistant to Fas-induced apoptosis (30). Peptide inhibitors of caspases prevent programmed cell death when administered to tissue culture cells and animals (31). These results indicate that the ICE family plays important roles in mammalian apoptosis. The roles played by individual members of the caspase family in controlling apoptosis are the subjects of intensive debates and investigations.Nedd-2, the murine caspase-2, was identified by Kumar et al. (32) as a mRNA expressed mostly during early embryonic brain development and down-regulated in ad...

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