Apopain/CPP32 cleaves proteins that are essential for cellular repair: a fundamental principle of apoptotic death.

Abstract

SummaryProteolysis mediated by the interleukin 1[3-converting enzyme (ICE) homologues is an important mechanism of the apoptotic process. The ICE homologue apopain/CPP-32/Yama (subsequently referred to as apopain) cleaves poly(ADP-ribose)polymerase (PAR.P) early during apoptosis. Additional apoptosis-specific protein cleavages have been observed in which the direct involvement of ICE-like proteases has been postulated. These substrates include the 70-kD protein component of the Ul-ribonucleoprotein (U1-70kD), and the catalytic subunit of the DNAdependent protein kinase (DNA-PKcs). The present studies demonstrate that U1-70kD and DNA-PK~ are excellent substrates for apopain, with cleavage occurring at sites that are highly similar to the cleavage site within PAR.P. The fragments generated from isolated protein substrates by apopain are identical to those observed in intact apoptotic cells, in apoptotic cell extracts, and in normal cell extracts to which apopain has been added. Like PARP, cleavage of these substrates in apoptotic cell extracts is abolished by nanomolar concentrations of Ac-DEVD-CHO and micromolar amounts of Ac-YVAD-CHO, confirming the involvement of apopain or an apopain-like activity. We propose that a central function of apopain or similar homologues in apoptosis is the cleavage of nuclear repair proteins, thereby abolishing their critical homeostatic functions.

Proteolytic Cleavage of the Catalytic Subunit of DNA-Dependent Protein Kinase during Poliovirus Infection

“…Cleavage of DNA-PKcs during poliovirus infection is not prevented by caspase inhibitors. It has previously been reported by several groups that the DNA-PK CS autoantigen is cleaved during apoptosis (10,13,35,40,50), and it has also been shown that DNA-PK CS is a substrate for caspase-3 in vitro (10,50). The results presented above strongly suggest that caspase-3 is not involved in the proteolysis of DNA-PK CS during poliovirus infection.…”

“…They and others have reported that a number of autoantigens are in fact caspase substrates and are cleaved during apoptosis (10,13,14,56). Furthermore, many of these autoantigens are translocated to apoptotic bodies, or apoptotic blebs, suggesting a mechanism whereby they could be presented to effector cells in the immune system (12).…”

Channels formed by subnanomolar concentrations of the toxin aerolysin trigger apoptosis of T lymphomas

“…The samples were analysed for classical indicators of apoptosis: DNA fragmentation by the method of Martin et al (1990) and CPP32 protease activity (Casciola-Rosen et al, 1994) using an ApoAlert CPP32 protease assay (Clontech). For¯ow cytometric analysis of apoptosis, cells were ®xed in 70% ethanol, permeabilized with 0.1% Triton X-100, treated with 5 mg ml À1 RNase and incubated at 378C for 15 min before staining with 50 mg ml À1 propidium iodide for 60 min at 48C.…”

Alterations of Extracellular Calcium Elicit Selective Modes of Cell Death and Protease Activation in SH‐SY5Y Human Neuroblastoma Cells

“…We have previously demonstrated the activation of two families of cysteine proteases, caspases and calpains, in SH-SY5Y cells undergoing staurosporineinduced apoptosis Posmantur et al, 1997). Caspase-3 has been studied as a putative mediator of mammalian apoptosis (Fernandes-Alnemri et al, 1994;Casciola-Rosen et al, 1996;Posmantur et al, Alterations in intracellular calcium (Ca2+,) homeostasis have been implicated in the onset of cell death in many cell types. However, the mode of cell death and direction of change in Ca2fi differ among cell types McConkey and Orrenius, 1997).…”