Execution of Apoptosis Signal-regulating Kinase 1 (ASK1)-induced Apoptosis by the Mitochondria-dependent Caspase Activation

TLDR

ASK1 activates JNK and p38 MAPKs and is a key pathway in cytokine- and stress-induced apoptosis, yet the mechanism by which it executes cell death remains poorly understood. This study examined whether caspases and mitochondria mediate ASK1-induced apoptosis. The data show that caspase inhibition blocks ASK1‑driven cell death, that ASK1ΔN triggers cytochrome c release and caspase‑9/3 activation but not caspase‑8, that caspase‑9, but not caspase‑8, is essential, and that mitochondrial cytochrome c release precedes caspase‑3 activation, indicating that ASK1 executes apoptosis mainly through mitochondria‑dependent caspase activation.

Abstract

ASK1 activates JNK and p38 mitogen-activated protein kinases and constitutes a pivotal signaling pathway in cytokine- and stress-induced apoptosis. However, little is known about the mechanism of how ASK1 executes apoptosis. Here we investigated the roles of caspases and mitochondria in ASK1-induced apoptosis. We found that benzyloxycarbonyl-Val-Ala-Asp-fluoromethyl ketone (zVAD-fmk), a broad-spectrum caspase inhibitor, mostly inhibited ASK1-induced cell death, suggesting that caspases are required for ASK1-induced apoptosis. Overexpression of ASK1ΔN, a constitutively active mutant of ASK1, induced cytochrome <i>c</i> release from mitochondria and activation of caspase-9 and caspase-3 but not of caspase-8-like proteases. Consistently, caspase-8-deficient (Casp8 ^−/−) cells were sensitive to ASK1-induced caspase-3 activation and apoptosis, suggesting that caspase-8 is dispensable for ASK1-induced apoptosis, whereas ASK1 failed to activate caspase-3 in caspase-9-dificient (Casp9 ^−/−) cells. Moreover, mitochondrial cytochrome <i>c</i> release, which was not inhibited by zVAD-fmk, preceded the onset of caspase-3 activation and cell death induced by ASK1. ASK1 thus appears to execute apoptosis mainly by the mitochondria-dependent caspase activation.