Proteasome inhibitors induce cytochrome c-caspase-3-like protease-mediatedapoptosis in cultured cortical neurons

The ubiquitin-proteasome protein degradation pathway is crucial in controll ing intracellular levels of a variety of short-lived proteins and maintaini ng cellular growth and metabolism. In a previous study, we showed the accum ulation of conjugated ubiquitin in CA1 neurons of the gerbil after 5 min of forebrain ischemia (Morimoto et al., 1996; Ide et al., 1999). The accumula tion of conjugated ubiquitin may reflect proteasome malfunction. In the pre sent study, we investigated the effects of proteasome inhibitors on primary neuronal cultures to determine whether proteasomal malfunction induces neu ronal death. When carbobenzoxy-Leu-Leu-Leu-aldehyde or lactacystin, two dif ferent types of proteasome inhibitors, were separately used to suppress pro teasome activity, we observed induction of apoptotic neuronal cell death in both cases. During the apoptotic process, mitochondrial membrane potential was disrupted, cytochrome-c was released from mitochondria into the cytoso l, and caspase-3-like proteases were activated. Apoptosis was inhibited by pretreatment with acetyl-aspartyl-glutamylvalyl-aspart-1-aldehyde or overex pression of Bcl-x/L. These results demonstrated that suppression of proteas ome function induces neuronal apoptosis via the release of cytochrome c fro m mitochondria and activation of caspase-3-like proteases.