p53 mediates nontranscriptional cell death in dopaminergic cells in response to proteasome inhibition.

Proteasome dysfunction has been demonstrated in Parkinson disease (PD), and proteasome inhibitors have been shown to induce degeneration of dopaminergic neurons in vitro and in vivo. The mechanism whereby proteasome dysfunction leads to dopaminergic cell death, however, is unknown. In this study, we show that proteasome inhibition in both PC12 cells and dopaminergic neurons derived from embryonic stem cells is associated with mitochondrial membrane permeabilization, activation of caspase-3, and nuclear changes consistent with apoptosis. Prior to the emergence of apoptotic features, we found that proteasome inhibition induced increased levels of phosphorylated p53. Inhibition of p53 by pifithrin-alpha or by RNA interference prevented mitochondrial membrane permeabilization and cytotoxicity. There was no increase in p53 mRNA in proteasome-inhibited cells, suggesting that p53 was increased in a transcription-independent manner. Further, there was no increase in Puma or Bax mRNA and p53 co-immunoprecipitated with Bcl-xL and Mdm2. These findings suggest that p53 mediates cell death by way of a direct mitochondrial effect in this model. We also observed increased levels of phosphorylated p53 in dopamine neurons of the substantia nigra pars compacta of mice following systemic administration of a proteasome inhibitor. These changes preceded degeneration of dopaminergic neurons. Increased phosphorylated p53 was also demonstrated in the substantia nigra pars compacta of post-mortem PD brains. These results suggest that abnormalities in p53 signaling play a role in dopaminergic cell death induced by proteasome inhibition and may be relevant to neurodegeneration in PD.