An increasing body of evidence points to a key role of endoplasmic reticulum (ER) stress in acute and chronic neurodegenerative conditions. Extensive ER stress can trigger neuronal apoptosis, but the signaling pathways that regulate this cell death remain unclear. In the present study, we demonstrate that PUMA, a Bcl-2 homology 3 (BH3)-only member of the Bcl-2 family, is transcriptionally activated in cortical neurons by ER stress and is essential for ER-stress-induced cell death. PUMA is known to be a key transcriptional target of p53, but we have found that ER stress triggers PUMA induction and cell death through a p53-independent mechanism mediated by the ER-stress-inducible transcription factor ATF4 (activating transcription factor 4). Specifically, we demonstrate that ectopic expression of ATF4 sensitizes mouse cortical neurons to ER-stress-induced apoptosis and that ATF4-deficient neurons exhibit markedly reduced levels of PUMA expression and cell death. However, chromatin immunoprecipitation experiments suggest that ATF4 does not directly regulate the PUMA promoter. Rather, we found that ATF4 induces expression of the transcription factor CHOP (C/EBP homologous protein) and that CHOP in turn activates PUMA induction. Specifically, we demonstrate that CHOP binds to the PUMA promoter during ER stress and that CHOP knockdown attenuates PUMA induction and neuronal apoptosis. In summary, we have identified a key signaling pathway in ER-stress-induced neuronal death involving ATF4 -CHOP-mediated transactivation of the proapoptotic Bcl-2 family member PUMA. We propose that this pathway may be an important therapeutic target relevant to a number of neurodegenerative conditions.

Loss of parkin function is responsible for the majority of autosomal recessive parkinsonism. Here, we show that parkin is not only a stress-protective, but also a stress-inducible protein. Both mitochondrial and endoplasmic reticulum (ER) stress induce an increase in parkin-specific mRNA and protein levels. The stress-induced upregulation of parkin is mediated by ATF4, a transcription factor of the unfolded protein response (UPR) that binds to a specific CREB/ATF site within the parkin promoter. Interestingly, c-Jun can bind to the same site, but acts as a transcriptional repressor of parkin gene expression. We also present evidence that mitochondrial damage can induce ER stress, leading to the activation of the UPR, and thereby to an upregulation of parkin expression. Vice versa, ER stress results in mitochondrial damage, which can be prevented by parkin. Notably, the activity of parkin to protect cells from stress-induced cell death is independent of the proteasome, indicating that proteasomal degradation of parkin substrates cannot explain the cytoprotective activity of parkin. Our study supports the notion that parkin has a role in the interorganellar crosstalk between the ER and mitochondria to promote cell survival under stress, suggesting that both ER and mitochondrial stress can contribute to the pathogenesis of Parkinson's disease.

We previously reported that delayed administration of the general cyclin‐dependent kinase inhibitor flavopiridol following global ischemia provided transient neuroprotection and improved behavioral performance. However, it failed to provide longer term protection. In the present study, we investigate the ability of delayed flavopiridol in combination with delayed minocycline, another neuroprotectant to provide sustained protection following global ischemia. We report that a delayed combinatorial treatment of flavopiridol and minocycline provides synergistic protection both 2 and 10 weeks following ischemia. However, protected neurons in the hippocampal CA1 are synaptically impaired as assessed by electrophysio logical field potential recordings. This is likely because of the presence of degenerated processes in the CA1 even with combinatorial therapy. This indicates that while we have addressed one important pre‐clinical parameter by dramatically improving long‐term neuronal survival with delayed combinatorial therapy, the issue of synaptic preservation of protected neurons still exists. These results also highlight the important observation that protection does not always lead to proper function.

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