Inhibition of PKR protects against tunicamycin-induced apoptosis in neuroblastoma cells

• Inhibition of PKR protects neuronal cells from apoptosis.
• PKR activity is necessary for caspase-3 activation and CHOP induction.
• PKR inhibition by its catalytically inactive mutant is effective in neuronal cells.
• PKR may be a good target for neuroprotective therapeutics.

Endoplasmic reticulum (ER) dysfunction is thought to play a significant role in several neurological disorders, including Alzheimer's disease, Parkinson's disease, Huntington's disease, multiple sclerosis, amyotrophic lateral sclerosis, cerebral ischemia, and the prion diseases. ER dysfunction can be mimicked by cellular stress signals such as disruption of calcium homeostasis, inhibition of protein glycosylation, and reduction of disulfide bonds, which results in accumulation of misfolded proteins in the ER and leads to cell death by apoptosis. Tunicamycin, which is an inhibitor of protein glycosylation, induces ER stress and apoptosis. In this study, we examined the involvement of double stranded (ds) RNA-activated protein kinase PKR in tunicamycin-induced apoptosis. We used overexpression of the trans-dominant negative, catalytically inactive mutant K296R to inhibit PKR activity in neuroblastoma cells. We demonstrate that inhibition of PKR activation in response to tunicamycin protects neuronal cells from undergoing apoptosis. Furthermore, K296R overexpressing cells show defective PKR activation, delayed eIF2α phosphorylation, dramatically delayed ATF4 expression. In addition, both caspase-3 activation and C/EBP homologous protein (CHOP, also known as GADD153) induction, which are markers of apoptotic cells, are absent from K296R overexpression cells in response to tunicamycin. These results establish that PKR activation plays a major regulatory role in induction of apoptosis in response to ER stress and indicates the potential of PKR as possible target for neuroprotective therapeutics.