Phosphorylation of Eukaryotic Translation Initiation Factor 2 Mediates Apoptosis in Response to Activation of the Double-stranded RNA-dependent Protein Kinase

The interferon-inducible, double-stranded (ds) RNA-dependent serine/threonine protein kinase (PKR) plays a role in viral pathogenesis, cell growth, and differentiation and is implicated as a tumor suppressor gene. Expression of atrans-dominant negative, catalytically inactive mutant PKR protected NIH3T3 cells from apoptosis in response to either treatment with tumor necrosis factor α (TNFα), serum deprivation. In cells expressing mutant PKR, TNFα, but not dsRNA induced transcription from a nuclear factor κ B-dependent promoter, demonstrating specificity for dsRNA in signaling through the PKR pathway. Serum or platelet-derived growth factor addition to serum-deprived mutant PKR-expressing cells induced transcription of the early response genes c-fos and c-jun, indicating that the immediate early response signaling was intact. Overexpression of wild-type PKR in a transient DNA transfection system was sufficient to induce apoptosis. TNFα-induced apoptosis correlated with increased phosphorylation of the α subunit of eukaryotic translation initiation factor 2 (eIF-2α), the primary physiological substrate of the PKR. Furthermore, forced expression of a nonphosphorylatable S51A mutant eIF-2α partially protected cells from TNFα-induced apoptosis, and expression of a S51D mutant eIF-2α, a mutant that mimics phosphorylated eIF-2α, was sufficient to induce apoptosis. Taken together, these studies identify a novel requirement for PKR in stress-induced apoptosis that is mediated through eIF-2α phosphorylation.