Double-stranded RNA-dependent Protein Kinase Phosphorylation of the α-Subunit of Eukaryotic Translation Initiation Factor 2 Mediates Apoptosis

Abstract

As the molecular processes of complex cell stress signaling pathways are defined, the subsequent challenge is to elucidate how each individual event influences the final biological outcome. Phosphorylation of the translation initiation factor 2 (eIF2␣) at Ser 51 is a molecular signal that inhibits translation in response to activation of any of four diverse eIF2␣ stress kinases. We used gene targeting to replace the wild-type Ser 51 allele with an Ala in the eIF2␣ gene to test the hypothesis that translational control through eIF2␣ phosphorylation is a central death stimulus in eukaryotic cells. Homozygous eIF2␣ mutant mouse embryo fibroblasts were resistant to the apoptotic effects of dsRNA, tumor necrosis factor-␣, and serum deprivation. TNF␣ treatment induced eIF2␣ phosphorylation and activation of caspase 3 primarily through the dsRNA-activated eIF2␣ kinase PKR. In addition, expression of a phospho-mimetic Ser 51 to Asp mutant eIF2␣-activated caspase 3, indicating that eIF2␣ phosphorylation is sufficient to induce apoptosis. The proapoptotic effects of PKR-mediated eIF2␣ phosphorylation contrast with the anti-apoptotic response upon activation of the PKR-related endoplasmic reticulum eIF2␣ kinase, PERK. Therefore, divergent fates of death and survival can be mediated through phosphorylation at the same site within eIF2␣. We propose that eIF2␣ phosphorylation is fundamentally a death signal, yet it may promote either death or survival, depending upon coincident signaling events.

Roles of the translationally controlled tumour protein (TCTP) and the double-stranded RNA-dependent protein kinase, PKR, in cellular stress responses

“…Ca þ þ -stress-mediated downregulation of TCTP is dependent on PKR (Figure 1) and also on phosphorylation of eIF2a (Figures 2b and c). The latter result is in line with the demonstration that eIF2a phosphorylation is required for apoptosis induction (Scheuner et al, 2006). Consistent with the results obtained using the PKR À/À cells, the data from experiments with the eIF2a S51A-knock-in cells (Figure 2c) show that TCTP levels are substantially downregulated under Ca þ þ -stress conditions, but only to a much lesser extent by tunicamycin.…”

“…Interestingly, Mcl-1 has been described as a protein stabilizing TCTP (Zhang et al, 2002) and vice versa (Liu et al, 2005). These results are compatible with a model according to which activation of PKR by proapoptotic stimuli results in an increase in eIF2a phosphorylation and downregulation of antiapoptotic proteins (Scheuner et al, 2006), such as TCTP (Figure 7) or Mcl-1 (Fritsch et al, 2007). It will be important to investigate which other antiapoptotic proteins may be regulated in this way.…”

“…Phosphorylation of eIF2a has been shown to be important for PKR-dependent apoptosis (Garcia et al, 2006;Scheuner et al, 2006). However, since PKR is also involved in other apoptosis-regulating pathways, it was of interest to establish whether eIF2a phosphorylation is indeed essential for downregulation of TCTP under Ca þ þ stress.…”

“…A major role for PKR has recently emerged in the control of apoptosis, following activation of the tumour-suppressor p53 pathway (Yoon et al, 2009). Inhibition of protein synthesis through eIF2a phosphorylation plays a considerable part in PKR activity (Scheuner et al, 2006;Lee et al, 2007), but other pathways are most likely also involved (Garcia et al, 2006).…”

Ppp1r15 gene knockout reveals an essential role for translation initiation factor 2 alpha (eIF2α) dephosphorylation in mammalian development

“…Although an important adaptation to a variety of stressful conditions, sustained elevation of eIF2(␣P) is poorly tolerated (1,15,16). However, the role of dephosphorylation in protecting against the consequences of deregulated elevation in eIF2(␣P) has not been studied.…”