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Exposure of cells to various stresses such as hypoxia, hypoglycemia, and pH changes leads to the accumulation of unfolded proteins within the lumen of the endoplasmic reticulum (ER) and subsequent activation of the Unfolded Protein Response (UPR) pathway. One element of the UPR, activation of the PKR-like ER kinase (PERK), leads to phosphorylation of the eukaryotic initiation factor 2α (eIF2α), resulting in attenuation of mRNA translation, and cell cycle arrest in G1 phase. Inhibition of mRNA translation is thought to reduce the load of unfolded proteins in the ER, thereby contributing to cell adaptations. In addition, we have previously demonstrated that translation inhibition contributes to cell cycle arrest, allowing the cell to divert resources to the restoration of ER homeostasis that would normally be used for division. This cell cycle arrest is dependent on reduction of cellular levels of cyclin D1, as overexpression of cyclin D1 prevents cell cycle arrest due to treatment with the inhibitor of glycosylation, tunicamycin. While UPR-dependent protein translation is thought to rely solely on the PERK kinase, we have found that UPR-induced eIF2α and cell cycle arrest still occur in the absence of functional PERK. Tunicamycin-induced phosphorylation of eIF2α in PERK -/- cells occurs in a delayed fashion when compared to wild-type cells, and this delayed phosphorylation coincides temporally with the delayed reduction of cyclin D1 levels observed in these cells. Our work has revealed that the eIF2α kinase GCN2 acts in lieu of PERK and that elimination of both PERK and GCN2 results in the inability of cells to phosphorylate eIF2α or arrest in G1 during ER stress. Strikingly, although eIF2α phosphorylation, reduction of cyclin D1 levels, and cell cycle arrest are abolished in PERK/GCN2 -/- cells, inhibition of mRNA translation is not. Although eIF2α phosphorylation results in the inhibition of select transcripts such as that of cyclin D1, we have found that the bulk inhibition of mRNA translation is not dependent on eIF2α phosphorylation. In fact, UPR activation results in the accumulation of hypophosphorylated (active) eIF4E binding proteins (4EBPs), inhibitors of the cap-binding protein eukaryotic initiation factor 4E. This reduction of 4EBP phosphorylation coincides temporally with inactivation of the Akt-mTOR pathway, a pathway which when activated results in the phosphorylation of 4EBPs. We are currently studying the role that UPR-induced calcium release from the ER plays in controlling the Akt-mTOR pathway during ER stress. Elucidation of the mechanism by which the stressed ER communicates with the cytosolic GCN2 kinase and the Akt-mTOR pathway remains an important goal, as a full understanding of the manner by which cells respond to stresses of the ER will reveal targets of treatment for UPR-based pathologies.

[Proc Amer Assoc Cancer Res, Volume 47, 2006]