Protein misfolding in the endoplasmic reticulum (ER) leads to cell death through PERK-mediated phosphorylation of eIF2α, although the mechanism is not understood. ChIP-seq and mRNA-seq of activating transcription factor 4 (ATF4) and C/EBP homologous protein (CHOP), key transcription factors downstream of p-eIF2α, demonstrated that they interact to directly induce genes encoding protein synthesis and the unfolded protein response, but not apoptosis. Forced expression of ATF4 and CHOP increased protein synthesis and caused ATP depletion, oxidative stress and cell death. The increased protein synthesis and oxidative stress were necessary signals for cell death. We show that eIF2α-phosphorylation-attenuated protein synthesis, and not Atf4 mRNA translation, promotes cell survival. These results show that transcriptional induction through ATF4 and CHOP increases protein synthesis leading to oxidative stress and cell death. The findings suggest that limiting protein synthesis will be therapeutic for diseases caused by protein misfolding in the ER.

SUMMARY Integrated Stress Response is a homeostatic mechanism induced by endoplasmic reticulum (ER) stress. In acute/transient ER stress, decreased global protein synthesis and increased uORF mRNA translation are followed by normalization of protein synthesis. Here, we report a dramatically different response during chronic ER stress. This chronic ISR program is characterized by persistently elevated uORF mRNA translation and concurrent gene expression reprogramming, which permits simultaneous stress sensing and proteostasis. The program includes PERK-dependent switching to an eIF3-dependent translation initiation mechanism resulting in partial but not complete translational recovery, which, together with transcriptional reprogramming, selectively bolsters expression of proteins with ER functions. Coordination of transcriptional and translational reprogramming prevents ER dysfunction and inhibits “foamy cell” development, thus establishing a molecular basis for understanding human diseases associated with ER dysfunction.

The tumor suppressor p53, a sensor of multiple forms of cellular stress, is regulated by post-translational mechanisms to induce cell-cycle arrest, senescence, or apoptosis. We demonstrate that endoplasmic reticulum (ER) stress inhibits p53-mediated apoptosis. The mechanism of inhibition involves the increased cytoplasmic localization of p53 due to phosphorylation at serine 315 and serine 376, which is mediated by glycogen synthase kinase-3 ␤ (GSK-3␤). ER stress induces GSK-3␤ binding to p53 in the nucleus and enhances the cytoplasmic localization of the tumor suppressor. Inhibition of apoptosis caused by ER stress requires GSK-3␤ and does not occur in cells expressing p53 with mutation(s) of serine 315 and/or serine 376 to alanine(s). As a result of the increased cytoplasmic localization, ER stress prevents p53 stabilization and p53-mediated apoptosis upon DNA damage. It is concluded that inactivation of p53 is a protective mechanism utilized by cells to adapt to ER stress.[Keywords: Endoplasmic reticulum stress; p53; glycogen synthase kinase-3␤; protein phosphorylation; protein localization; apoptosis] Supplemental material is available at http://www.genesdev.org.

Although studies on viral gene expression were essential for the discovery of internal ribosome entry sites (IRESs), it is becoming increasingly clear that IRES activities are present in a significant number of cellular mRNAs. Remarkably, many of these IRES elements initiate translation of mRNAs encoding proteins that protect cells from stress (when the translation of the vast majority of cellular mRNAs is significantly impaired). The purpose of this review is to summarize the progress on the discovery and function of cellular IRESs. Recent findings on the structures of these IRESs and specifically regulation of their activity during nutritional stress, differentiation, and mitosis will be discussed.Initiation of protein synthesis in eukaryotes is a complex process requiring numerous accessory proteins called initiation factors (also termed canonical initiation factors) (1). Assembly of the 80 S ribosome at a start codon within the majority of eukaryotic mRNAs involves binding of the mRNA 5Ј-m 7 G cap structure to a group of proteins referred to as the cap-binding complex or eIF4F (which consists of three proteins: eIF4E, 1 eIF4G, and eIF4A) (1-3). This is followed by recruitment of the 40 S ribosomal subunit and associated initiation factors (43 S initiation complex comprising a 40 S subunit, eIF2⅐GTP⅐Met-tRNA i , and eIF3) and movement of the 43 S initiation complex along the 5Ј-untranslated region (5Ј-UTR) in search of the initiation codon (1-3) (Fig. 1, left panel). This mechanism of translation initiation is known as "ribosome scanning" (1-3). Initiation factor eIF4G functions as a scaffolding protein. It binds eIF4E (a capbinding protein) and eIF4A (an ATP-dependent RNA helicase, which is thought to unwind the secondary structure in the mRNA 5Ј-UTR) and bridges the mRNA and the ribosome via its interaction with the 40 S bound initiation factor eIF3 (1-3). eIF3 is a multiprotein complex directly associated with the small ribosomal subunit and was shown to impede the association of the 40 S and 60 S ribosomal subunits in the absence of eIF2⅐GTP⅐Met-tRNA i ternary complex (1-3). eIF2 binds GTP and Met-tRNA i and transfers Met-tRNA i to the 40 S ribosomal subunit (1-4). It should be noted that the availability of eIF4E for binding to eIF4G is regulated in eukaryotic cells by the phosphorylation of a small family of eIF4E-binding proteins (the 4E-BPs). Furthermore, proteins that bind the poly(A) tails of the mRNA (PABPs) were shown to facilitate initiation and recycling of ribosomes through interaction with eIF4G (1-3). However, it has become clear that some viral and eukaryotic cellular mRNAs can be translated via internal initiation, a process that involves direct binding of the ribosome to specific mRNA regions termed internal ribosome entry sites (IRESs). This translation initiation mechanism is generally independent of recognition of the 5Ј-mRNA end and involves direct recruitment of the 40 S ribosomes to the vicinity of the initiation codon (5-10) (Fig. 1, right panel). Although the debate continues over ...