Explore the vast range of titles available.

When protein folding in the endoplasmic reticulum (ER) is disrupted by alterations in homeostasis in the ER lumen, eucaryotic cells activate a series of signal transduction cascades that are collectively termed the unfolded protein response (UPR). Here we summarize our current understanding of how the UPR functions upon acute and severe stress. We discuss the mechanism of UPR receptor activation, UPR signal transduction to translational and transcriptional responses, UPR termination, and UPR signals that activate upon irreversible damage. Further, we review recent studies that have revealed that UPR provides a wide spectrum of physiological roles. Each individual UPR subpathway provides a unique and specialized role in diverse developmental and metabolic processes. This is especially observed for professional secretory cells, such as plasma cells, pancreatic β cells, hepatocytes, and osteoblasts, where high-level secretory protein synthesis requires a highly evolved mechanism to properly fold, process, and secrete proteins. There is a growing body of data that suggest that different subpathways of the UPR are required throughout the entire life of eucaryotic organisms, from regulation of differentiation to induction of apoptosis.

Expanded polyglutamine 72 repeat (polyQ72) aggregates induce endoplasmic reticulum (ER) stress-mediated cell death with caspase-12 activation and vesicular formation (autophagy). We examined this relationship and the molecular mechanism of autophagy formation. Rapamycin, a stimulator of autophagy, inhibited the polyQ72-induced cell death with caspase-12 activation. PolyQ72, but not polyQ11, stimulated Atg5-Atg12-Atg16 complex-dependent microtubule-associated protein 1 (MAP1) light chain 3 (LC3) conversion from LC3-I to -II, which plays a key role in autophagy. The eucaryotic translation initiation factor 2 α (eIF2 α ) A/A mutation, a knock-in to replace a phosphorylatable Ser 51 with Ala 51 , and dominant-negative PERK inhibited polyQ72-induced LC3 conversion. PolyQ72 as well as ER stress stimulators upregulated Atg12 mRNA and proteins via eIF2 α phosphorylation. Furthermore, Atg5 deficiency as well as the eIF2 α A/A mutation increased the number of cells showing polyQ72 aggregates and polyQ72-induced caspase-12 activation. Thus, autophagy formation is a cellular defense mechanism against polyQ72-induced ER-stress-mediated cell death by degrading polyQ72 aggregates, with PERK/eIF2 α phosphorylation being involved in polyQ72-induced LC3 conversion.

In the endoplasmic reticulum (ER), secretory and transmembrane proteins fold into their native conformation and undergo posttranslational modifications important for their activity and structure. When protein folding in the ER is inhibited, signal transduction pathways, which increase the biosynthetic capacity and decrease the biosynthetic burden of the ER to maintain the homeostasis of this organelle, are activated. These pathways are called the unfolded protein response (UPR). In this review, we briefly summarize principles of protein folding and molecular chaperone function important for a mechanistic understanding of UPR-signaling events. We then discuss mechanisms of signal transduction employed by the UPR in mammals and our current understanding of the remodeling of cellular processes by the UPR. Finally, we summarize data that demonstrate that UPR signaling feeds into decision making in other processes previously thought to be unrelated to ER function, e.g., eukaryotic starvation responses and differentiation programs.In the endoplasmic reticulum (ER), secretory and transmembrane proteins fold into their native conformation and undergo posttranslational modifications important for their activity and structure. When protein folding in the ER is inhibited, signal transduction pathways, which increase the biosynthetic capacity and decrease the biosynthetic burden of the ER to maintain the homeostasis of this organelle, are activated. These pathways are called the unfolded protein response (UPR). In this review, we briefly summarize principles of protein folding and molecular chaperone function important for a mechanistic understanding of UPR-signaling events. We then discuss mechanisms of signal transduction employed by the UPR in mammals and our current understanding of the remodeling of cellular processes by the UPR. Finally, we summarize data that demonstrate that UPR signaling feeds into decision making in other processes previously thought to be unrelated to ER function, e.g., eukaryotic starvation responses and differentiation programs.

Aims/hypothesis Although obesity is associated with endoplasmic reticulum (ER) stress and activation of the unfolded protein response (UPR) in adipose tissue, it is not known how UPR signalling affects adipogenesis. To test whether signalling through protein kinase RNA-like ER kinase/eukaryotic initiation factor 2 alpha (PERK/eIF2α) or inositol-requiring enzyme 1 alpha/X-box binding protein 1 (IRE1α/XBP1) is required for adipogenesis, we studied the role of UPR signalling in adipocyte differentiation in vitro and in vivo in mice. Methods The role of UPR signalling in adipogenesis was investigated using 3T3-L1 cells and primary mouse embryonic fibroblasts (MEFs) by activation or inhibition of PERK-mediated phosphorylation of the eIF2α- and IRE1α-mediated splicing of Xbp1 mRNA. Body weight change, fat mass composition and adipocyte number and size were measured in wild-type and genetically engineered mice fed a control or high-fat diet (HFD). Results ER stress repressed adipocyte differentiation in 3T3-L1 cells. Impaired eIF2α phosphorylation enhanced adipocyte differentiation in MEFs, as well as in mice. In contrast, increased eIF2α phosphorylation reduced adipocyte differentiation in 3T3-L1 cells. Forced production of CCAAT/enhancer binding protein (C/EBP) homologous protein (CHOP), a downstream target of eIF2α phosphorylation, inhibited adipogenesis in 3T3-L1 cells. Mice with deletion of Chop (also known as Ddit3 ) ( Chop −/− ) gained more fat mass than wild-type mice on HFD. In addition, Chop deletion in genetically obese Lepr db/db mice increased body fat mass without altering adipocyte size. In contrast to the eIF2α–CHOP pathway, activation or deletion of Ire1a (also known as Ern1 ) did not alter adipocyte differentiation in 3T3-L1 cells. Conclusions/interpretation These results demonstrate that eIF2α–CHOP suppresses adipogenesis and limits expansion of fat mass in vivo in mice, rendering this pathway a potential therapeutic target.

Type 2 diabetes is a disorder of hyperglycemia resulting from failure of beta cells to produce adequate insulin to accommodate an increased metabolic demand. Here we show that regulation of mRNA translation through phosphorylation of eukaryotic initiation factor 2 (eIF2α) is essential to preserve the integrity of the endoplasmic reticulum (ER) and to increase insulin production to meet the demand imposed by a high-fat diet. Accumulation of unfolded proteins in the ER activates phosphorylation of eIF2α at Ser51 and inhibits translation. To elucidate the role of this pathway in beta-cell function we studied glucose homeostasis in Eif2s1 tm1Rjk mutant mice, which have an alanine substitution at Ser51. Heterozygous ( Eif2s1 +/tm1Rjk ) mice became obese and diabetic on a high-fat diet. Profound glucose intolerance resulted from reduced insulin secretion accompanied by abnormal distension of the ER lumen, defective trafficking of proinsulin, and a reduced number of insulin granules in beta cells. We propose that translational control couples insulin synthesis with folding capacity to maintain ER integrity and that this signal is essential to prevent diet-induced type 2 diabetes.

Macrophages are pivotal constituents of the innate immune system, vital for recognition and elimination of microbial pathogens 1 . Macrophages use Toll-like receptors (TLRs) to detect pathogen-associated molecular patterns—including bacterial cell wall components, such as lipopolysaccharide or lipoteichoic acid, and viral nucleic acids, such as double-stranded (ds)RNA—and in turn activate effector functions, including anti-apoptotic signalling pathways 2 . Certain pathogens, however, such as Salmonella spp., Shigellae spp. and Yersiniae spp., use specialized virulence factors to overcome these protective responses and induce macrophage apoptosis 3 . We found that the anthrax bacterium, Bacillus anthracis , selectively induces apoptosis of activated macrophages 4 through its lethal toxin, which prevents activation of the anti-apoptotic p38 mitogen-activated protein kinase 4 . We now demonstrate that macrophage apoptosis by three different bacterial pathogens depends on activation of TLR4. Dissection of anti- and pro-apoptotic signalling events triggered by TLR4 identified the dsRNA responsive protein kinase PKR as a critical mediator of pathogen-induced macrophage apoptosis. The pro-apoptotic actions of PKR are mediated both through inhibition of protein synthesis and activation of interferon response factor 3.

Transient phosphorylation of the α‐subunit of translation initiation factor 2 (eIF2α) represses translation and activates select gene expression under diverse stressful conditions. Defects in the eIF2α phosphorylation‐dependent integrated stress response impair resistance to accumulation of malfolded proteins in the endoplasmic reticulum (ER stress), to oxidative stress and to nutrient deprivations. To study the hypothesized protective role of eIF2α phosphorylation in isolation of parallel stress signaling pathways, we fused the kinase domain of pancreatic endoplasmic reticulum kinase (PERK), an ER stress‐inducible eIF2α kinase that is normally activated by dimerization, to a protein module that binds a small dimerizer molecule. The activity of this artificial eIF2α kinase, Fv2E‐PERK, is subordinate to the dimerizer and is uncoupled from upstream stress signaling. Fv2E‐PERK activation enhanced the expression of numerous stress‐induced genes and protected cells from the lethal effects of oxidants, peroxynitrite donors and ER stress. Our findings indicate that eIF2α phosphorylation can initiate signaling in a cytoprotective gene expression pathway independently of other parallel stress‐induced signals and that activation of this pathway can single‐handedly promote a stress‐resistant preconditioned state.

The expression of cloned genes in mammalian cells is a basic tool for understanding gene expression, protein structure, and function, and biological regulatory mechanisms. The level of protein expression from heterologous genes introduced into mammalian cells depends upon multiple factors including DNA copy number, efficiency of transportation, mRNA processing, mRNA transport, mRNA stability, and translational efficiency, and protein processing, transport, and stability. Different genes exhibit different rate limiting steps for efficient expression. Multiple strategies are available to obtain high level expression in mammalian cells. This article reviews vector design for expression of foreign genes in mammalian cells.

The eIF2α kinases are a family of evolutionarily conserved serine/threonine kinases that regulate stress-induced translational arrest. Here, we demonstrate that the yeast eIF2α kinase, GCN2 , the target phosphorylation site of Gcn2p, Ser-51 of eIF2α, and the eIF2α-regulated transcriptional transactivator, GCN4 , are essential for another fundamental stress response, starvation-induced autophagy. The mammalian IFN-inducible eIF2α kinase, PKR, rescues starvation-induced autophagy in GCN2 -disrupted yeast, and pkr null and Ser-51 nonphosphorylatable mutant eIF2α murine embryonic fibroblasts are defective in autophagy triggered by herpes simplex virus infection. Furthermore, PKR and eIF2α Ser-51-dependent autophagy is antagonized by the herpes simplex virus neurovirulence protein, ICP34.5. Thus, autophagy is a novel evolutionarily conserved function of the eIF2α kinase pathway that is targeted by viral virulence gene products.