Synthetic embryonic lethality upon deletion of the ER cochaperone p58 and the ER stress sensor ATF6α

The unfolded protein response (UPR) is activated as a consequence of alterations to ER homeostasis. It upregulates a group of ER chaperones and cochaperones, as well as other genes that improve protein processing within the secretory pathway. The UPR effector ATF6α augments—but is not essential for—maximal induction of ER chaperones during stress, yet its role, if any, in protecting cellular function during normal development and physiology is unknown. A systematic analysis of multiple tissues from Atf6α−/− mice revealed that all tissues examined were grossly insensitive to loss of ATF6α. However, combined deletion of ATF6α and the ER cochaperone p58^IPK resulted in synthetic embryonic lethality. These findings reveal for the first time that an intact UPR can compensate for the genetic impairment of protein folding in the ER in vivo. The also expose a role for p58^IPK in normal embryonic development.     

Endoplasmic reticulum stress in glomerular epithelial cell injury

Focal segmental glomerulosclerosis (FSGS) may be associated with glomerular epithelial cell (GEC; podocyte) apoptosis due to acquired injury or mutations in specific podocyte proteins. This study addresses mediation of GEC injury, focusing on endoplasmic reticulum (ER) stress. We studied signaling in cultured GECs in the presence or absence of the extracellular matrix (ECM). Adhesion to collagen supports cell survival, but adhesion to plastic (loss of contact with ECM) leads to apoptosis. Compared with collagen-adherent cells, GECs on plastic showed increased protein misfolding in the ER, and an adaptive-protective ER stress response, including increased expression of ER chaperones, increased phosphorylation of eukaryotic translation initiation factor-2α (eIF2α), and a reduction in protein synthesis. Activation of these ER stress pathways counteracted apoptosis. However, tunicamycin (a potent stimulator of ER stress) changed the ER stress response from protective to cytotoxic, as tunicamycin induced the proapoptotic ER stress gene, C/EBP homologous protein-10, and exacerbated apoptosis in GECs adherent to plastic, but not collagen. In GECs adherent to plastic, adaptive ER stress was associated with an increase in polyubiquitinated proteins and "choking" of the proteasome. Furthermore, pharmacological inhibition of the proteasome induced ER stress in GECs. Finally, we show that ER stress (induction of ER chaperones and eIF2α phosphorylation) was evident in experimental FSGS in vivo. Thus interactions of GECs with ECM may regulate protein folding and induction of the ER stress response. FSGS is associated with induction of ER stress. Enhancing protective aspects of the ER stress response may reduce apoptosis and possibly glomerulosclerosis.     

Peroxisome deficiency-induced ER stress and SREBP-2 pathway activation in the liver of newborn PEX2 knock-out mice

Disruption of the Pex2 gene leads to peroxisome deficiency and widespread metabolic dysfunction. We previously demonstrated that peroxisomes are critical for maintaining cholesterol homeostasis, using peroxisome-deficient Pex2(-/-) mice on a hybrid Swiss Webster×129S6/SvEv (SW/129) genetic background. Peroxisome deficiency activates hepatic endoplasmic reticulum (ER) stress pathways, leading to dysregulation of the endogenous sterol response mechanism. Herein, we demonstrate a more profound dysregulation of cholesterol homeostasis in newborn Pex2(-/-) mice congenic on a 129S6/SvEv (129) genetic background, and substantial differences between newborn versus postnatal Pex2(-/-) mice in factors that activate ER stress. These differences extend to relationships between activation of genes regulated by SREBP-2 versus PPARα. The SREBP-2 pathway is induced in neonatal Pex2(-/-) livers from 129 and SW/129 strains, despite normal hepatic cholesterol levels. ER stress markers are increased in newborn 129 Pex2(-/-) livers, which occurs in the absence of hepatic steatosis or accumulation of peroxins in the ER. Moreover, the induction of SREBP-2 and ER stress pathways is independent of PPARα activation in livers of newborn 129 and SW/129 Pex2(-/-) mice. Two-week-old wild-type mice treated with the peroxisome proliferator WY-14,643 show strong induction of PPARα-regulated genes and decreased expression of SREBP-2 and its target genes, further demonstrating that SREBP-2 pathway induction is not dependent on PPARα activation. Lastly, there is no activation of either SREBP-2 or ER stress pathways in kidney and lung of newborn Pex2(-/-) mice, suggesting a parallel induction of these pathways in peroxisome-deficient mice. These findings establish novel associations between SREBP-2, ER stress and PPARα pathway inductions.     

IRE1α disruption causes histological abnormality of exocrine tissues, increase of blood glucose level, and decrease of serum immunoglobulin level

Accumulation of unfolded proteins in the endoplasmic reticulum (ER) causes ER stress. As a cellular adaptive response to ER stress, unfolded protein response (UPR) activates molecules for the quality control of ER proteins. One enzyme that plays an important role in UPR is Inositol Requiring Enzyme-1 (IRE1), which is highly conserved from yeast to humans. In particular, mammalian IRE1α activates X-box-binding protein 1 (XBP1) by unconventional splicing of XBP1 mRNA during ER stress. From analysis of knockout mice, both IRE1α and XBP1 have been shown to be essential for development and that XBP1 is necessary for the secretory machinery of exocrine glands, plasma cell differentiation, and hepatic lipogenesis. However, the essentiality of IRE1α in specific organs and tissues remains incompletely understood. Here, we analyzed the phenotype of IRE1α conditional knockout mice and found that IRE1α-deficient mice exhibit mild hypoinsulinemia, hyperglycemia, and a low-weight trend. Moreover, IRE1α disruption causes histological abnormality of the pancreatic acinar and salivary serous tissues and decrease of serum level of immunoglobulin produced in the plasma cells, but not dysfunction of liver. Comparison of this report with previous reports regarding XBP1 conditional knockout mice might provide some clues for the discovery of the novel functions of IRE1α and XBP1.     

Treadmill exercise represses neuronal cell death and inflammation during Aβ-induced ER stress by regulating unfolded protein response in aged presenilin 2 mutant mice

Alzheimer’s disease (AD) is characterized by the deposition of aggregated amyloid-beta (Aβ), which triggers a cellular stress response called the unfolded protein response (UPR). The UPR signaling pathway is a cellular defense system for dealing with the accumulation of misfolded proteins but switches to apoptosis when endoplasmic reticulum (ER) stress is prolonged. ER stress is involved in neurodegenerative diseases including AD, but the molecular mechanisms of neuronal apoptosis and inflammation by Aβ-induced ER stress to exercise training are not fully understood. Here, we demonstrated that treadmill exercise (TE) prevented PS2 mutation-induced memory impairment and reduced Aβ-42 deposition through the inhibition of β-secretase (BACE-1) and its product, C-99 in cortex and/or hippocampus of aged PS2 mutant mice. We also found that TE down-regulated the expression of GRP78/Bip and PDI proteins and inhibited activation of PERK, eIF2α, ATF6α, sXBP1 and JNK-p38 MAPK as well as activation of CHOP, caspase-12 and caspase-3. Moreover, TE up-regulated the expression of Bcl-2 and down-regulated the expressions of Bax in the hippocampus of aged PS2 mutant mice. Finally, the generation of TNFα and IL-1α and the number of TUNEL-positive cells in the hippocampus of aged PS2 mutant mice was also prevented or decreased by TE. These results showed that TE suppressed the activation of UPR signaling pathways as well as inhibited the apoptotic pathways of the UPR and inflammatory response following Aβ-induced ER stress. Thus, therapeutic strategies that modulate Aβ-induced ER stress through TE could represent a promising approach for the prevention or treatment of AD.     

Endoplasmic reticulum (ER) stress in cumulus-oocyte complexes impairs pentraxin-3 secretion, mitochondrial membrane potential (DeltaPsi m), and embryo development

Fatty acids such as palmitic acid at high levels are known to induce endoplasmic reticulum (ER) stress and lipotoxicity in numerous cell types and thereby contribute to cellular dysfunctions in obesity. To understand the impact of high fatty acids on oocytes, ER stress and lipotoxicity were induced in mouse cumulus-oocyte complexes during in vitro maturation using the ER Ca(2+) channel blocker thapsigargin or high physiological levels of palmitic acid; both of which significantly induced ER stress marker genes (Atf4, Atf6, Xbp1s, and Hspa5) and inositol-requiring protein-1α phosphorylation, demonstrating an ER stress response that was reversible with the ER stress inhibitor salubrinal. Assessment of pentraxin-3, an extracellular matrix protein essential for fertilization, by immunocytochemistry and Western blotting showed dramatically impaired secretion concurrent with ER stress. Mitochondrial activity in oocytes was assessed by 5,5',6,6'-tetrachloro-1,1',3,3'-tetraethylbenzimidazolylcarbocyanine iodide staining of inner mitochondrial membrane potential, and oocytes matured in thapsigargin or high-dose palmitic acid had significantly reduced mitochondrial activity, reduced in vitro fertilization rates, and were slower to develop to blastocysts. The deficiencies in protein secretion, mitochondrial activity, and oocyte developmental competence were each normalized by salubrinal, demonstrating that ER stress is a key mechanism mediating fatty acid-induced defects in oocyte developmental potential.     

Modulating stress responses by the UPRosome: a matter of life and death

The accumulation of unfolded proteins in the endoplasmic reticulum (ER) triggers the unfolded protein response (UPR) through the activation of specialized sensors including inositol-requiring enzyme-1α (IRE1α). IRE1α signals by assembling a dynamic protein platform referred to as the UPRosome, where different modulator and adaptor proteins assemble to regulate the kinetics and amplitude of UPR effector responses. Conversely, chronic ER stress can cause apoptosis. Recent evidence indicates that several apoptosis-related proteins interact with IRE1α, regulating its prosurvival activities and performing a dual function in the regulation of cell death and adaptation to stress. Based on the increasing relevance of ER stress to the occurrence of diverse pathological conditions, strategies to target and modulate the assembly and composition of the UPRosome could have therapeutic benefits for disease intervention.