Role of the unfolded protein response in cell death

Unfolded protein response (UPR) is an important genomic response to endoplasmic reticulum (ER) stress. The ER chaperones, GRP78 and Gadd153, play critical roles in cell survival or cell death as part of the UPR, which is regulated by three signaling pathways: PERK/ATF4, IRE1/XBP1 and ATF6. During the UPR, accumulated unfolded protein is either correctly refolded, or unsuccessfully refolded and degraded by the ubiquitin-proteasome pathway. When the unfolded protein exceeds a threshold, damaged cells are committed to cell death, which is mediated by ATF4 and ATF6, as well as activation of the JNK/AP-1/Gadd153-signaling pathway. Gadd153 suppresses activation of Bcl-2 and NF-κB. UPR-mediated cell survival or cell death is regulated by the balance of GRP78 and Gadd153 expression, which is coregulated by NF-κB in accordance with the magnitude of ER stress. Less susceptibility to cell death upon activation of the UPR may contribute to tumor progression and drug resistance of solid tumors.     

The unfolded protein response to endoplasmic reticulum stress in cultured astrocytes and rat brain during experimental diabetes

Oxidative-nitrosative stress and inflammatory responses are associated with endoplasmic reticulum (ER) stress in diabetic retinopathy, raising the possibility that disturbances in ER protein processing may contribute to CNS dysfunction in diabetics. Upregulation of the unfolded protein response (UPR) is a homeostatic response to accumulation of abnormal proteins in the ER, and the present study tested the hypothesis that the UPR is upregulated in two models for diabetes, cultured astrocytes grown in 25 mmol/L glucose for up to 4 weeks and brain of streptozotocin (STZ)-treated rats with diabetes for 1–7 months. Markers associated with translational blockade (phospho-eIF2α and apoptosis (CHOP), inflammatory response (inducible nitric oxide synthase, iNOS), and nitrosative stress (nuclear translocation of glyceraldehyde-3-phosphate dehydrogenase, GAPDH) were not detected in either model. Nrf2 was present in nuclei of low- and high-glucose cultures, consistent with oxidative stress. Astrocytic ATF4 expression was not altered by culture glucose concentration, whereas phospho-IRE and ATF6 levels were higher in low- compared with high-glucose cultures. The glucose-regulated chaperones, GRP78 and GRP94, were also expressed at higher levels in low- than high-glucose cultures, probably due to recurrent glucose depletion between feeding cycles. In STZ-rat cerebral cortex, ATF4 level was transiently reduced at 4 months, and p-IRE levels were transiently elevated at 3 months. However, GRP78 and GRP94 expression was not upregulated, and iNOS, amyloid-β, and nuclear accumulation of GAPDH were not evident in STZ-diabetic brain. High-glucose cultured astrocytes and STZ-diabetic brain are relatively resistant to diabetes-induced ER stress, in sharp contrast with cultured retinal Müller cells and diabetic rodent retina.     

HCV causes chronic endoplasmic reticulum stress leading to adaptation and interference with the unfolded protein response

Background

The endoplasmic reticulum (ER) is the cellular site for protein folding. ER stress occurs when protein folding capacity is exceeded. This stress induces a cyto-protective signaling cascades termed the unfolded protein response (UPR) aimed at restoring homeostasis. While acute ER stress is lethal, chronic sub-lethal ER stress causes cells to adapt by attenuation of UPR activation. Hepatitis C virus (HCV), a major human pathogen, was shown to cause ER stress, however it is unclear whether HCV induces chronic ER stress, and if so whether adaptation mechanisms are initiated. We wanted to characterize the kinetics of HCV-induced ER stress during infection and assess adaptation mechanisms and their significance.

Methods and Findings

The HuH7.5.1 cellular system and HCV-transgenic (HCV-Tg) mice were used to characterize HCV-induced ER stress/UPR pathway activation and adaptation. HCV induced a wave of acute ER stress peaking 2–5 days post-infection, which rapidly subsided thereafter. UPR pathways were activated including IRE1 and EIF2α phosphorylation, ATF6 cleavage and XBP-1 splicing. Downstream target genes including GADD34, ERdj4, p58ipk, ATF3 and ATF4 were upregulated. CHOP, a UPR regulated protein was activated and translocated to the nucleus. Remarkably, UPR activity did not return to baseline but remained elevated for up to 14 days post infection suggesting that chronic ER stress is induced. At this time, cells adapted to ER stress and were less responsive to further drug-induced ER stress. Similar results were obtained in HCV-Tg mice. Suppression of HCV by Interferon-α 2a treatment, restored UPR responsiveness to ER stress tolerant cells.

Conclusions

Our study shows, for the first time, that HCV induces adaptation to chronic ER stress which was reversed upon viral suppression. These finding represent a novel viral mechanism to manipulate cellular response pathways.     

Infusion of glucose and lipids at physiological rates causes acute endoplasmic reticulum stress in rat liver

Endoplasmic reticulum (ER) stress has recently been implicated as a cause for obesity-related insulin resistance; however, what causes ER stress in obesity has remained uncertain. Here, we have tested the hypothesis that macronutrients can cause acute (ER) stress in rat liver. Examined were the effects of intravenously infused glucose and/or lipids on proximal ER stress sensor activation (PERK, eIF2-α, ATF4, Xbox protein 1 (XBP1s)), unfolded protein response (UPR) proteins (GRP78, calnexin, calreticulin, protein disulphide isomerase (PDI), stress kinases (JNK, p38 MAPK) and insulin signaling (insulin/receptor substrate (IRS) 1/2 associated phosphoinositol-3-kinase (PI3K)) in rat liver. Glucose and/or lipid infusions, ranging from 23.8 to 69.5 kJ/4 h (equivalent to between ~17% and ~50% of normal daily energy intake), activated the proximal ER stress sensor PERK and ATF6 increased the protein abundance of calnexin, calreticulin and PDI and increased two GRP78 isoforms. Glucose and glucose plus lipid infusions induced comparable degrees of ER stress, but only infusions containing lipid activated stress kinases (JNK and p38 MAPK) and inhibited insulin signaling (PI3K). In summary, physiologic amounts of both glucose and lipids acutely increased ER stress in livers 12-h fasted rats and dependent on the presence of fat, caused insulin resistance. We conclude that this type of acute ER stress is likely to occur during normal daily nutrient intake.     

Effect of Vitamin D on Peripheral Blood Mononuclear Cells from Patients with Psoriasis Vulgaris and Psoriatic Arthritis

BackgroundPsoriasis, a chronic skin disease with or without joint inflammation, has increased circulating proinflammatory cytokine levels. Vitamin D is involved in calcium homeostasis, bone formation, osteoclastogenesis and osteoclast activity, as well as regulation of immune response. We aimed to study osteoclast differentiation and cytokine secretion of peripheral blood mononuclear cells (PBMCs) from patients with psoriasis vulgaris and psoriatic arthritis, in response to 1,25(OH)₂D₃.MethodsSerum levels of bone turnover markers were measured by ELISA in patients with psoriasis vulgaris and psoriatic arthritis, and healthy controls. PBMCs were isolated and cultured with or without RANKL/M-CSF and 1,25(OH)₂D₃. Osteoclast differentiation and cytokine secretion were assessed.ResultsPsoriatic arthritis patients had lower osteocalcin, as well as higher C-telopeptide of type I collagen and cathepsin K serum levels compared with psoriasis vulgaris patients and controls. RANKL/M-CSF-stimulated PBMCs from psoriatic arthritis patients produced higher proinflammatory cytokine levels and had a differential secretion profile in response to 1,25(OH)₂D₃, compared with psoriasis vulgaris and control PBMCs.ConclusionsOur data confirmed altered bone turnover in psoriatic arthritis patients, and demonstrated increased osteoclastogenic potential and proinflammatory cytokine secretion capacity of these PBMCs compared with psoriasis vulgaris and controls. 1,25(OH)₂D₃ abrogated these effects.     

HSP105 interacts with GRP78 and GSK3 and promotes ER stress-induced caspase-3 activation

Stress of the endoplasmic reticulum (ER stress) is caused by the accumulation of misfolded proteins, which occurs in many neurodegenerative diseases. ER stress can lead to adaptive responses or apoptosis, both of which follow activation of the unfolded protein response (UPR). Heat shock proteins (HSP) support the folding and function of many proteins, and are important components of the ER stress response, but little is known about the role of one of the major large HSPs, HSP105. We identified several new partners of HSP105, including glycogen synthase kinase-3 (GSK3), a promoter of ER stress-induced apoptosis, and GRP78, a key component of the UPR. Knockdown of HSP105 did not alter UPR signaling after ER stress, but blocked caspase-3 activation after ER stress. In contrast, caspase-3 activation induced by genotoxic stress was unaffected by knockdown of HSP105, suggesting ER stress-specificity in the apoptotic action of HSP105. However, knockdown of HSP105 did not alter cell survival after ER stress, but instead diverted signaling to a caspase-3-independent cell death pathway, indicating that HSP105 is necessary for apoptotic signaling after UPR activation by ER stress. Thus, HSP105 appears to chaperone the responses to ER stress through its interactions with GRP78 and GSK3, and without HSP105 cell death following ER stress proceeds by a non-caspase-3-dependent process.