bims-unfpre Biomed News
on Unfolded protein response
Issue of 2020‒12‒13
thirteen papers selected by
Susan Logue
University of Manitoba

  1. ACS Chem Biol. 2020 Dec 08.
      Inositol-requiring enzyme 1α (IRE1α) is one of three endoplasmic reticulum stress sensors. Upon activation of its kinase domain, IRE1α splices the mRNA substrate XBP1, which activates the unfolded protein response. IRE1α has emerged as a therapeutic target as its hyperactivation is implicated in various diseases. Kinase inhibiting RNase attenuator 6 (KIRA6) is an allosteric IRE1α inhibitor targeting the ATP binding pocket, resulting in effective blockage of the IRE1α-XBP1 pathway in mouse models of diabetes and pain. However, recent studies indicate that KIRA6 is not as selective as initially thought. Here, we developed a photoaffinity-based KIRA6 probe to reveal its selectivity. Surprisingly, the majority of off-targets that we identified were not protein kinases but mostly nucleotide-binding proteins. Furthermore, we found that the promiscuous off-target profile of KIRA6 is not cell-line-dependent. Overall, this study calls for caution when KIRA6 is used in IRE1α-targeted studies and illustrates the power of kinase photoaffinity probes.
  2. Cell Death Discov. 2020 Nov 19. 6(1): 128
      Glomerular epithelial cell (GEC)/podocyte proteostasis is dysregulated in glomerular diseases. The unfolded protein response (UPR) is an adaptive pathway in the endoplasmic reticulum (ER) that upregulates proteostasis resources. This study characterizes mechanisms by which inositol requiring enzyme-1α (IRE1α), a UPR transducer, regulates proteostasis in GECs. Mice with podocyte-specific deletion of IRE1α (IRE1α KO) were produced and nephrosis was induced with adriamycin. Compared with control, IRE1α KO mice had greater albuminuria. Adriamycin increased glomerular ER chaperones in control mice, but this upregulation was impaired in IRE1α KO mice. Likewise, autophagy was blunted in adriamycin-treated IRE1α KO animals, evidenced by reduced LC3-II and increased p62. Mitochondrial ultrastructure was markedly disrupted in podocytes of adriamycin-treated IRE1α KO mice. To pursue mechanistic studies, GECs were cultured from glomeruli of IRE1α flox/flox mice and IRE1α was deleted by Cre-lox recombination. In GECs incubated with tunicamycin, deletion of IRE1α attenuated upregulation of ER chaperones, LC3 lipidation, and LC3 transcription, compared with control GECs. Deletion of IRE1α decreased maximal and ATP-linked oxygen consumption, as well as mitochondrial membrane potential. In summary, stress-induced chaperone production, autophagy, and mitochondrial health are compromised by deletion of IRE1α. The IRE1α pathway is cytoprotective in glomerular disease associated with podocyte injury and ER stress.
  3. Elife. 2020 Dec 09. pii: e62601. [Epub ahead of print]9
      The metazoan endoplasmic reticulum (ER) serves both as a hub for maturation of secreted proteins and as an intracellular calcium storage compartment, facilitating calcium release-dependent cellular processes. ER calcium depletion robustly activates the unfolded protein response (UPR). However, it is unclear how fluctuations in ER calcium impact organellar proteostasis. Here we report that calcium selectively affects the dynamics of the abundant metazoan ER Hsp70 chaperone BiP, by enhancing its affinity for ADP. In the calcium-replete ER, ADP rebinding to post-ATP hydrolysis BiP-substrate complexes competes with ATP binding during both spontaneous and co-chaperone-assisted nucleotide exchange, favouring substrate retention. Conversely, in the calcium-depleted ER, relative acceleration of ADP-to-ATP exchange favours substrate release. These findings explain the rapid dissociation of certain substrates from BiP observed in the calcium-depleted ER and suggest a mechanism for tuning ER quality control and coupling UPR activity to signals that mobilise ER calcium in secretory cells.
    Keywords:  biochemistry; cell biology; chemical biology; none
  4. Cell Rep. 2020 Dec 08. pii: S2211-1247(20)31478-9. [Epub ahead of print]33(10): 108489
      In multicellular organisms, neurons integrate a diverse array of external cues to affect downstream changes in organismal health. Specifically, activation of the endoplasmic reticulum (ER) unfolded protein response (UPRER) in neurons increases lifespan by preventing age-onset loss of ER proteostasis and driving lipid depletion in a cell non-autonomous manner. The mechanism of this communication is dependent on the release of small clear vesicles from neurons. We find dopaminergic neurons are necessary and sufficient for activation of cell non-autonomous UPRER to drive lipid depletion in peripheral tissues, whereas serotonergic neurons are sufficient to drive protein homeostasis in peripheral tissues. These signaling modalities are unique and independent and together coordinate the beneficial effects of neuronal cell non-autonomous ER stress signaling upon health and longevity.
    Keywords:  UPRER; aging; non-autonomous signaling; stress response
  5. Mol Med Rep. 2021 Feb;pii: 116. [Epub ahead of print]23(2):
      Endoplasmic reticulum stress (ERS) contributes to the pathogenesis of myocardial ischemia/reperfusion injury and myocardial infarction (MI). Long non-coding RNAs (lncRNAs) serve an important role in cardiovascular diseases, and lncRNA discrimination antagonizing non-protein coding RNA (Dancr) alleviates cardiomyocyte damage. microRNA (miR)-6324 was upregulated in MI model rats and was predicted to bind to Dancr. The present study aimed to investigate the role of Dancr in ERS-induced cardiomyocytes and the potential underlying mechanisms. Tunicamycin (Tm) was used to induce ERS. Cell viability, apoptosis and levels of associated proteins, ERS and autophagy in Dancr-overexpression H9C2 cells and miR-6234 mimic-transfected H9C2 cells were assessed using Cell Counting Kit-8, TUNEL staining and western blot assay, respectively. The results suggested that Dancr expression levels and cell viability were downregulated by Tm in a concentration-dependent manner compared with the control group. Tm induced apoptosis, ERS and autophagy, as indicated by an increased ratio of apoptotic cells, increased expression levels of Bax, cleaved (c)-caspase-3/9, glucose-regulated protein 78 kDa (GRP78), phosphorylated (p)-inositol-requiring enzyme-1α (IRE1α), spliced X-box-binding protein 1 (Xbp1s), IRE1α, activating transcription factor (ATF)6, ATF4, Beclin 1 and microtubule associated protein 1 light chain 3α (LC3)II/I, and decreased expression levels of Bcl-2, unspliced Xbp1 (Xbp1u) and p62 in the Tm group compared with the control group. Moreover, the results indicated that compared with the Tm + overexpression (Oe)-negative control (NC) group, the Tm + Oe-Dancr group displayed decreased apoptosis, but enhanced ERS and autophagy to restore cellular homeostasis. Compared with the Tm + Oe-NC group, the Tm + Oe-Dancr group decreased the ratio of apoptotic cells, decreased expression levels of Bax, c-caspase-3/9 and Xbp1u, and increased expression levels of Bcl-2, p-IRE1α, Xbp1s, Beclin 1 and LC3II/I. Dancr overexpression also significantly downregulated miR-6324 expression compared with Oe-NC. The dual-luciferase reporter assay further indicated an interaction between Dancr and miR-6324. In addition, miR-6324 mimic partially reversed the effects of Dancr overexpression on Tm-induced apoptosis, ERS and autophagy. In conclusion, lncRNA Dancr overexpression protected cardiomyocytes against ERS injury via sponging miR-6324, thus inhibiting apoptosis, enhancing autophagy and restoring ER homeostasis.
  6. Nat Commun. 2020 12 10. 11(1): 6330
      Endothelial cell (EC) metabolism is thought to be one of the driving forces for angiogenesis. Here we report the identification of the hexosamine D-mannosamine (ManN) as an EC mitogen and survival factor for bovine and human microvascular EC, with an additivity with VEGF. ManN inhibits glycosylation in ECs and induces significant changes in N-glycan and O-glycan profiles. We further demonstrate that ManN and two N-glycosylation inhibitors stimulate EC proliferation via both JNK activation and the unfolded protein response caused by ER stress. ManN results in enhanced angiogenesis in a mouse skin injury model. ManN also promotes angiogenesis in a mouse hindlimb ischemia model, with accelerated limb blood flow recovery compared to controls. In addition, intraocular injection of ManN induces retinal neovascularization. Therefore, activation of stress pathways following inhibition of protein glycosylation can promote EC proliferation and angiogenesis and may represent a therapeutic strategy for treatment of ischemic disorders.
  7. Am J Pathol. 2020 Dec 08. pii: S0002-9440(20)30553-8. [Epub ahead of print]
      ER stress is shown to promote nucleus pulposus (NP) cell apoptosis and intervertebral disc degeneration. However, little is known about ER stress regulation by the hypoxic disc microenvironment and its contribution to extracellular matrix homeostasis. NP cells were cultured under hypoxia (1% pO2) to assess ER stress status and gain- and loss-of-function approaches were used to assess the role of HIF-1α in this pathway. Additionally, the contribution of ER stress induction on the NP cell secretome was assessed by a non-targeted quantitative proteomic analysis by SWATH-mass spectrometry. NP cells exhibited a lower ER stress burden under hypoxia. Knockdown of HIF-1α increased CHOP, PERK, and ATF6 levels; whereas HIF-1α stabilization decreased the expression of ER stress markers Ddit3, Hsp5a, Atf6, and Eif2a. Interestingly, ER stress inducers Tunicamycin and Thapsigargin induced HIF-1α activity under hypoxia while promoting the unfolded protein response (UPR). NP cell secretome analysis demonstrated an impact of ER stress induction on extracellular matrix secretion, with decreases in collagens and cell adhesion-related proteins. Moreover, analysis of transcriptomic data of NP tissues from aged mice and degenerated human discs showed higher levels of UPR markers and decreased levels of matrix components. Our study shows for the first time that hypoxia and HIF-1α attenuate ER stress responses in NP cells, and ER stress promotes inefficient extracellular matrix secretion under hypoxia.
    Keywords:  ER stress; HIF-1α; Nucleus pulposus; collagens; extracellular matrix; hypoxia; intervertebral disc; proteomics; secretome
  8. J Mater Chem B. 2020 Dec 08.
      The endoplasmic reticulum (ER) apparatus is a part of the secretory pathway that transports proteins to the plasma membrane through vesicle trafficking, enabling post-translational modification of the newly synthesized proteins. Several diseases such as inflammation, neurodegenerative disorder, and bipolar disorder are closely associated with dysfunction of the ER stress response. Herein, we present an ER-targeting, intracellular delivery approach that utilized cell-penetrating peptide (CPP)-conjugated lipid/polymer hybrid nanovehicles (LPNVs). For this, we patched Penetratin, a type of CPP, onto the LPNVs with vesicular membranes formulated with poly(ethylene oxide)-b-poly(ε-caprolactone)-b-poly(ethylene oxide) (PEO-b-PCL-b-PEO) and 1,2-dipalmitoyl-sn-glycero-3-phosphocholine (DPPC). We found that the Penetratin-conjugated LPNV (LPNVPnt) was readily taken up by cells and showed specific ER-targeting ability, which was comparable to that of LPNVs conjugated with other types of CPPs. Moreover, we observed that remarkable lysosomal escape of the LPNVs occurred due to effective pH buffering with the aid of PEO-b-PCL-b-PEO. These results highlighted that our LPNVPnt system could pave the way for the development of an elaborate drug delivery technology for ER-targeting at the intracellular level.
  9. iScience. 2020 Dec 18. 23(12): 101810
      Endoplasmic reticulum (ER) stress is known to induce pro-inflammatory response and ultimately leads to cell death. Mesencephalic astrocyte-derived neurotrophic factor (MANF) is an ER-localized protein whose expression and secretion is induced by ER stress and a crucial survival factor. However, the underlying mechanism of how MANF exerts its cytoprotective activity remains unclear due to the lack of knowledge of its receptor. Here we show that Neuroplastin (NPTN) is such a receptor for MANF. Biochemical analysis shows the physiological interaction between MANF and NPTN on the cell surface. Binding of MANF to NPTN mitigates the inflammatory response and apoptosis via suppression of NF-kβ signaling. Our results demonstrate that NPTN is a cell surface receptor for MANF, which modulates inflammatory responses and cell death, and that the MANF-NPTN survival signaling described here provides potential therapeutic targets for the treatment of ER stress-related disorders, including diabetes mellitus, neurodegeneration, retinal degeneration, and Wolfram syndrome.
    Keywords:  Biochemistry; Cell Biology; Molecular Biology
  10. Nutrients. 2020 Dec 08. pii: E3771. [Epub ahead of print]12(12):
      Growth differentiation factor-15 (GDF-15) and its receptor GFRAL are both involved in the development of obesity and insulin resistance. Plasmatic GDF-15 level increases with obesity and is positively associated with disease progression. Despite macrophages have been recently suggested as a key source of GDF-15 in obesity, little is known about the regulation of GDF-15 in these cells. In the present work, we sought for potential pathophysiological activators of GDF15 expression in human macrophages and identified saturated fatty acids (SFAs) as strong inducers of GDF15 expression and secretion. SFAs increase GDF15 expression through the induction of an ER stress and the activation of the PERK/eIF2/CHOP signaling pathway in both PMA-differentiated THP-1 cells and in primary monocyte-derived macrophages. The transcription factor CHOP directly binds to the GDF15 promoter region and regulates GDF15 expression. Unlike SFAs, unsaturated fatty acids do not promote GDF15 expression and rather inhibit both SFA-induced GDF15 expression and ER stress. These results suggest that free fatty acids may be involved in the control of GDF-15 and provide new molecular insights about how diet and lipid metabolism may regulate the development of obesity and T2D.
    Keywords:  CHOP; ER stress; GDF15; macrophage; obesity; saturated fatty acids; stearate
  11. Cancer Biol Med. 2020 Nov 15. 17(4): 842-863
      Despite advances in cancer therapeutics and the integration of personalized medicine, the development of chemoresistance in many patients remains a significant contributing factor to cancer mortality. Upon treatment with chemotherapeutics, the disruption of homeostasis in cancer cells triggers the adaptive response which has emerged as a key resistance mechanism. In this review, we summarize the mechanistic studies investigating the three major components of the adaptive response, autophagy, endoplasmic reticulum (ER) stress signaling, and senescence, in response to cancer chemotherapy. We will discuss the development of potential cancer therapeutic strategies in the context of these adaptive resistance mechanisms, with the goal of stimulating research that may facilitate the development of effective cancer therapy.
    Keywords:  Cancer; ER stress signaling; adaptive response; autophagy; chemoresistance; chemotherapy; senescence
  12. Cell Signal. 2020 Dec 08. pii: S0898-6568(20)30357-0. [Epub ahead of print] 109880
      In cartilage, chondrocytes are responsible for the biogenesis and maintenance of the extracellular matrix (ECM) composed of proteins, glycoproteins and proteoglycans. Various cellular stresses, such as hypoxia, nutrient deprivation, oxidative stress or the accumulation of advanced glycation end products (AGEs) during aging, but also translational errors or mutations in cartilage components or chaperone proteins affect the synthesis and secretion of ECM proteins, causing protein aggregates to accumulate in the endoplasmic reticulum (ER). This condition, referred to as ER stress, interferes with cartilage cell homeostasis and initiates the unfolded protein response (UPR), a rescue mechanism to regain cell viability and function. Chronic or irreversible ER stress, however, triggers UPR-initiated cell death. Due to unresolved ER stress in chondrocytes, diseases of the skeletal system, such as chondrodysplasias, arise. ER stress has also been identified as a contributing factor to the pathogenesis of cartilage degeneration processes such as osteoarthritis (OA). This review provides current knowledge about the biogenesis of ECM components in chondrocytes, describes possible causes for the impairment of involved processes and focuses on the ER stress-induced cell death in articular cartilage during OA. Targeting of the ER stress itself or intervention in UPR signaling to reduce death of chondrocytes may be promising for future osteoarthritis therapy.
    Keywords:  Apoptosis; Cartilage; Chondroptosis; ER stress; Necroptosis; Osteoarthritis; Unfolded protein response
  13. Aging (Albany NY). 2020 Dec 03. 12
      Genome-wide association studies have revealed that multiple single-nucleotide polymorphisms in the intergenic region between estrogen receptor 1 and coiled-coil domain containing 170 (CCDC170) are associated with breast cancer risk. We performed microarray and bioinformatics analyses to identify genes that were induced upon CCDC170 overexpression, and confirmed our findings by evaluating paraffin-embedded breast cancer tissues and conducting cellular assays. In CCDC170-overexpressing MCF7 breast cancer cells, microarray analyses revealed that inositol-requiring enzyme 1 (IRE1) was the most elevated gene in enriched pathways. In breast cancer tissues, IRE1 expression correlated positively with CCDC170 and X-box binding protein 1 expression at both the mRNA and protein levels. In a survival analysis, patients with higher CCDC170 levels exhibited better disease-free survival. Western blotting indicated that overexpressing CCDC170 in MCF7 cells increased protein levels of IRE1α, estrogen receptor α and X-box binding protein 1, while silencing CCDC170 reduced them. CCDC170 overexpression promoted apoptosis in MCF7 cells, and this effect was more obvious under endoplasmic reticulum stress. MCF7 cells overexpressing CCDC170 were more sensitive to paclitaxel. Our study showed that higher CCDC170 expression is associated with a better prognosis in breast cancer patients and that CCDC170 may promote apoptosis through the IRE1α pathway.
    Keywords:  CCDC170; IRE1α; XBP1; apoptosis; breast cancer