bims-unfpre Biomed News
on Unfolded protein response
Issue of 2019‒12‒29
twelve papers selected by
Susan Logue
University of Manitoba


  1. Proc Natl Acad Sci U S A. 2019 Dec 23. pii: 201915311. [Epub ahead of print]
    Belyy V, Tran NH, Walter P.
      The endoplasmic reticulum (ER) membrane-resident stress sensor inositol-requiring enzyme 1 (IRE1) governs the most evolutionarily conserved branch of the unfolded protein response. Upon sensing an accumulation of unfolded proteins in the ER lumen, IRE1 activates its cytoplasmic kinase and ribonuclease domains to transduce the signal. IRE1 activity correlates with its assembly into large clusters, yet the biophysical characteristics of IRE1 clusters remain poorly characterized. We combined superresolution microscopy, single-particle tracking, fluorescence recovery, and photoconversion to examine IRE1 clustering quantitatively in living human and mouse cells. Our results revealed that: 1) In contrast to qualitative impressions gleaned from microscopic images, IRE1 clusters comprise only a small fraction (∼5%) of the total IRE1 in the cell; 2) IRE1 clusters have complex topologies that display features of higher-order organization; 3) IRE1 clusters contain a diffusionally constrained core, indicating that they are not phase-separated liquid condensates; 4) IRE1 molecules in clusters remain diffusionally accessible to the free pool of IRE1 molecules in the general ER network; 5) when IRE1 clusters disappear at later time points of ER stress as IRE1 signaling attenuates, their constituent molecules are released back into the ER network and not degraded; 6) IRE1 cluster assembly and disassembly are mechanistically distinct; and 7) IRE1 clusters' mobility is nearly independent of cluster size. Taken together, these insights define the clusters as dynamic assemblies with unique properties. The analysis tools developed for this study will be widely applicable to investigations of clustering behaviors in other signaling proteins.
    Keywords:  IRE1; clustering; quantitative microscopy; signaling; unfolded protein response
    DOI:  https://doi.org/10.1073/pnas.1915311117
  2. Elife. 2019 Dec 24. pii: e50793. [Epub ahead of print]8
    Amin-Wetzel N, Neidhardt L, Yan Y, Mayer MP, Ron D.
      Coupling of endoplasmic reticulum stress to dimerisation‑dependent activation of the UPR transducer IRE1 is incompletely understood. Whilst the luminal co-chaperone ERdj4 promotes a complex between the Hsp70 BiP and IRE1's stress-sensing luminal domain (IRE1LD) that favours the latter's monomeric inactive state and loss of ERdj4 de-represses IRE1, evidence linking these cellular and in vitro observations is presently lacking. We report that enforced loading of endogenous BiP onto endogenous IRE1α repressed UPR signalling in CHO cells and deletions in the IRE1α locus that de-repressed the UPR in cells, encode flexible regions of IRE1LD that mediated BiP‑induced monomerisation in vitro. Changes in the hydrogen exchange mass spectrometry profile of IRE1LD induced by ERdj4 and BiP confirmed monomerisation and were consistent with active destabilisation of the IRE1LD dimer. Together, these observations support a competition model whereby waning ER stress passively partitions ERdj4 and BiP to IRE1LD to initiate active repression of UPR signalling.
    Keywords:  E. coli; cell biology
    DOI:  https://doi.org/10.7554/eLife.50793
  3. Cell Struct Funct. 2019 Dec 26.
    Jin B, Ishikawa T, Taniguchi M, Ninagawa S, Okada T, Kagaya S, Mori K.
      Three types of transmembrane protein, IRE1α/IRE1β, PERK, and ATF6α/ATF6β, are expressed ubiquitously in vertebrates as transducers of the unfolded protein response (UPR), which maintains the homeostasis of the endoplasmic reticulum. IRE1 is highly conserved from yeast to mammals, and transmits a signal by a unique mechanism, namely splicing of mRNA encoding XBP1, the transcription factor downstream of IRE1 in metazoans. IRE1 contains a ribonuclease domain in its cytoplasmic region which initiates splicing reaction by direct cleavage of XBP1 mRNA at the two stem loop structures. As the UPR is considered to be involved in the development and progression of various diseases, as well as in the survival and growth of tumor cells, UPR inhibitors have been sought. To date, IRE1 inhibitors have been screened using cell-based reporter assays and fluorescent-based in vitro cleavage assays. Here, we used medaka fish to develop an in vivo assay for IRE1α inhibitors. IRE1α, IRE1β, ATF6α and ATF6β are ubiquitously expressed in medaka. We found that IRE1α/ATF6α-double knockout is lethal, similarly to IRE1α/IRE1β- and ATF6α/ATF6β-double knockout. Therefore, IRE1 inhibitors are expected to confer lethality to ATF6α-knockout medaka but not to wild-type medaka. One compound named K114 was obtained from 1,280 compounds using this phenotypic screening. K114 inhibited ER stress-induced splicing of XBP1 mRNA as well as reporter luciferase expression in HCT116 cells derived from human colorectal carcinoma, and inhibited ribonuclease activity of human IRE1α in vitro. Thus, this phenotypic assay can be used as a quick test for the efficacy of IRE1α inhibitors in vivo.Key Words: endoplasmic reticulum, inhibitor screening, mRNA splicing, phenotypic assay, unfolded protein response.
    Keywords:  endoplasmic reticulum; inhibitor screening; mRNA splicing; phenotypic assay; unfolded protein response
    DOI:  https://doi.org/10.1247/csf.19032
  4. Front Immunol. 2019 ;10 2825
    Coleman OI, Haller D.
      An imbalance in the correct protein folding milieu of the endoplasmic reticulum (ER) can cause ER stress, which leads to the activation of the unfolded protein response (UPR). The UPR constitutes a highly conserved and intricately regulated group of pathways that serve to restore ER homeostasis through adaptation or apoptosis. Numerous studies over the last decade have shown that the UPR plays a critical role in shaping immunity and inflammation, resulting in the recognition of the UPR as a key player in pathological processes including complex inflammatory, autoimmune and neoplastic diseases. The intestinal epithelium, with its many highly secretory cells, forms an important barrier and messenger between the luminal environment and the host immune system. It is not surprising, that numerous studies have associated ER stress and the UPR with intestinal diseases such as inflammatory bowel disease (IBD) and colorectal cancer (CRC). In this review, we discuss our current understanding of the roles of ER stress and the UPR in shaping immune responses and maintaining tissue homeostasis. Furthermore, the role played by the UPR in disease, with emphasis on IBD and CRC, is described here. As a key player in immunity and inflammation, the UPR has been increasingly recognized as an important pharmacological target in the development of therapeutic strategies for immune-mediated pathologies. We summarize available strategies targeting the UPR and their therapeutic implications. Understanding the balance between homeostasis and pathophysiology, as well as means of manipulating this balance, provides an important avenue for future research.
    Keywords:  CRC; IBD; UPR; immunity; tissue homeostasis
    DOI:  https://doi.org/10.3389/fimmu.2019.02825
  5. Viruses. 2019 Dec 21. pii: E17. [Epub ahead of print]12(1):
    Johnston BP, McCormick C.
      Herpesviruses usurp cellular stress responses to promote viral replication and avoid immune surveillance. The unfolded protein response (UPR) is a conserved stress response that is activated when the protein load in the ER exceeds folding capacity and misfolded proteins accumulate. The UPR aims to restore protein homeostasis through translational and transcriptional reprogramming; if homeostasis cannot be restored, the UPR switches from "helper" to "executioner", triggering apoptosis. It is thought that the burst of herpesvirus glycoprotein synthesis during lytic replication causes ER stress, and that these viruses may have evolved mechanisms to manage UPR signaling to create an optimal niche for replication. The past decade has seen considerable progress in understanding how herpesviruses reprogram the UPR. Here we provide an overview of the molecular events of UPR activation, signaling and transcriptional outputs, and highlight key evidence that herpesviruses hijack the UPR to aid infection.
    Keywords:  ATF4; ATF6; GADD34; IRE1; Kaposi’s sarcoma-associated herpesvirus (KSHV); PERK; XBP1; cytomegalovirus (CMV); herpes simplex virus (HSV); herpesvirus; integrated stress response (ISR); unfolded protein response (UPR)
    DOI:  https://doi.org/10.3390/v12010017
  6. Cell Rep. 2019 Dec 24. pii: S2211-1247(19)31605-5. [Epub ahead of print]29(13): 4525-4539.e4
    Huang C, Hedl M, Ranjan K, Abraham C.
      LACC1 genetic variants are associated with multiple immune-mediated diseases. However, laccase domain containing-1 (LACC1) functions are incompletely defined. We find that upon stimulation of the pattern-recognition receptor (PRR) NOD2, LACC1 localizes to the endoplasmic reticulum (ER) and forms a complex with ER-stress sensors. All three ER-stress branches, PERK, IRE1α, and ATF6, are required for NOD2-induced signaling, cytokines, and antimicrobial pathways in human macrophages. LACC1, and its localization to the ER, is required for these outcomes. Relative to wild-type (WT) LACC1, transfection of the common Val254 and rare Arg284 immune-mediated disease-risk LACC1 variants into HeLa cells and macrophages, as well as macrophages from LACC1 Val254 carriers, shows reduced NOD2-induced ER stress-associated outcomes; these downstream outcomes are restored by rescuing ER stress. Therefore, we identify ER stress to be essential in PRR-induced outcomes in macrophages, define a critical role for LACC1 in these ER stress-dependent events, and elucidate how LACC1 disease-risk variants mediate these outcomes.
    Keywords:  Crohn's disease; ER stress; genetics; inflammatory bowel disease; innate immunity; macrophages; unfolded protein response
    DOI:  https://doi.org/10.1016/j.celrep.2019.11.105
  7. FEBS J. 2019 Dec 27.
    Bartoszewski R, Gebert M, Janaszak-Jasiecka A, Cabaj A, Króliczewski J, Bartoszewska S, Sobolewska A, Crossman DK, Ochocka R, Kamysz W, Kalinowski L, Dąbrowski M, Collawn JF.
      Endoplasmic reticulum (ER) stress conditions promote a cellular adaptive mechanism called the unfolded protein response (UPR) that utilizes three stress sensors, IRE1, PERK, and ATF6. These sensors activate a number of pathways to reduce the stress and facilitate cell survival. While much is known about the mechanisms involved that modulate apoptosis during chronic stress, less is known about the transition between the pro-survival and pro-apoptotic factors that determine cell fate. Here we employed a genetic screen that utilized three different pharmacological stressors to induce ER stress in a human immortalized airway epithelial cell line, 16HBE14o- cells. We followed the stress responses over an 18-hour time course and utilized real time monitoring of cell survival, next generation sequencing (NGS), and qRT-PCR to identify and validate genes that were upregulated with all three commonly employed ER stressors, ALLN, tunicamycin, and thapsigargin. GADD45A (Growth Arrest and DNA Damage Inducible Alpha), a pro-apoptotic factor, and RCAN1 (Regulator of Calcineurin 1) mRNAs were identified and verified by showing that siRNA knockdown of GADD45A decreased CHOP, PUMA, and NOXA expression, 3 pro-apoptotic factors, and increased cell viability during ER stress conditions, whereas siRNA knockdown of RCAN1 dramatically decreased cell viability. These results suggest that the relative levels of these two genes regulate cell fate decisions during ER stress independent of the type of ER stressor.
    Keywords:   CHOP ; DDIT1 ; GAD45A ; NOXA ; PUMA ; RCAN1 ; ER stress; RNA-seq; UPR
    DOI:  https://doi.org/10.1111/febs.15195
  8. Proc Natl Acad Sci U S A. 2019 Dec 27. pii: 201905641. [Epub ahead of print]
    Albert S, Wietrzynski W, Lee CW, Schaffer M, Beck F, Schuller JM, Salomé PA, Plitzko JM, Baumeister W, Engel BD.
      To promote the biochemical reactions of life, cells can compartmentalize molecular interaction partners together within separated non-membrane-bound regions. It is unknown whether this strategy is used to facilitate protein degradation at specific locations within the cell. Leveraging in situ cryo-electron tomography to image the native molecular landscape of the unicellular alga Chlamydomonas reinhardtii, we discovered that the cytosolic protein degradation machinery is concentrated within ∼200-nm foci that contact specialized patches of endoplasmic reticulum (ER) membrane away from the ER-Golgi interface. These non-membrane-bound microcompartments exclude ribosomes and consist of a core of densely clustered 26S proteasomes surrounded by a loose cloud of Cdc48. Active proteasomes in the microcompartments directly engage with putative substrate at the ER membrane, a function canonically assigned to Cdc48. Live-cell fluorescence microscopy revealed that the proteasome clusters are dynamic, with frequent assembly and fusion events. We propose that the microcompartments perform ER-associated degradation, colocalizing the degradation machinery at specific ER hot spots to enable efficient protein quality control.
    Keywords:  ERAD; cdc48; cryo-electron tomography; phase separation; proteasome
    DOI:  https://doi.org/10.1073/pnas.1905641117
  9. PLoS Genet. 2019 Dec 23. 15(12): e1008563
    Hong ZH, Qing T, Schubert D, Kleinmanns JA, Liu JX.
      The unfolded protein response (UPR) is required for protein homeostasis in the endoplasmic reticulum (ER) when plants are challenged by adverse environmental conditions. Inositol-requiring enzyme 1 (IRE1), the bifunctional protein kinase / ribonuclease, is an important UPR regulator in plants mediating cytoplasmic splicing of the mRNA encoding the transcription factor bZIP60. This activates the UPR signaling pathway and regulates canonical UPR genes. However, how the protein activity of IRE1 is controlled during plant growth and development is largely unknown. In the present study, we demonstrate that the nuclear and Golgi-localized protein BLISTER (BLI) negatively controls the activity of IRE1A/IRE1B under normal growth condition in Arabidopsis. Loss-of-function mutation of BLI results in chronic up-regulation of a set of both canonical UPR genes and non-canonical UPR downstream genes, leading to cell death and growth retardation. Genetic analysis indicates that BLI-regulated vegetative growth phenotype is dependent on IRE1A/IRE1B but not their canonical splicing target bZIP60. Genetic complementation with mutation analysis suggests that the D570/K572 residues in the ATP-binding pocket and N780 residue in the RNase domain of IRE1A are required for the activation of canonical UPR gene expression, in contrast, the D570/K572 residues and D590 residue in the protein kinase domain of IRE1A are important for the induction of non-canonical UPR downstream genes in the BLI mutant background, which correlates with the shoot growth phenotype. Hence, our results reveal the important role of IRE1A in plant growth and development, and BLI negatively controls IRE1A's function under normal growth condition in plants.
    DOI:  https://doi.org/10.1371/journal.pgen.1008563
  10. Int J Mol Sci. 2019 Dec 25. pii: E169. [Epub ahead of print]21(1):
    Ramirez MU, Hernandez SR, Soto-Pantoja DR, Cook KL.
      Despite advances in cancer therapy, several persistent issues remain. These include cancer recurrence, effective targeting of aggressive or therapy-resistant cancers, and selective treatments for transformed cells. This review evaluates the current findings and highlights the potential of targeting the unfolded protein response to treat cancer. The unfolded protein response, an evolutionarily conserved pathway in all eukaryotes, is initiated in response to misfolded proteins accumulating within the lumen of the endoplasmic reticulum. This pathway is initially cytoprotective, allowing cells to survive stressful events; however, prolonged activation of the unfolded protein response also activates apoptotic responses. This balance is key in successful mammalian immune response and inducing cell death in malignant cells. We discuss how the unfolded protein response affects cancer progression, survival, and immune response to cancer cells. The literature shows that targeting the unfolded protein response as a monotherapy or in combination with chemotherapy or immunotherapies increases the efficacy of these drugs; however, systemic unfolded protein response targeting may yield deleterious effects on immune cell function and should be taken into consideration. The material in this review shows the promise of both approaches, each of which merits further research.
    Keywords:  Activating transcription factor 6 (ATF6); Glucose-regulated protein 78 (GRP78); Inositol-requiring enzyme 1 (IRE1); PKR-like endoplasmic reticulum kinase (PERK); T cell; immune cells; macrophage; tumor microenvironment; unfolded protein response
    DOI:  https://doi.org/10.3390/ijms21010169
  11. Cytokine. 2019 Dec 23. pii: S1043-4666(19)30388-6. [Epub ahead of print]127 154959
    Oh HJ, Lee S, Park PH.
      Adiponectin, the most abundant adipokine, exhibits various physiological functions. In addition to its critical role in lipid metabolism, recent studies have demonstrated its potent anti-inflammatory and cytoprotective properties. Accumulating evidence suggests that autophagy plays a critical role in various biological responses by adiponectin. However, the underlying mechanisms remain elusive. Herein, we investigated the role of ER stress in adiponectin-induced autophagy and its functional roles in biological responses by adiponectin in macrophages. In this study, globular adiponectin (gAcrp) significantly increased the expression of various ER stress markers in both RAW 264.7 and primary peritoneal macrophages. In addition, inhibition of ER stress by treatment with tauroursodeoxycholic acid (TUDCA) or gene silencing of CHOP prominently suppressed gAcrp-induced autophagy. Treatment with gAcrp also induced significant increase in sestrin2 expression. Interestingly, knockdown of sestrin2 prevented autophagy induction and inhibition of ER stress abrogated sestrin2 induction by gAcrp, collectively implying that ER stress critically contributes to gAcrp-induced autophagy activation via sestrin2 induction. Moreover, pretreatment with TUDCA restored suppression of TNF-α and IL-1β expression and attenuated the enhanced viability of macrophages induced by gAcrp. Taken together, these findings indicate the potential role of ER stress in autophagy activation, modulation of inflammatory responses, and cell survival by gAcrp in macrophages.
    Keywords:  Adiponectin; Autophagy; ER stress; Inflammation
    DOI:  https://doi.org/10.1016/j.cyto.2019.154959
  12. Front Immunol. 2019 ;10 2797
    Wang LX, Zhu XM, Yao YM.
      Sestrin2 (SESN2), a highly evolutionarily conserved protein, is critically involved in cellular responses to various stresses. SESN2 has a protective effect on physiological and pathological states mainly via regulating oxidative stress, endoplasmic reticulum stress, autophagy, metabolism, and inflammation. In recent years, breakthrough investigations with regard to the regulation and signaling mechanisms of SESN2 have markedly deepened our understanding of its potential role as well as its significance in host response. However, the functions of SESN2 in the immune system and inflammation remain elusive. It has been documented that many immune cells positively express SESN2 and, in turn, that SESN2 might modulate cellular activities. This review incorporates recent progress and aims to provide novel insight into the protective role and regulatory pathway of SESN2, which acts as a potential biomarker and therapeutic target in the context of various diseases.
    Keywords:  Sestrin2; autophagy; endoplasmic reticulum stress; immune cell; immune response
    DOI:  https://doi.org/10.3389/fimmu.2019.02797