bims-unfpre 2020-02-02 papers

Issue of 2020‒02‒02
six papers selected by
Susan Logue
University of Manitoba

Sci Rep. 2020 Jan 29. 10(1): 1438

Activation of pH-Sensing Receptor OGR1 (GPR68) Induces ER Stress Via the IRE1α/JNK Pathway in an Intestinal Epithelial Cell Model.

Chiaki Maeyashiki, Hassan Melhem, Larissa Hering, Katharina Baebler, Jesus Cosin-Roger, Fabian Schefer, Bruce Weder, Martin Hausmann, Michael Scharl, Gerhard Rogler, Cheryl de Vallière, Pedro A Ruiz.

Proton-sensing ovarian cancer G-protein coupled receptor (OGR1) plays an important role in pH homeostasis. Acidosis occurs at sites of intestinal inflammation and can induce endoplasmic reticulum (ER) stress and the unfolded protein response (UPR), an evolutionary mechanism that enables cells to cope with stressful conditions. ER stress activates autophagy, and both play important roles in gut homeostasis and contribute to the pathogenesis of inflammatory bowel disease (IBD). Using a human intestinal epithelial cell model, we investigated whether our previously observed protective effects of OGR1 deficiency in experimental colitis are associated with a differential regulation of ER stress, the UPR and autophagy. Caco-2 cells stably overexpressing OGR1 were subjected to an acidic pH shift. pH-dependent OGR1-mediated signalling led to a significant upregulation in the ER stress markers, binding immunoglobulin protein (BiP) and phospho-inositol required 1α (IRE1α), which was reversed by a novel OGR1 inhibitor and a c-Jun N-terminal kinase (JNK) inhibitor. Proton-activated OGR1-mediated signalling failed to induce apoptosis, but triggered accumulation of total microtubule-associated protein 1 A/1B-light chain 3, suggesting blockage of late stage autophagy. Our results show novel functions for OGR1 in the regulation of ER stress through the IRE1α-JNK signalling pathway, as well as blockage of autophagosomal degradation. OGR1 inhibition might represent a novel therapeutic approach in IBD.

DOI: https://doi.org/10.1038/s41598-020-57657-9

J Cell Biol. 2020 Feb 03. pii: e201904048. [Epub ahead of print]219(2):

IRE1β negatively regulates IRE1α signaling in response to endoplasmic reticulum stress.

Michael J Grey, Eva Cloots, Mariska S Simpson, Nicole LeDuc, Yevgeniy V Serebrenik, Heidi De Luca, Delphine De Sutter, Phi Luong, Jay R Thiagarajah, Adrienne W Paton, James C Paton, Markus A Seeliger, Sven Eyckerman, Sophie Janssens, Wayne I Lencer.

IRE1β is an ER stress sensor uniquely expressed in epithelial cells lining mucosal surfaces. Here, we show that intestinal epithelial cells expressing IRE1β have an attenuated unfolded protein response to ER stress. When modeled in HEK293 cells and with purified protein, IRE1β diminishes expression and inhibits signaling by the closely related stress sensor IRE1α. IRE1β can assemble with and inhibit IRE1α to suppress stress-induced XBP1 splicing, a key mediator of the unfolded protein response. In comparison to IRE1α, IRE1β has relatively weak XBP1 splicing activity, largely explained by a nonconserved amino acid in the kinase domain active site that impairs its phosphorylation and restricts oligomerization. This enables IRE1β to act as a dominant-negative suppressor of IRE1α and affect how barrier epithelial cells manage the response to stress at the host-environment interface.

DOI: https://doi.org/10.1083/jcb.201904048

Cell Commun Signal. 2020 Jan 27. 18(1): 12

Cell death induced by the ER stressor thapsigargin involves death receptor 5, a non-autophagic function of MAP1LC3B, and distinct contributions from unfolded protein response components.

Paula Lindner, Søren Brøgger Christensen, Poul Nissen, Jesper Vuust Møller, Nikolai Engedal.

BACKGROUND: Cell death triggered by unmitigated endoplasmic reticulum (ER) stress plays an important role in physiology and disease, but the death-inducing signaling mechanisms are incompletely understood. To gain more insight into these mechanisms, the ER stressor thapsigargin (Tg) is an instrumental experimental tool. Additionally, Tg forms the basis for analog prodrugs designed for cell killing in targeted cancer therapy. Tg induces apoptosis via the unfolded protein response (UPR), but how apoptosis is initiated, and how individual effects of the various UPR components are integrated, is unclear. Furthermore, the role of autophagy and autophagy-related (ATG) proteins remains elusive.METHODS: To systematically address these key questions, we analyzed the effects of Tg and therapeutically relevant Tg analogs in two human cancer cell lines of different origin (LNCaP prostate- and HCT116 colon cancer cells), using RNAi and inhibitory drugs to target death receptors, UPR components and ATG proteins, in combination with measurements of cell death by fluorescence imaging and propidium iodide staining, as well as real-time RT-PCR and western blotting to monitor caspase activity, expression of ATG proteins, UPR components, and downstream ER stress signaling.
RESULTS: In both cell lines, Tg-induced cell death depended on death receptor 5 and caspase-8. Optimal cytotoxicity involved a non-autophagic function of MAP1LC3B upstream of procaspase-8 cleavage. PERK, ATF4 and CHOP were required for Tg-induced cell death, but surprisingly acted in parallel rather than as a linear pathway; ATF4 and CHOP were independently required for Tg-mediated upregulation of death receptor 5 and MAP1LC3B proteins, whereas PERK acted via other pathways. Interestingly, IRE1 contributed to Tg-induced cell death in a cell type-specific manner. This was linked to an XBP1-dependent activation of c-Jun N-terminal kinase, which was pro-apoptotic in LNCaP but not HCT116 cells. Molecular requirements for cell death induction by therapy-relevant Tg analogs were identical to those observed with Tg.
CONCLUSIONS: Together, our results provide a new, integrated understanding of UPR signaling mechanisms and downstream mediators that induce cell death upon Tg-triggered, unmitigated ER stress. Video Abstract.

Keywords: ATF4; Apoptosis; Autophagy; CHOP; Caspase-8; Cell death; DR5; IRE1; JNK; LC3B; PERK; SERCA; Thapsigargin; Unfolded protein response; XBP1s

DOI: https://doi.org/10.1186/s12964-019-0499-z

J Cell Sci. 2020 Jan 31. pii: jcs.241539. [Epub ahead of print]

Retrograde signaling mediates an adaptive survival response to endoplasmic reticulum stress.

Imadeddin Hijazi, Jeffrey Knupp, Amy Chang.

One major cause of endoplasmic reticulum (ER) stress is homeostatic imbalance between biosynthetic protein folding and protein folding capacity. Cells utilize mechanisms such as the unfolded protein response (UPR) to cope with ER stress. Nevertheless, when ER stress is prolonged or severe, cell death may occur, accompanied by production of mitochondrial reactive oxygen species (ROS). Using a yeast model, we describe an innate, adaptive response to ER stress to increase select mitochondrial proteins, O2 consumption, and cell survival. The mitochondrial response allows cells to resist additional ER stress. ER stress-induced mitochondrial response is mediated by activation of retrograde (RTG) signaling to enhance anapleurotic reactions of the TCA cycle. Mitochondrial response to ER stress is accompanied by inactivation of the conserved TORC1 pathway, and activation of Snf1/AMPK, the conserved energy sensor and regulator of metabolism. Our results provide new insight into the role of respiration in cell survival in the face of ER stress, and should help in developing therapeutic strategies to limit cell death in disorders linked to ER stress.

Keywords: ER stress; Endoplasmic reticulum; Mitochondria; Yeast

DOI: https://doi.org/10.1242/jcs.241539

Cell Death Dis. 2020 Jan 27. 11(1): 66

Lipid desaturation-associated endoplasmic reticulum stress regulates MYCN gene expression in hepatocellular carcinoma cells.

Xian-Yang Qin, Ting Su, Wenkui Yu, Soichi Kojima.

Hepatocellular carcinoma (HCC) is the second leading cause of cancer-related deaths worldwide due to its high rate of recurrence, in part because of cancer stem cell (CSC)-dependent "field cancerization". Recently, we identified that the oncogene v-myc avian myelocytomatosis viral oncogene neuroblastoma derived homolog (MYCN) marked CSC-like subpopulations in heterogeneous HCC and served as a therapeutic target and prognostic marker for HCC. In this study, we explored the molecular basis of upregulated MYCN gene expression in HCC cells. Liquid chromatograph time-of-flight mass spectrometry-based metabolome analysis demonstrated that the content of unsaturated fatty acids was increased in MYCN high expression (MYCNhigh) CSC-like HCC cells. Inhibition of lipid desaturation using either the chemical inhibitor or siRNA/shRNA against stearoyl-CoA desaturase-1 (SCD1) suppressed cell proliferation as well as MYCN gene expression in MYCNhigh HCC cells, grown as both monolayer and spheres. Further mechanistic study using RNA-seq based transcriptome analysis revealed that endoplasmic reticulum (ER) stress related signaling networks such as endocannabinoid cancer inhibition pathway were under the control of SCD1 in MYCNhigh HCC cells. Furthermore, the expression of ER stress-inducible transcription suppressor cyclic AMP-dependent transcription factor (ATF3) was downregulated in MYCNhigh CSC-like HCC cells and CSC-rich spheroids, which was upregulated by inhibition of lipid desaturation or treatment with acyclic retinoid (ACR). Lipid profiling using NMR spectroscopy revealed that the ACR dramatically reduced the content of unsaturated fatty acids in HCC cells. The chemical inducer of ER stress inhibited MYCN gene expression, while the chemical inhibitor of ER stress or knockdown of ATF3 gene expression partially rescued the suppression of MYCN gene expression by ACR in MYCNhigh HCC cells. These data suggested that lipid desaturation-mediated ER stress signaling regulates MYCN gene expression in HCC cells and serves as a promising therapeutic target for the treatment and prevention of HCC.

DOI: https://doi.org/10.1038/s41419-020-2257-y

J Biol Chem. 2020 Jan 31. pii: jbc.RA119.010589. [Epub ahead of print]

Glycolytic reprogramming of macrophages activated by NOD1 and TLR4 agonists: no association with proinflammatory cytokine production in normoxia.

Nina E Murugina, Anna S Budikhina, Yulia A Dagil, Polina V Maximchik, Lyudmila S Balyasova, Vladimir V Murugin, Mikhail V Melnikov, Viktoriya S Sharova, Anna M Nikolaeva, Georgy Z Chkadua, Boris V Pinegin, Mikhail V Pashenkov.

Upon activation with pathogen-associated molecular patterns, metabolism of macrophages and dendritic cells is shifted from oxidative phosphorylation to aerobic glycolysis, which is considered important for proinflammatory cytokine production. Fragments of bacterial peptidoglycan (muramyl peptides) activate innate immune cells through nucleotide-binding oligomerization domain (NOD) 1 and/or NOD2 receptors. Here, we show that NOD1 and NOD2 agonists induce early glycolytic reprogramming of human monocyte-derived macrophages (MDM), which is similar to that induced by the Toll-like receptor (TLR) 4 agonist, lipopolysaccharide. This glycolytic reprogramming is dependent on Akt kinases, independent of mTOR complex 1, and is efficiently inhibited by 2-deoxy-D-glucose (2-DG) or by glucose starvation. 2-DG inhibits proinflammatory cytokine production by MDM and monocyte-derived dendritic cells activated by NOD1 or TLR4 agonists, except for tumor necrosis factor (TNF) production by MDM, which is inhibited initially, but augmented 4 hours after addition of agonists and later. However, 2-DG exerts these effects by inducing unfolded protein response (UPR) rather than by inhibiting glycolysis. By contrast, glucose starvation does not cause UPR and, in normoxic conditions, only marginally affects proinflammatory cytokine production triggered through NOD1 or TLR4. In hypoxia mimicked by treating MDM with oligomycin (a mitochondrial ATP synthase inhibitor), both 2-DG and glucose starvation strongly suppress TNF and interleukin-6 production, and compromise cell viability. In summary, the requirement of glycolytic reprogramming for proinflammatory cytokine production in normoxia is not obvious, and effects of 2-DG on cytokine responses should be interpreted cautiously. In hypoxia, however, glycolysis becomes critical for cytokine production and cell survival.

Keywords: 2-deoxy-D-glucose; NOD1; NOD2; Nod-like receptor (NLR); cytokine; dendritic cell; glucose metabolism; macrophage; metabolic reprogramming; unfolded protein response (UPR)

DOI: https://doi.org/10.1074/jbc.RA119.010589