bims-unfpre 2024-01-07 papers

bims-unfpre

Biomed News

on Unfolded protein response

Issue of 2024–01–07
eleven papers selected by
Susan Logue, University of Manitoba

UBXN1 maintains ER proteostasis and represses UPR activation by modulating translation.
The IRE1β-mediated unfolded protein response is repressed by the chaperone AGR2 in mucin producing cells.
Activation of goblet-cell stress sensor IRE1β is controlled by the mucin chaperone AGR2.
A beneficial adaptive role for CHOP in driving cell fate selection during ER stress.
Canonical IRE1 function needed to sustain vigorous natural killer cell proliferation during viral infection.
FANNING THE FLAMES OF ER STRESS: Can sphingolipid metabolism be targeted to enhance ER stress associated immunogenic cell death in cancer?
Endoplasmic reticulum stress and its role in various neurodegenerative diseases.
The integrated stress response protects against ER stress but is not required for altered translation and lifespan from dietary restriction in Caenorhabditis elegans.
Changes in tissue protein N-glycosylation and associated molecular signature occur in the human Parkinsonian brain in a region-specific manner.
Methotrimeprazine is a neuroprotective antiviral in JEV infection via adaptive ER stress and autophagy.
Quantitative Measurement of Transthyretin Mistargeting by Proximity Labeling and Parallel Reaction Monitoring.

EMBO Rep. 2024 Jan 02.

UBXN1 maintains ER proteostasis and represses UPR activation by modulating translation.

Brittany A Ahlstedt, Rakesh Ganji, Sirisha Mukkavalli, Joao A Paulo, Steve P Gygi, Malavika Raman.

  ER protein homeostasis (proteostasis) is essential for proper folding and maturation of proteins in the secretory pathway. Loss of ER proteostasis can lead to the accumulation of misfolded or aberrant proteins in the ER and triggers the unfolded protein response (UPR). In this study, we find that the p97 adaptor UBXN1 is an important negative regulator of the UPR. Loss of UBXN1 sensitizes cells to ER stress and activates the UPR. This leads to widespread upregulation of the ER stress transcriptional program. Using comparative, quantitative proteomics we show that deletion of UBXN1 results in a significant enrichment of proteins involved in ER-quality control processes including those involved in protein folding and import. Notably, we find that loss of UBXN1 does not perturb p97-dependent ER-associated degradation (ERAD). Our studies indicate that loss of UBXN1 increases translation in both resting and ER-stressed cells. Surprisingly, this process is independent of p97 function. Taken together, our studies have identified a new role for UBXN1 in repressing translation and maintaining ER proteostasis in a p97 independent manner.

Keywords:  ER Stress; Translation; Ubiquitin; Unfolded Protein Response; p97

DOI:  https://doi.org/10.1038/s44319-023-00027-z
EMBO J. 2023 Dec 18.

The IRE1β-mediated unfolded protein response is repressed by the chaperone AGR2 in mucin producing cells.

Lisa Neidhardt, Eva Cloots, Natalie Friemel, Caroline A M Weiss, Heather P Harding, Stephen H McLaughlin, Sophie Janssens, David Ron.

  Effector mechanisms of the unfolded protein response (UPR) in the endoplasmic reticulum (ER) are well-characterised, but how ER proteostasis is sensed is less well understood. Here, we exploited the beta isoform of the UPR transducer IRE1, that is specific to mucin-producing cells in order to gauge the relative regulatory roles of activating ligands and repressing chaperones of the specialised ER of goblet cells. Replacement of the stress-sensing luminal domain of endogenous IRE1α in CHO cells (normally expressing neither mucin nor IRE1β) with the luminal domain of IRE1β deregulated basal IRE1 activity. The mucin-specific chaperone AGR2 repressed IRE1 activity in cells expressing the domain-swapped IRE1β/α chimera, but had no effect on IRE1α. Introduction of the goblet cell-specific client MUC2 reversed AGR2-mediated repression of the IRE1β/α chimera. In vitro, AGR2 actively de-stabilised the IRE1β luminal domain dimer and formed a reversible complex with the inactive monomer. These features of the IRE1β-AGR2 couple suggest that active repression of IRE1β by a specialised mucin chaperone subordinates IRE1 activity to a proteostatic challenge unique to goblet cells, a challenge that is otherwise poorly recognised by the pervasive UPR transducers.

Keywords:  Endoplasmic Reticulum (ER); Molecular Chaperones; Mucin; Protein Multimerisation; Unfolded Protein Response (UPR)

DOI:  https://doi.org/10.1038/s44318-023-00014-z
EMBO J. 2023 Dec 20.

Activation of goblet-cell stress sensor IRE1β is controlled by the mucin chaperone AGR2.

Eva Cloots, Phaedra Guilbert, Mathias Provost, Lisa Neidhardt, Evelien Van de Velde, Farzaneh Fayazpour, Delphine De Sutter, Savvas N Savvides, Sven Eyckerman, Sophie Janssens.

  Intestinal goblet cells are secretory cells specialized in the production of mucins, and as such are challenged by the need for efficient protein folding. Goblet cells express Inositol-Requiring Enzyme-1β (IRE1β), a unique sensor in the unfolded protein response (UPR), which is part of an adaptive mechanism that regulates the demands of mucin production and secretion. However, how IRE1β activity is tuned to mucus folding load remains unknown. We identified the disulfide isomerase and mucin chaperone AGR2 as a goblet cell-specific protein that crucially regulates IRE1β-, but not IRE1α-mediated signaling. AGR2 binding to IRE1β disrupts IRE1β oligomerization, thereby blocking its downstream endonuclease activity. Depletion of endogenous AGR2 from goblet cells induces spontaneous IRE1β activation, suggesting that alterations in AGR2 availability in the endoplasmic reticulum set the threshold for IRE1β activation. We found that AGR2 mutants lacking their catalytic cysteine, or displaying the disease-associated mutation H117Y, were no longer able to dampen IRE1β activity. Collectively, these results demonstrate that AGR2 is a central chaperone regulating the goblet cell UPR by acting as a rheostat of IRE1β endonuclease activity.

Keywords:  AGR2; Goblet Cells; IRE1β; Mucus Homeostasis; Unfolded Protein Response (UPR)

DOI:  https://doi.org/10.1038/s44318-023-00015-y
EMBO Rep. 2024 Jan 02.

A beneficial adaptive role for CHOP in driving cell fate selection during ER stress.

Kaihua Liu, Chaoxian Zhao, Reed C Adajar, Diane DeZwaan-McCabe, D Thomas Rutkowski.

  Cellular stresses elicit signaling cascades that are capable of either mitigating the inciting dysfunction or initiating cell death. During endoplasmic reticulum (ER) stress, the transcription factor CHOP is widely recognized to promote cell death. However, it is not clear whether CHOP also has a beneficial role during adaptation. Here, we combine a new, versatile, genetically modified Chop allele with single cell analysis and with stresses of physiological intensity, to rigorously examine the contribution of CHOP to cell fate. Paradoxically, we find that CHOP promotes death in some cells, but proliferation-and hence recovery-in others. Strikingly, this function of CHOP confers to cells a stress-specific competitive growth advantage. The dynamics of CHOP expression and UPR activation at the single cell level suggest that CHOP maximizes UPR activation, which in turn favors stress resolution, subsequent UPR deactivation, and proliferation. Taken together, these findings suggest that CHOP's function can be better described as a "stress test" that drives cells into either of two mutually exclusive fates-adaptation or death-during stresses of physiological intensity.

Keywords:  Adaptation; Cell Fate; ER Stress; Cell Proliferation; Unfolded Protein Response

DOI:  https://doi.org/10.1038/s44319-023-00026-0
iScience. 2023 Dec 15. 26(12): 108570

Canonical IRE1 function needed to sustain vigorous natural killer cell proliferation during viral infection.

Jessica Vetters, Mary van Helden, Clint De Nolf, Sofie Rennen, Eva Cloots, Evelien Van De Velde, Farzaneh Fayazpour, Justine Van Moorleghem, Manon Vanheerswynghels, Karl Vergote, Louis Boon, Eric Vivier, Bart N Lambrecht, Sophie Janssens.

  The unfolded protein response (UPR) aims to restore ER homeostasis under conditions of high protein folding load, a function primarily serving secretory cells. Additional, non-canonical UPR functions have recently been unraveled in immune cells. We addressed the function of the inositol-requiring enzyme 1 (IRE1) signaling branch of the UPR in NK cells in homeostasis and microbial challenge. Cell-intrinsic compound deficiency of IRE1 and its downstream transcription factor XBP1 in NKp46+ NK cells, did not affect basal NK cell homeostasis, or overall outcome of viral MCMV infection. However, mixed bone marrow chimeras revealed a competitive advantage in the proliferation of IRE1-sufficient Ly49H+ NK cells after viral infection. CITE-Seq analysis confirmed strong induction of IRE1 early upon infection, concomitant with the activation of a canonical UPR signature. Therefore, we conclude that IRE1/XBP1 activation is required during vigorous NK cell proliferation early upon viral infection, as part of a canonical UPR response.

Keywords:  Immunology; Microbiology; Transcriptomics; Virology

DOI:  https://doi.org/10.1016/j.isci.2023.108570
Mol Pharmacol. 2023 Dec 21. pii: MOLPHARM-AR-2023-000786. [Epub ahead of print]

FANNING THE FLAMES OF ER STRESS: Can sphingolipid metabolism be targeted to enhance ER stress associated immunogenic cell death in cancer?

Jeremy A Hengst, Asvelt J Nduwumwami, Arati Sharma, Jong K Yun.

  The three arms of the unfolded protein response (UPR) surveil the lumenal environment of the endoplasmic reticulum (ER) and transmit information through the lipid bilayer to the cytoplasm to alert the cell of stress conditions within the ER lumen. That same lipid bilayer is the site of de novo synthesis of phospholipids and sphingolipids. Thus, it is no surprise that lipids are modulated by and modulators of ER stress that intersect with the UPR. Given that sphingolipids have both pro-survival and pro-apoptotic effects, they also exert opposing effects on life/death decisions in the face of prolonged ER stress detected by the UPR. In this review, we will focus on several recent studies that demonstrate how sphingolipids affect each arm of the UPR. We will also discuss the role of sphingolipids in the process of immunogenic cell death (ICD) downstream of the protein kinase RNA-like endoplasmic reticulum kinase (PERK)/ eukaryotic initiating factor 2α (eIF2α) arm of the UPR. Furthermore, we will discuss strategies to target the sphingolipid metabolic pathway that could potentially act synergistically with agents that induce ER stress as novel anti-cancer treatments. Significance Statement This review provides the readers with a brief discussion of the sphingolipid metabolic pathway and the unfolded protein response. The primary focus of the review is the mechanism(s) by which sphingolipids modulate the ER stress response pathways and the critical role of sphingolipids in the process of immunogenic cell death associated with the ER stress response.

Keywords:  Ceramide; D-Sphingosine; Endoplasmic reticulum stress; immunogenicity; mechanisms of cell killing/apoptosis; sphingolipids; sphingosine 1-phosphate

DOI:  https://doi.org/10.1124/molpharm.123.000786
Brain Res. 2023 Dec 29. pii: S0006-8993(23)00513-9. [Epub ahead of print]1826 148742

Endoplasmic reticulum stress and its role in various neurodegenerative diseases.

Rimaljot Singh, Navpreet Kaur, Vinay Choubey, Neelima Dhingra, Tanzeer Kaur.

  The Endoplasmic reticulum (ER), a critical cellular organelle, maintains cellular homeostasis by regulating calcium levels and orchestrating essential functions such as protein synthesis, folding, and lipid production. A pivotal aspect of ER function is its role in protein quality control. When misfolded proteins accumulate within the ER due to factors like protein folding chaperone dysfunction, toxicity, oxidative stress, or inflammation, it triggers the Unfolded protein response (UPR). The UPR involves the activation of chaperones like calnexin, calreticulin, glucose-regulating protein 78 (GRP78), and Glucose-regulating protein 94 (GRP94), along with oxidoreductases like protein disulphide isomerases (PDIs). Cells employ the Endoplasmic reticulum-associated degradation (ERAD) mechanism to counteract protein misfolding. ERAD disruption causes the detachment of GRP78 from transmembrane proteins, initiating a cascade involving Inositol-requiring kinase/endoribonuclease 1 (IRE1), Activating transcription factor 6 (ATF6), and Protein kinase RNA-like endoplasmic reticulum kinase (PERK) pathways. The accumulation and deposition of misfolded proteins within the cell are hallmarks of numerous neurodegenerative diseases. These aberrant proteins disrupt normal neuronal signalling and contribute to impaired cellular homeostasis, including oxidative stress and compromised protein degradation pathways. In essence, ER stress is defined as the cellular response to the accumulation of misfolded proteins in the endoplasmic reticulum, encompassing a series of signalling pathways and molecular events that aim to restore cellular homeostasis. This comprehensive review explores ER stress and its profound implications for the pathogenesis and progression of neurodegenerative diseases.

Keywords:  Calcium homeostasis; ER stress; Misfolded proteins; Neurodegeneration; Unfolded protein response

DOI:  https://doi.org/10.1016/j.brainres.2023.148742
Front Cell Dev Biol. 2023 ;11 1263344

The integrated stress response protects against ER stress but is not required for altered translation and lifespan from dietary restriction in Caenorhabditis elegans.

Zhengxin Ma, Jordan Horrocks, Dilawar A Mir, Matthew Cox, Marissa Ruzga, Jarod Rollins, Aric N Rogers.

  The highly conserved integrated stress response (ISR) reduces and redirects mRNA translation in response to certain forms of stress and nutrient limitation. It is activated when kinases phosphorylate a key residue in the alpha subunit of eukaryotic translation initiation factor 2 (eIF2). General Control Nonderepressible-2 (GCN2) is activated to phosphorylate eIF2α by the presence of uncharged tRNA associated with nutrient scarcity, while protein kinase R-like ER kinase-1 (PERK) is activated during the ER unfolded protein response (UPRER). Here, we investigated the role of the ISR during nutrient limitation and ER stress with respect to changes in protein synthesis, translationally driven mRNA turnover, and survival in Caenorhabditis elegans. We found that, while GCN2 phosphorylates eIF2α when nutrients are restricted, the ability to phosphorylate eIF2α is not required for changes in translation, nonsense-mediated decay, or lifespan associated with dietary restriction (DR). Interestingly, loss of both GCN2 and PERK abolishes increased lifespan associated with dietary restriction, indicating the possibility of other substrates for these kinases. The ISR was not dispensable under ER stress conditions, as demonstrated by the requirement for PERK and eIF2α phosphorylation for decreased translation and wild type-like survival. Taken together, results indicate that the ISR is critical for ER stress and that other translation regulatory mechanisms are sufficient for increased lifespan under dietary restriction.

Keywords:  aging; dietary restriction; endoplasmic reticulum stress; integrated stress response; nonsense mediated decay

DOI:  https://doi.org/10.3389/fcell.2023.1263344
PNAS Nexus. 2024 Jan;3(1): pgad439

Changes in tissue protein N-glycosylation and associated molecular signature occur in the human Parkinsonian brain in a region-specific manner.

Ana Lúcia Rebelo, Richard R Drake, Martina Marchetti-Deschmann, Radka Saldova, Abhay Pandit.

  Parkinson's disease (PD) associated state of neuroinflammation due to the aggregation of aberrant proteins is widely reported. One type of post-translational modification involved in protein stability is glycosylation. Here, we aimed to characterize the human Parkinsonian nigro-striatal N-glycome, and related transcriptome/proteome, and its correlation with endoplasmic reticulum (ER) stress and unfolded protein response (UPR), providing a comprehensive characterization of the PD molecular signature. Significant changes were seen upon a PD: a 3% increase in sialylation and 5% increase in fucosylation in both regions, and a 2% increase in oligomannosylated N-glycans in the substantia nigra. In the latter, a decrease in the mRNA expression of sialidases and an upregulation in the UPR pathway were also seen. To show the correlation between these, we also describe a small in vitro study where changes in specific glycosylation trait enzymes (inhibition of sialyltransferases) led to impairments in cell mitochondrial activity, changes in glyco-profile, and upregulation in UPR pathways. This complete characterization of the human nigro-striatal N-glycome provides an insight into the glycomic profile of PD through a transversal approach while combining the other PD "omics" pieces, which can potentially assist in the development of glyco-focused therapeutics.

Keywords:  Biological Sciences; N-glycosylation; Neuroscience; Parkinson's disease; glycomics; human brain; protein glycosylation

DOI:  https://doi.org/10.1093/pnasnexus/pgad439
EMBO Mol Med. 2024 Jan 02.

Methotrimeprazine is a neuroprotective antiviral in JEV infection via adaptive ER stress and autophagy.

Surendra K Prajapat, Laxmi Mishra, Sakshi Khera, Shadrack D Owusu, Kriti Ahuja, Puja Sharma, Eira Choudhary, Simran Chhabra, Niraj Kumar, Rajan Singh, Prem S Kaushal, Dinesh Mahajan, Arup Banerjee, Rajender K Motiani, Sudhanshu Vrati, Manjula Kalia.

  Japanese encephalitis virus (JEV) pathogenesis is driven by a combination of neuronal death and neuroinflammation. We tested 42 FDA-approved drugs that were shown to induce autophagy for antiviral effects. Four drugs were tested in the JE mouse model based on in vitro protective effects on neuronal cell death, inhibition of viral replication, and anti-inflammatory effects. The antipsychotic phenothiazines Methotrimeprazine (MTP) & Trifluoperazine showed a significant survival benefit with reduced virus titers in the brain, prevention of BBB breach, and inhibition of neuroinflammation. Both drugs were potent mTOR-independent autophagy flux inducers. MTP inhibited SERCA channel functioning, and induced an adaptive ER stress response in diverse cell types. Pharmacological rescue of ER stress blocked autophagy and antiviral effect. MTP did not alter translation of viral RNA, but exerted autophagy-dependent antiviral effect by inhibiting JEV replication complexes. Drug-induced autophagy resulted in reduced NLRP3 protein levels, and attenuation of inflammatory cytokine/chemokine release from infected microglial cells. Our study suggests that MTP exerts a combined antiviral and anti-inflammatory effect in JEV infection, and has therapeutic potential for JE treatment.

Keywords:  Antipsychotic; Microglia; Neuroinflammation; Phenothiazines; Trifluoperazine

DOI:  https://doi.org/10.1038/s44321-023-00014-w
Front Chem Biol. 2023 ;pii: 1288188. [Epub ahead of print]2

Quantitative Measurement of Transthyretin Mistargeting by Proximity Labeling and Parallel Reaction Monitoring.

Ziqi Lyu, Joseph C Genereux.

  Proximity labeling is a powerful approach for characterizing subcellular proteomes. We recently demonstrated that proximity labeling can be used to identify mistrafficking of secretory proteins, such as occurs during pre-emptive quality control (pre-QC) following endoplasmic reticulum (ER) stress. This assay depends on protein quantification by immunoblotting and densitometry, which sometimes suffers from poor sensitivity. Here, we integrate parallel reaction monitoring (PRM) mass spectrometry to enable a more quantitative platform, and assess how chemical ER stressors impact pre-QC of the model secretory protein transthyretin in HEK293T cells. We find that some drug treatments affect labeling efficiency, which can be controlled for by normalizing to APEX2 auto-labeling. While some chemical ER stress inducers including Brefeldin A and thapsigargin induce pre-QC, tunicamycin and dithiothreitol do not, indicating ER stress alone is not sufficient. This finding contrasts with the canonical model of pre-QC induction, and establishes the utility of our platform.

Keywords:  APEX2 proximity labeling; endoplasmic reticulum stress; parallel reaction monitoring; pre-emptive quality control; protein mislocalization

DOI:  https://doi.org/10.3389/fchbi.2023.1288188