Small molecule proteostasis regulators that reprogram the ER to reduce extracellular protein aggregation

Imbalances in endoplasmic reticulum (ER) proteostasis are associated with etiologically-diverse degenerative diseases linked to excessive extracellular protein misfolding and aggregation. Reprogramming of the ER proteostasis environment through genetic activation of the Unfolded Protein Response (UPR)-associated transcription factor ATF6 attenuates secretion and extracellular aggregation of amyloidogenic proteins. Here, we employed a screening approach that included complementary arm-specific UPR reporters and medium-throughput transcriptional profiling to identify non-toxic small molecules that phenocopy the ATF6-mediated reprogramming of the ER proteostasis environment. The ER reprogramming afforded by our molecules requires activation of endogenous ATF6 and occurs independent of global ER stress. Furthermore, our molecules phenocopy the ability of genetic ATF6 activation to selectively reduce secretion and extracellular aggregation of amyloidogenic proteins. These results show that small molecule-dependent ER reprogramming, achieved through preferential activation of the ATF6 transcriptional program, is a promising strategy to ameliorate imbalances in ER function associated with degenerative protein aggregation diseases.

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Suppression of Mouse AApoAII Amyloidosis Progression by Daily Supplementation with Oxidative Stress Inhibitors

Oxidative Medicine and Cellular Longevity ◽

10.1155/2019/1263274 ◽

2019 ◽

Vol 2019 ◽

pp. 1-14

Author(s):

Jian Dai ◽

Xin Ding ◽

Hiroki Miyahara ◽

Zhe Xu ◽

Xiaoran Cui ◽

...

Keyword(s):

Oxidative Stress ◽

Protein Misfolding ◽

Amyloid Fibrils ◽

Treatment Strategies ◽

Amyloid Deposition ◽

Unfolded Protein ◽

Antioxidative Effects ◽

The Unfolded Protein Response ◽

Protein Response ◽

Protein Misfolding And Aggregation

Amyloidosis is a group of diseases characterized by protein misfolding and aggregation to form amyloid fibrils and subsequent deposition within various tissues. Previous studies have indicated that amyloidosis is often associated with oxidative stress. However, it is not clear whether oxidative stress is involved in the progression of amyloidosis. We administered the oxidative stress inhibitors tempol and apocynin via drinking water to the R1.P1-Apoa2c mouse strain induced to develop mouse apolipoprotein A-II (AApoAII) amyloidosis and found that treatment with oxidative stress inhibitors led to reduction in AApoAII amyloidosis progression compared to an untreated group after 12 weeks, especially in the skin, stomach, and liver. There was no effect on ApoA-II plasma levels or expression of Apoa2 mRNA. Detection of the lipid peroxidation markers 4-hydroxynonenal (4-HNE) and malondialdehyde (MDA) revealed that the antioxidative effects of the treatments were most obvious in the skin, stomach, and liver, which contained higher levels of basal oxidative stress. Moreover, the unfolded protein response was reduced in the liver and was associated with a decrease in oxidative stress and amyloid deposition. These results suggest that antioxidants can suppress the progression of AApoAII amyloid deposition in the improved microenvironment of tissues and that the effect may be related to the levels of oxidative stress in local tissues. This finding provides insights for antioxidative stress treatment strategies for amyloidosis.

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ER stress causes widespread protein aggregation and prion formation

The Journal of Cell Biology ◽

10.1083/jcb.201612165 ◽

2017 ◽

Vol 216 (8) ◽

pp. 2295-2304 ◽

Cited By ~ 28

Author(s):

Norfadilah Hamdan ◽

Paraskevi Kritsiligkou ◽

Chris M. Grant

Keyword(s):

Protein Aggregation ◽

Er Stress ◽

Secretory Pathway ◽

Cellular Protein ◽

Protein Homeostasis ◽

Er Quality Control ◽

Unfolded Protein ◽

The Unfolded Protein Response ◽

Protein Response ◽

Er Homeostasis

Disturbances in endoplasmic reticulum (ER) homeostasis create a condition termed ER stress. This activates the unfolded protein response (UPR), which alters the expression of many genes involved in ER quality control. We show here that ER stress causes the aggregation of proteins, most of which are not ER or secretory pathway proteins. Proteomic analysis of the aggregated proteins revealed enrichment for intrinsically aggregation-prone proteins rather than proteins which are affected in a stress-specific manner. Aggregation does not arise because of overwhelming proteasome-mediated degradation but because of a general disruption of cellular protein homeostasis. We further show that overexpression of certain chaperones abrogates protein aggregation and protects a UPR mutant against ER stress conditions. The onset of ER stress is known to correlate with various disease processes, and our data indicate that widespread amorphous and amyloid protein aggregation is an unanticipated outcome of such stress.

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Beyond the cell factory: Homeostatic regulation of and by the UPRER

Science Advances ◽

10.1126/sciadv.abb9614 ◽

2020 ◽

Vol 6 (29) ◽

pp. eabb9614

Author(s):

Melissa G. Metcalf ◽

Ryo Higuchi-Sanabria ◽

Gilberto Garcia ◽

C. Kimberly Tsui ◽

Andrew Dillin

Keyword(s):

Protein Misfolding ◽

Lipid Synthesis ◽

Transcriptional Response ◽

Cell Factory ◽

Unfolded Protein ◽

Membrane Bound ◽

Emerging Themes ◽

The Unfolded Protein Response ◽

Protein Response ◽

Cellular Components

The endoplasmic reticulum (ER) is commonly referred to as the factory of the cell, as it is responsible for a large amount of protein and lipid synthesis. As a membrane-bound organelle, the ER has a distinct environment that is ideal for its functions in synthesizing these primary cellular components. Many different quality control machineries exist to maintain ER stability under the stresses associated with synthesizing, folding, and modifying complex proteins and lipids. The best understood of these mechanisms is the unfolded protein response of the ER (UPRER), in which transmembrane proteins serve as sensors, which trigger a coordinated transcriptional response of genes dedicated for mitigating the stress. As the name suggests, the UPRER is most well described as a functional response to protein misfolding stress. Here, we focus on recent findings and emerging themes in additional roles of the UPRER outside of protein homeostasis, including lipid homeostasis, autophagy, apoptosis, and immunity.

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Protein Misfolding Induces Hypoxic Preconditioning via a Subset of the Unfolded Protein Response Machinery

Molecular and Cellular Biology ◽

10.1128/mcb.00922-10 ◽

2010 ◽

Vol 30 (21) ◽

pp. 5033-5042 ◽

Cited By ~ 39

Author(s):

Xianrong R. Mao ◽

C. Michael Crowder

Keyword(s):

Unfolded Protein Response ◽

Protein Misfolding ◽

Transcriptional Response ◽

Hypoxic Preconditioning ◽

Misfolded Proteins ◽

Hypoxic Exposure ◽

Hypoxic Injury ◽

Unfolded Protein ◽

The Unfolded Protein Response ◽

Protein Response

ABSTRACT Prolonged cellular hypoxia results in energy failure and ultimately cell death. However, less-severe hypoxia can induce a cytoprotective response termed hypoxic preconditioning (HP). The unfolded protein response pathway (UPR) has been known for some time to respond to hypoxia and regulate hypoxic sensitivity; however, the role of the UPR, if any, in HP essentially has been unexplored. We have shown previously that a sublethal hypoxic exposure of the nematode Caenorhabditis elegans induces a protein chaperone component of the UPR (L. L. Anderson, X. Mao, B. A. Scott, and C. M. Crowder, Science 323:630-633, 2009). Here, we show that HP induces the UPR and that the pharmacological induction of misfolded proteins is itself sufficient to stimulate a delayed protective response to hypoxic injury that requires the UPR pathway proteins IRE-1, XBP-1, and ATF-6. HP also required IRE-1 but not XBP-1 or ATF-6; instead, GCN-2, which is known to suppress translation and induce an adaptive transcriptional response under conditions of UPR activation or amino acid deprivation, was required for HP. The phosphorylation of the translation factor eIF2α, an established mechanism of GCN-2-mediated translational suppression, was not necessary for HP. These data suggest a model where hypoxia-induced misfolded proteins trigger the activation of IRE-1, which along with GCN-2 controls an adaptive response that is essential to HP.

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ER stress and the unfolded protein response in intestinal inflammation

AJP Gastrointestinal and Liver Physiology ◽

10.1152/ajpgi.00063.2010 ◽

2010 ◽

Vol 298 (6) ◽

pp. G820-G832 ◽

Cited By ~ 107

Author(s):

Michael A. McGuckin ◽

Rajaraman D. Eri ◽

Indrajit Das ◽

Rohan Lourie ◽

Timothy H. Florin

Keyword(s):

Er Stress ◽

Unfolded Protein Response ◽

Protein Misfolding ◽

Intestinal Inflammation ◽

Secretory Cells ◽

Unfolded Protein ◽

Bowel Diseases ◽

Stressed Cells ◽

The Unfolded Protein Response ◽

Protein Response

Endoplasmic reticulum (ER) stress is a phenomenon that occurs when excessive protein misfolding occurs during biosynthesis. ER stress triggers a series of signaling and transcriptional events known as the unfolded protein response (UPR). The UPR attempts to restore homeostasis in the ER but if unsuccessful can trigger apoptosis in the stressed cells and local inflammation. Intestinal secretory cells are susceptible to ER stress because they produce large amounts of complex proteins for secretion, most of which are involved in mucosal defense. This review focuses on ER stress in intestinal secretory cells and describes how increased protein misfolding could occur in these cells, the process of degradation of misfolded proteins, the major molecular elements of the UPR pathway, and links between the UPR and inflammation. Evidence is reviewed from mouse models and human inflammatory bowel diseases that ties ER stress and activation of the UPR with intestinal inflammation, and possible therapeutic approaches to ameliorate ER stress are discussed.

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The Unfolded Protein Response and Chemical Chaperones Reduce Protein Misfolding and Colitis in Mice

Gastroenterology ◽

10.1053/j.gastro.2013.01.023 ◽

2013 ◽

Vol 144 (5) ◽

pp. 989-1000.e6 ◽

Cited By ~ 111

Author(s):

Stewart Siyan Cao ◽

Ellen M. Zimmermann ◽

Brandy–Mengchieh Chuang ◽

Benbo Song ◽

Anosike Nwokoye ◽

...

Keyword(s):

Unfolded Protein Response ◽

Protein Misfolding ◽

Chemical Chaperones ◽

Unfolded Protein ◽

The Unfolded Protein Response ◽

Protein Response

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Age-related decline of the unfolded protein response in the heart promotes protein misfolding and cardiac pathology

10.1101/2021.06.16.448596 ◽

2021 ◽

Author(s):

Christoph Hofmann ◽

Erik A Blackwood ◽

Tobias Jakobi ◽

Clara Sandmann ◽

Julia Groß ◽

...

Keyword(s):

Heart Failure ◽

Unfolded Protein Response ◽

Cardiac Myocytes ◽

Protein Misfolding ◽

Cardiac Myocyte ◽

Myocardial Dysfunction ◽

Disease Process ◽

Unfolded Protein ◽

The Unfolded Protein Response ◽

Protein Response

Cardiac myocyte death during heart failure is particularly detrimental, given that cardiac muscle exhibits limited regenerative potential. Protein aggregation was previously observed in end-stage heart failure, suggesting protein-misfolding in cardiac myocytes as a contributor to the disease process. However, the relationship between protein-misfolding, cardiac myocyte death, and myocardial dysfunction is yet to be clearly established. Here, we showed that protein synthesis and the unfolded protein response (UPR) declined as a function of mammalian postnatal development, especially in tissues with low mitotic activity, such as the heart. A deeper examination in animals models showed that compared to neonatal cardiac myocytes, adult cardiac myocytes expressed lower levels of the adaptive UPR transcription factor, ATF6, as well as lower levels of numerous ATF6-regulated genes, which was associated with susceptibility to ER stress-induced cell death. Further reduction of the ATF6-dependent gene program in ATF6 knock-out mice led to the accumulation of misfolded proteins in the myocardium and impaired myocardial function in response to cardiac stress, indicating that ATF6 plays a critical adaptive role in the setting of cardiac disease. Thus, strategies to increase ATF6 aimed at balancing proteostasis in cardiac myocytes might be a fruitful avenue for the development of novel therapies for heart disease and other age-associated diseases.

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Induction of the Unfolded Protein Response but Normal Basal Granulopoiesis in Mice Expressing G192X ELA2

Blood ◽

10.1182/blood.v112.11.314.314 ◽

2008 ◽

Vol 112 (11) ◽

pp. 314-314

Author(s):

Mark Murakami ◽

Jill Woloszynek ◽

Jun Xia ◽

Fulu Liu ◽

Daniel Link

Keyword(s):

Unfolded Protein Response ◽

Protein Misfolding ◽

Embryonic Stem ◽

Clinical Samples ◽

Developmental Defect ◽

Granulocytic Differentiation ◽

Transgenic Mouse Line ◽

Unfolded Protein ◽

The Unfolded Protein Response ◽

Protein Response

Abstract Severe congenital neutropenia (SCN) is an inborn disorder of granulopoiesis characterized by chronic neutropenia, a block in granulocytic differentiation at the promyelocyte/myelocyte stage, and a marked propensity to develop acute myeloid leukemia. Most cases of SCN are associated with germline heterozygous mutations of ELA2, encoding neutrophil elastase (NE). To date, 59 different, mostly missense, mutations of ELA2 have been reported. A unifying mechanism by which all of the different ELA2 mutants disrupt granulopoiesis is lacking. We and others previously proposed a model in which the ELA2 mutations result in NE protein misfolding, induction of the unfolded protein response (UPR), and ultimately apoptosis of granulocytic precursors. Testing this (and other) models has been limited by the rarity of SCN and difficulty in obtaining clinical samples for testing. Herein, we report the preliminary description of a novel transgenic mouse line that expresses G192X Ela2, reproducing the G193X ELA2 mutation found in some patients with SCN. The G192X mutation was introduced into the murine Ela2 locus by homologous recombination in embryonic stem cells. Heterozygous or homozygous G192 Ela2 “knock-in” mice were healthy with no apparent developmental defect. While expression of Ela2 mRNA was normal, no mature NE protein was detected in the neutrophils of homozygous G192X Ela2 mice. However, in granulocytic precursors (mainly promyelocytes/myelocytes) a small amount of heavily glycosylated mutant NE protein was detected. Together, these observations suggest that G192X NE protein is retained in the endoplasmic reticulum (ER) and rapidly degraded. Consistent with ER stress and induction of the UPR, a significant increase in BiP/GRP78 and ATF6 mRNA expression in mutant granulocytic precursors were observed. Surprisingly, G192X Ela2 mice have normal basal granulopoiesis. The number of circulating neutrophils, granulocytic differentiation in the bone marrow, and number and cytokine responsiveness of myeloid progenitors were comparable to wild type mice. In summary, the G192X Ela2 mice appear to reproduce the NE protein misfolding and UPR activation observed in human SCN granulocytic precursors. However, expression of G192X Ela2 is not sufficient to disrupt basal granulopoiesis in mice. Studies of stress granulopoiesis are underway.

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