Mutation in FBXO32 causes dilated cardiomyopathy through up-regulation of ER-stress mediated apoptosis

AbstractEndoplasmic reticulum (ER) stress induction of cell death is implicated in cardiovascular diseases. Sustained activation of ER-stress induces the unfolded protein response (UPR) pathways, which in turn activate three major effector proteins. We previously reported a missense homozygous mutation in FBXO32 (MAFbx, Atrogin-1) causing advanced heart failure by impairing autophagy. In the present study, we performed transcriptional profiling and biochemical assays, which unexpectedly revealed a reduced activation of UPR effectors in patient mutant hearts, while a strong up-regulation of the CHOP transcription factor and of its target genes are observed. Expression of mutant FBXO32 in cells is sufficient to induce CHOP-associated apoptosis, to increase the ATF2 transcription factor and to impair ATF2 ubiquitination. ATF2 protein interacts with FBXO32 in the human heart and its expression is especially high in FBXO32 mutant hearts. These findings provide a new underlying mechanism for FBXO32-mediated cardiomyopathy, implicating abnormal activation of CHOP. These results suggest alternative non-canonical pathways of CHOP activation that could be considered to develop new therapeutic targets for the treatment of FBXO32-associated DCM.

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Effectors Targeting the Unfolded Protein Response during Intracellular Bacterial Infection

Microorganisms ◽

10.3390/microorganisms9040705 ◽

2021 ◽

Vol 9 (4) ◽

pp. 705

Author(s):

Manal H. Alshareef ◽

Elizabeth L. Hartland ◽

Kathleen McCaffrey

Keyword(s):

Bacterial Infection ◽

Er Stress ◽

Unfolded Protein Response ◽

Host Defense ◽

Effector Proteins ◽

Dual Nature ◽

Unfolded Protein ◽

Bacterial Replication ◽

The Unfolded Protein Response ◽

Protein Response

The unfolded protein response (UPR) is a homeostatic response to endoplasmic reticulum (ER) stress within eukaryotic cells. The UPR initiates transcriptional and post-transcriptional programs to resolve ER stress; or, if ER stress is severe or prolonged, initiates apoptosis. ER stress is a common feature of bacterial infection although the role of the UPR in host defense is only beginning to be understood. While the UPR is important for host defense against pore-forming toxins produced by some bacteria, other bacterial effector proteins hijack the UPR through the activity of translocated effector proteins that facilitate intracellular survival and proliferation. UPR-mediated apoptosis can limit bacterial replication but also often contributes to tissue damage and disease. Here, we discuss the dual nature of the UPR during infection and the implications of UPR activation or inhibition for inflammation and immunity as illustrated by different bacterial pathogens.

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Ceapins are a new class of unfolded protein response inhibitors, selectively targeting the ATF6α branch

eLife ◽

10.7554/elife.11878 ◽

2016 ◽

Vol 5 ◽

Cited By ~ 71

Author(s):

Ciara M Gallagher ◽

Carolina Garri ◽

Erica L Cain ◽

Kenny Kean-Hooi Ang ◽

Christopher G Wilson ◽

...

Keyword(s):

Transcription Factor ◽

Er Stress ◽

Unfolded Protein Response ◽

Proteolytic Processing ◽

Unfolded Protein ◽

New Class ◽

Cancer Models ◽

Membrane Bound ◽

The Unfolded Protein Response ◽

Protein Response

The membrane-bound transcription factor ATF6α plays a cytoprotective role in the unfolded protein response (UPR), required for cells to survive ER stress. Activation of ATF6α promotes cell survival in cancer models. We used cell-based screens to discover and develop Ceapins, a class of pyrazole amides, that block ATF6α signaling in response to ER stress. Ceapins sensitize cells to ER stress without impacting viability of unstressed cells. Ceapins are highly specific inhibitors of ATF6α signaling, not affecting signaling through the other branches of the UPR, or proteolytic processing of its close homolog ATF6β or SREBP (a cholesterol-regulated transcription factor), both activated by the same proteases. Ceapins are first-in-class inhibitors that can be used to explore both the mechanism of activation of ATF6α and its role in pathological settings. The discovery of Ceapins now enables pharmacological modulation all three UPR branches either singly or in combination.

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Activation of the ATF6 branch of the unfolded protein response in neurons improves stroke outcome

Journal of Cerebral Blood Flow & Metabolism ◽

10.1177/0271678x16650218 ◽

2016 ◽

Vol 37 (3) ◽

pp. 1069-1079 ◽

Cited By ~ 33

Author(s):

Zhui Yu ◽

Huaxin Sheng ◽

Shuai Liu ◽

Shengli Zhao ◽

Christopher C Glembotski ◽

...

Keyword(s):

Transcription Factor ◽

Er Stress ◽

Unfolded Protein Response ◽

Short Form ◽

Infarct Volume ◽

Stroke Outcome ◽

Early Reperfusion ◽

Unfolded Protein ◽

The Unfolded Protein Response ◽

Protein Response

Impaired function of the endoplasmic reticulum (ER stress) is a hallmark of many human diseases including stroke. To restore ER function in stressed cells, the unfolded protein response (UPR) is induced, which activates 3 ER stress sensor proteins including activating transcription factor 6 (ATF6). ATF6 is then cleaved by proteases to form the short-form ATF6 (sATF6), a transcription factor. To determine the extent to which activation of the ATF6 UPR branch defines the fate and function of neurons after stroke, we generated a conditional and tamoxifen-inducible sATF6 knock-in mouse. To express sATF6 in forebrain neurons, we crossed our sATF6 knock-in mouse line with Emx1-Cre mice to generate ATF6-KI mice. After the ATF6 branch was activated in ATF6-KI mice with tamoxifen, mice were subjected to transient middle cerebral artery occlusion. Forced activation of the ATF6 UPR branch reduced infarct volume and improved functional outcome at 24 h after stroke. Increased autophagic activity at early reperfusion time after stroke may contribute to the ATF6-mediated neuroprotection. We concluded that the ATF6 UPR branch is crucial to ischemic stroke outcome. Therefore, boosting UPR pro-survival pathways may be a promising therapeutic strategy for stroke.

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Endoplasmic reticulum stress regulation of the Kar2p/BiP chaperone alleviates proteotoxicity via dual degradation pathways

Molecular Biology of the Cell ◽

10.1091/mbc.e11-04-0297 ◽

2012 ◽

Vol 23 (4) ◽

pp. 630-641 ◽

Cited By ~ 6

Author(s):

Chia-Ling Hsu ◽

Rupali Prasad ◽

Christie Blackman ◽

Davis T. W. Ng

Keyword(s):

Endoplasmic Reticulum ◽

Er Stress ◽

Target Genes ◽

Regulatory Element ◽

Unfolded Protein ◽

Single Target ◽

Transcriptional Regulatory ◽

The Unfolded Protein Response ◽

Protein Response ◽

Stress Activation

The unfolded protein response (UPR) monitors and maintains protein homeostasis in the endoplasmic reticulum (ER). In budding yeast, the UPR is a transcriptional regulatory pathway that is quiescent under normal conditions. Under conditions of acute ER stress, activation of UPR targets is essential for cell viability. How individual target genes contribute to stress tolerance is unclear. Uncovering these roles is hampered because most targets also play important functions in the absence of stress. To differentiate stress-specific roles from everyday functions, a single target gene was uncoupled from UPR control by eliminating its UPR-specific regulatory element. Through this approach, the UPR remains intact, aside from its inability to induce the designated target. Applying the strategy to the major ER chaperone Kar2p/BiP revealed the physiological function of increasing its cellular concentration. Despite hundreds of target genes under UPR control, we show that activation of KAR2 is indispensable to alleviate some forms of ER stress. Specifically, activation is essential to dispose misfolded proteins that are otherwise toxic. Surprisingly, induced BiP/Kar2p molecules are dedicated to alleviating stress. The inability to induce KAR2 under stress had no effect on its known housekeeping functions.

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Specificity in endoplasmic reticulum-stress signaling in yeast entails a step-wise engagement of HAC1 mRNA to clusters of the stress sensor Ire1

eLife ◽

10.7554/elife.05031 ◽

2014 ◽

Vol 3 ◽

Cited By ~ 23

Author(s):

Eelco van Anken ◽

David Pincus ◽

Scott Coyle ◽

Tomás Aragón ◽

Christof Osman ◽

...

Keyword(s):

Endoplasmic Reticulum ◽

Er Stress ◽

Target Genes ◽

Transmembrane Domain ◽

Transcription Activator ◽

Stress Sensor ◽

Unfolded Protein ◽

Oligomeric Assembly ◽

The Unfolded Protein Response ◽

Linker Domain

Insufficient protein-folding capacity in the endoplasmic reticulum (ER) induces the unfolded protein response (UPR). In the ER lumen, accumulation of unfolded proteins activates the transmembrane ER-stress sensor Ire1 and drives its oligomerization. In the cytosol, Ire1 recruits HAC1 mRNA, mediating its non-conventional splicing. The spliced mRNA is translated into Hac1, the key transcription activator of UPR target genes that mitigate ER-stress. In this study, we report that oligomeric assembly of the ER-lumenal domain is sufficient to drive Ire1 clustering. Clustering facilitates Ire1's cytosolic oligomeric assembly and HAC1 mRNA docking onto a positively charged motif in Ire1's cytosolic linker domain that tethers the kinase/RNase to the transmembrane domain. By the use of a synthetic bypass, we demonstrate that mRNA docking per se is a pre-requisite for initiating Ire1's RNase activity and, hence, splicing. We posit that such step-wise engagement between Ire1 and its mRNA substrate contributes to selectivity and efficiency in UPR signaling.

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Transcription factor ATF4 directs basal and stress-induced gene expression in the unfolded protein response and cholesterol metabolism in the liver

Molecular Biology of the Cell ◽

10.1091/mbc.e16-01-0039 ◽

2016 ◽

Vol 27 (9) ◽

pp. 1536-1551 ◽

Cited By ~ 72

Author(s):

Michael E. Fusakio ◽

Jeffrey A. Willy ◽

Yongping Wang ◽

Emily T. Mirek ◽

Rana J. T. Al Baghdadi ◽

...

Keyword(s):

Oxidative Stress ◽

Gene Expression ◽

Transcription Factor ◽

Er Stress ◽

Unfolded Protein Response ◽

Cholesterol Metabolism ◽

Unfolded Protein ◽

Expression Of Genes ◽

The Unfolded Protein Response ◽

Protein Response

Disturbances in protein folding and membrane compositions in the endoplasmic reticulum (ER) elicit the unfolded protein response (UPR). Each of three UPR sensory proteins—PERK (PEK/EIF2AK3), IRE1, and ATF6—is activated by ER stress. PERK phosphorylation of eIF2 represses global protein synthesis, lowering influx of nascent polypeptides into the stressed ER, coincident with preferential translation of ATF4 (CREB2). In cultured cells, ATF4 induces transcriptional expression of genes directed by the PERK arm of the UPR, including genes involved in amino acid metabolism, resistance to oxidative stress, and the proapoptotic transcription factor CHOP (GADD153/DDIT3). In this study, we characterize whole-body and tissue-specific ATF4-knockout mice and show in liver exposed to ER stress that ATF4 is not required for CHOP expression, but instead ATF6 is a primary inducer. RNA-Seq analysis indicates that ATF4 is responsible for a small portion of the PERK-dependent UPR genes and reveals a requirement for expression of ATF4 for expression of genes involved in oxidative stress response basally and cholesterol metabolism both basally and under stress. Consistent with this pattern of gene expression, loss of ATF4 resulted in enhanced oxidative damage, and increased free cholesterol in liver under stress accompanied by lowered cholesterol in sera.

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Low-Dose Radiation Suppresses Pokemon Expression under Hypoxic Conditions

Zeitschrift für Naturforschung C ◽

10.5560/znc.2013-0035 ◽

2014 ◽

Vol 69 (1-2) ◽

pp. 68-74

Author(s):

Seung-Whan Kim ◽

Kweon Yu ◽

Kee-Sun Shin ◽

Kisang Kwon ◽

Tae-Sik Hwang ◽

...

Keyword(s):

Transcription Factor ◽

Er Stress ◽

Low Dose ◽

Differentially Expressed Gene ◽

Low Dose Radiation ◽

Unfolded Protein ◽

Hypoxic Conditions ◽

The Unfolded Protein Response ◽

Protein Response ◽

Dose Radiation

Our previous data demonstrated that CoCl2-induced hypoxia controls endoplasmic reticulum (ER) stress-associated and other intracellular factors. One of them, the transcription factor Pokemon, was differentially regulated by low-dose radiation (LDR). There are limited data regarding how this transcription factor is involved in expression of the unfolded protein response (UPR) under hypoxic conditions. The purpose of this study was to obtain clues on how Pokemon is involved in the UPR. Pokemon was selected as a differentially expressed gene under hypoxic conditions; however, its regulation was clearly repressed by LDR. It was also demonstrated that both expression of ER chaperones and ER stress sensors were affected by hypoxic conditions, and the same results were obtained when cells in which Pokemon was up- or down-regulated were used. The current state of UPR and LDR research associated with the Pokemon pathway offers an important opportunity to understand the oncogenesis, senescence, and differentiation of cells, as well as to facilitate introduction of new therapeutic radiopharmaceuticals

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Coxsackievirus B3 Infection Activates the Unfolded Protein Response and Induces Apoptosis through Downregulation of p58IPK and Activation of CHOP and SREBP1

Journal of Virology ◽

10.1128/jvi.01416-09 ◽

2010 ◽

Vol 84 (17) ◽

pp. 8446-8459 ◽

Cited By ~ 56

Author(s):

Huifang M. Zhang ◽

Xin Ye ◽

Yue Su ◽

Ji Yuan ◽

Zhen Liu ◽

...

Keyword(s):

Transcription Factor ◽

Er Stress ◽

Unfolded Protein Response ◽

Hela Cells ◽

Coxsackievirus B3 ◽

Unfolded Protein ◽

Caspase 12 ◽

Cvb3 Infection ◽

The Unfolded Protein Response ◽

Protein Response

ABSTRACT Cardiomyocyte apoptosis is a hallmark of coxsackievirus B3 (CVB3)-induced myocarditis. We used cardiomyocytes and HeLa cells to explore the cellular response to CVB3 infection, with a focus on pathways leading to apoptosis. CVB3 infection triggered endoplasmic reticulum (ER) stress and differentially regulated the three arms of the unfolded protein response (UPR) initiated by the proximal ER stress sensors ATF6a (activating transcription factor 6a), IRE1-XBP1 (X box binding protein 1), and PERK (PKR-like ER protein kinase). Upon CVB3 infection, glucose-regulated protein 78 expression was upregulated, and in turn ATF6a and XBP1 were activated via protein cleavage and mRNA splicing, respectively. UPR activity was further confirmed by the enhanced expression of UPR target genes ERdj4 and EDEM1. Surprisingly, another UPR-associated gene, p58IPK, which often is upregulated during infections with other types of viruses, was downregulated at both mRNA and protein levels after CVB3 infection. These findings were observed similarly for uninfected Tet-On HeLa cells induced to overexpress ATF6a or XBP1. In exploring potential connections between the three UPR pathways, we found that the ATF6a-induced downregulation of p58IPK was associated with the activation of PKR (PERK) and the phosphorylation of eIF2α, suggesting that p58IPK, a negative regulator of PERK and PKR, mediates cross-talk between the ATF6a/IRE1-XBP1 and PERK arms. Finally, we found that CVB3 infection eventually produced the induction of the proapoptoic transcription factor CHOP and the activation of SREBP1 and caspase-12. Taken together, these data suggest that CVB3 infection activates UPR pathways and induces ER stress-mediated apoptosis through the suppression of P58IPK and induction/activation of CHOP, SREBP1, and caspase-12.

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Endoplasmic Reticulum Stress-induced mRNA Splicing Permits Synthesis of Transcription Factor Hac1p/Ern4p That Activates the Unfolded Protein Response

Molecular Biology of the Cell ◽

10.1091/mbc.8.10.1845 ◽

1997 ◽

Vol 8 (10) ◽

pp. 1845-1862 ◽

Cited By ~ 202

Author(s):

Tetsushi Kawahara ◽

Hideki Yanagi ◽

Takashi Yura ◽

Kazutoshi Mori

Keyword(s):

Endoplasmic Reticulum ◽

Transcription Factor ◽

Er Stress ◽

Unfolded Protein Response ◽

Intracellular Signaling ◽

Mrna Splicing ◽

Unfolded Protein ◽

The Unfolded Protein Response ◽

Protein Response ◽

Precursor Mrna

An intracellular signaling from the endoplasmic reticulum (ER) to the nucleus, called the unfolded protein response (UPR), is activated when unfolded proteins are accumulated in the ER under a variety of stress conditions (“ER stress”). We and others recently identified Hac1p/Ern4p as a transcription factor responsible for the UPR inSaccharomyces cerevisiae. It was further reported that Hac1p (238 aa) is detected only in ER-stressed cells, and its expression is mediated by unconventional splicing ofHAC1 precursor mRNA. The splicing replaces the C-terminal portion of Hac1p; it was proposed that precursor mRNA is also translated but the putative product of 230 aa is rapidly degraded by the ubiquitin–proteasome pathway. We have identified and characterized the same regulated splicing and confirmed its essential features. Contrary to the above proposal, however, we find that the 238-aa product of mature mRNA and the 230-aa-type protein tested are highly unstable with little or no difference in stability. Furthermore, we demonstrate that the absence of Hac1p in unstressed cells is due to the lack of translation of precursor mRNA. We conclude that Hac1p is synthesized as the result of ER stress-induced mRNA splicing, leading to activation of the UPR.

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IRE1 and efferent signaling from the endoplasmic reticulum

Journal of Cell Science ◽

10.1242/jcs.113.21.3697 ◽

2000 ◽

Vol 113 (21) ◽

pp. 3697-3702 ◽

Cited By ~ 4

Author(s):

F. Urano ◽

A. Bertolotti ◽

D. Ron

Keyword(s):

Gene Expression ◽

Endoplasmic Reticulum ◽

Transcription Factor ◽

Er Stress ◽

Signaling Pathway ◽

Unfolded Protein ◽

Genes Encoding ◽

The Unfolded Protein Response ◽

Protein Response ◽

Stress Signaling Pathway

Genetic analysis of the cellular adaptation to malfolded proteins in the endoplasmic reticulum (the unfolded protein response - UPR) has revealed a novel signaling pathway initiated by activation of IRE1, an ER-resident protein kinase and endonuclease. In yeast, Ire1p activates gene expression by promoting a non-conventional splicing event that converts the mRNA encoding the Hac1p transcription factor from an inefficiently translated inactive mRNA to an actively translated one. Hac1p binds to the promoters of genes encoding chaperones and other targets of the UPR and activates them. Recently, mammalian IRE1 homologues have been identified and their response to ER stress is regulated by binding to the ER chaperone BiP. The mechanisms by which mammalian IRE1 activates gene expression have not been completely characterized and mammalian HAC1 homologues have not been identified. Surprisingly, mammalian IRE1s are able to activate both JUN N-terminal kinases and an alternative ER-stress signaling pathway mediated by the transcription factor ATF6. This indicates that the mammalian UPR is more complex than that found in yeast.

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