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1.
J Virol ; 96(1): e0169521, 2022 01 12.
Article in English | MEDLINE | ID: covidwho-1816694

ABSTRACT

The replication of coronaviruses, including severe acute respiratory syndrome coronavirus (SARS-CoV), Middle East respiratory syndrome coronavirus (MERS-CoV), and the recently emerged severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), is closely associated with the endoplasmic reticulum (ER) of infected cells. The unfolded protein response (UPR), which is mediated by ER stress (ERS), is a typical outcome in coronavirus-infected cells and is closely associated with the characteristics of coronaviruses. However, the interaction between virus-induced ERS and coronavirus replication is poorly understood. Here, we demonstrate that infection with the betacoronavirus porcine hemagglutinating encephalomyelitis virus (PHEV) induced ERS and triggered all three branches of the UPR signaling pathway both in vitro and in vivo. In addition, ERS suppressed PHEV replication in mouse neuro-2a (N2a) cells primarily by activating the protein kinase R-like ER kinase (PERK)-eukaryotic initiation factor 2α (eIF2α) axis of the UPR. Moreover, another eIF2α phosphorylation kinase, interferon (IFN)-induced double-stranded RNA-dependent protein kinase (PKR), was also activated and acted cooperatively with PERK to decrease PHEV replication. Furthermore, we demonstrate that the PERK/PKR-eIF2α pathways negatively regulated PHEV replication by attenuating global protein translation. Phosphorylated eIF2α also promoted the formation of stress granules (SGs), which in turn repressed PHEV replication. In summary, our study presents a vital aspect of the host innate response to invading pathogens and reveals attractive host targets (e.g., PERK, PKR, and eIF2α) for antiviral drugs. IMPORTANCE Coronavirus diseases are caused by different coronaviruses of importance in humans and animals, and specific treatments are extremely limited. ERS, which can activate the UPR to modulate viral replication and the host innate response, is a frequent occurrence in coronavirus-infected cells. PHEV, a neurotropic betacoronavirus, causes nerve cell damage, which accounts for the high mortality rates in suckling piglets. However, it remains incompletely understood whether the highly developed ER in nerve cells plays an antiviral role in ERS and how ERS regulates viral proliferation. In this study, we found that PHEV infection induced ERS and activated the UPR both in vitro and in vivo and that the activated PERK/PKR-eIF2α axis inhibited PHEV replication through attenuating global protein translation and promoting SG formation. A better understanding of coronavirus-induced ERS and UPR activation may reveal the pathogenic mechanism of coronavirus and facilitate the development of new treatment strategies for these diseases.


Subject(s)
Betacoronavirus 1/physiology , Coronavirus Infections/metabolism , Eukaryotic Initiation Factor-2/metabolism , Virus Replication/physiology , eIF-2 Kinase/metabolism , Animals , Betacoronavirus 1/metabolism , Cell Line , Coronavirus Infections/virology , Endoplasmic Reticulum/metabolism , Endoplasmic Reticulum/ultrastructure , Endoplasmic Reticulum Stress , Mice , Phosphorylation , Protein Biosynthesis , Signal Transduction , Unfolded Protein Response
2.
Sci Adv ; 8(8): eabi6110, 2022 Feb 25.
Article in English | MEDLINE | ID: covidwho-1714330

ABSTRACT

The spread of SARS-CoV-2 and ongoing COVID-19 pandemic underscores the need for new treatments. Here we report that cannabidiol (CBD) inhibits infection of SARS-CoV-2 in cells and mice. CBD and its metabolite 7-OH-CBD, but not THC or other congeneric cannabinoids tested, potently block SARS-CoV-2 replication in lung epithelial cells. CBD acts after viral entry, inhibiting viral gene expression and reversing many effects of SARS-CoV-2 on host gene transcription. CBD inhibits SARS-CoV-2 replication in part by up-regulating the host IRE1α RNase endoplasmic reticulum (ER) stress response and interferon signaling pathways. In matched groups of human patients from the National COVID Cohort Collaborative, CBD (100 mg/ml oral solution per medical records) had a significant negative association with positive SARS-CoV-2 tests. This study highlights CBD as a potential preventative agent for early-stage SARS-CoV-2 infection and merits future clinical trials. We caution against use of non-medical formulations including edibles, inhalants or topicals as a preventative or treatment therapy at the present time.


Subject(s)
Antiviral Agents/pharmacology , Cannabidiol/pharmacology , Host-Pathogen Interactions/drug effects , Immunity, Innate/drug effects , SARS-CoV-2/drug effects , A549 Cells , Animals , Antiviral Agents/chemistry , COVID-19/drug therapy , COVID-19/virology , Cannabidiol/chemistry , Cannabidiol/metabolism , Chlorocebus aethiops , Endoplasmic Reticulum Stress/drug effects , Endoribonucleases/genetics , Endoribonucleases/metabolism , Epithelial Cells/virology , Female , Gene Expression Regulation, Viral/drug effects , Host-Pathogen Interactions/physiology , Humans , Interferons/metabolism , Mice , /metabolism , SARS-CoV-2/physiology , Vero Cells , Virus Internalization/drug effects , Virus Replication/drug effects
3.
Mol Biol Rep ; 49(2): 1545-1549, 2022 Feb.
Article in English | MEDLINE | ID: covidwho-1653637

ABSTRACT

Coronavirus disease 2019 (COVID-19) is caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) and has resulted in more than 4.4 million deaths worldwide as of August 24, 2021. Viral infections such as SARS-CoV2 are associated with endoplasmic reticulum (ER) stress and also increased the level of reactive oxygen species. Activating transcription factor 4 (ATF4) is preferentially translated under integrated stress conditions and controls the genes involved in protein homeostasis, amino acid transport and metabolism, and also protection from oxidative stress. The GRP78, regulated either directly or indirectly by ATF4, is an essential chaperone in the ER and overexpressed and appears on the surface of almost all cells during stress and function as a SARS-CoV2 receptor. In this mini-review article, we briefly discuss the effects of SARS-CoV2 infection on the ER stress, and then the stress modulator functions of ATF4 and GRP78 as novel therapeutic targets were highlighted. Finally, the effects of GRP78 inhibitory components as potential factors for targeted therapies for COVID-19 critical cases were discussed.


Subject(s)
Activating Transcription Factor 4/metabolism , COVID-19/metabolism , Endoplasmic Reticulum Stress/physiology , Humans , SARS-CoV-2/pathogenicity
4.
Sci Rep ; 11(1): 24432, 2021 12 24.
Article in English | MEDLINE | ID: covidwho-1585772

ABSTRACT

Despite the initial success of some drugs and vaccines targeting COVID-19, understanding the mechanism underlying SARS-CoV-2 disease pathogenesis remains crucial for the development of further approaches to treatment. Some patients with severe Covid-19 experience a cytokine storm and display evidence of inflammasome activation leading to increased levels of IL-1ß and IL-18; however, other reports have suggested reduced inflammatory responses to Sars-Cov-2. In this study we have examined the effects of the Sars-Cov-2 envelope (E) protein, a virulence factor in coronaviruses, on inflammasome activation and pulmonary inflammation. In cultured macrophages the E protein suppressed inflammasome priming and NLRP3 inflammasome activation. Similarly, in mice transfected with E protein and treated with poly(I:C) to simulate the effects of viral RNA, the E protein, in an NLRP3-dependent fashion, reduced expression of pro-IL-1ß, levels of IL-1ß and IL-18 in broncho-alveolar lavage fluid, and macrophage infiltration in the lung. To simulate the effects of more advanced infection, macrophages were treated with both LPS and poly(I:C). In this setting the E protein increased NLRP3 inflammasome activation in both murine and human macrophages. Thus, the Sars-Cov-2 E protein may initially suppress the host NLRP3 inflammasome response to viral RNA while potentially increasing NLRP3 inflammasome responses in the later stages of infection. Targeting the Sars-Cov-2 E protein especially in the early stages of infection may represent a novel approach to Covid-19 therapy.


Subject(s)
Coronavirus Envelope Proteins/metabolism , Inflammasomes/metabolism , NLR Family, Pyrin Domain-Containing 3 Protein/metabolism , SARS-CoV-2/metabolism , Animals , Bronchoalveolar Lavage Fluid/chemistry , COVID-19/pathology , COVID-19/virology , Coronavirus Envelope Proteins/genetics , Down-Regulation/drug effects , Endoplasmic Reticulum Stress , Humans , Inflammasomes/drug effects , Interleukin-1beta/genetics , Interleukin-1beta/metabolism , Janus Kinases/genetics , Janus Kinases/metabolism , Lipopolysaccharides/pharmacology , Macrophages/cytology , Macrophages/drug effects , Macrophages/metabolism , Mice , Mice, Inbred C57BL , Mice, Knockout , NLR Family, Pyrin Domain-Containing 3 Protein/deficiency , NLR Family, Pyrin Domain-Containing 3 Protein/genetics , Poly I-C/pharmacology , RNA, Viral/metabolism , SARS-CoV-2/drug effects , SARS-CoV-2/isolation & purification
5.
Nat Commun ; 12(1): 5536, 2021 09 20.
Article in English | MEDLINE | ID: covidwho-1428813

ABSTRACT

Coronaviruses (CoVs) are important human pathogens for which no specific treatment is available. Here, we provide evidence that pharmacological reprogramming of ER stress pathways can be exploited to suppress CoV replication. The ER stress inducer thapsigargin efficiently inhibits coronavirus (HCoV-229E, MERS-CoV, SARS-CoV-2) replication in different cell types including primary differentiated human bronchial epithelial cells, (partially) reverses the virus-induced translational shut-down, improves viability of infected cells and counteracts the CoV-mediated downregulation of IRE1α and the ER chaperone BiP. Proteome-wide analyses revealed specific pathways, protein networks and components that likely mediate the thapsigargin-induced antiviral state, including essential (HERPUD1) or novel (UBA6 and ZNF622) factors of ER quality control, and ER-associated protein degradation complexes. Additionally, thapsigargin blocks the CoV-induced selective autophagic flux involving p62/SQSTM1. The data show that thapsigargin hits several central mechanisms required for CoV replication, suggesting that this compound (or derivatives thereof) may be developed into broad-spectrum anti-CoV drugs.


Subject(s)
Endoplasmic Reticulum Stress , SARS-CoV-2/physiology , Virus Replication/physiology , Animals , Autophagy/drug effects , Bronchi/pathology , COVID-19/pathology , COVID-19/virology , Cell Differentiation/drug effects , Cell Extracts , Cell Line , Cell Survival/drug effects , Chlorocebus aethiops , Coronavirus 229E, Human/physiology , Down-Regulation/drug effects , Endoplasmic Reticulum Stress/drug effects , Endoplasmic Reticulum Stress/genetics , Endoplasmic Reticulum-Associated Degradation/drug effects , Epithelial Cells/drug effects , Epithelial Cells/virology , Heat-Shock Proteins/metabolism , Humans , Macrolides/pharmacology , Middle East Respiratory Syndrome Coronavirus/drug effects , Middle East Respiratory Syndrome Coronavirus/physiology , Protein Biosynthesis/drug effects , Proteome/metabolism , RNA, Messenger/genetics , RNA, Messenger/metabolism , Reproducibility of Results , SARS-CoV-2/drug effects , Thapsigargin/pharmacology , Unfolded Protein Response/drug effects , Vero Cells , Virus Replication/drug effects
6.
Food Chem Toxicol ; 153: 112286, 2021 Jul.
Article in English | MEDLINE | ID: covidwho-1385569

ABSTRACT

Higher selenium status has been shown to improve the clinical outcome of infections caused by a range of evolutionally diverse viruses, including SARS-CoV-2. However, the impact of SARS-CoV-2 on host-cell selenoproteins remains elusive. The present study investigated the influence of SARS-CoV-2 on expression of selenoprotein mRNAs in Vero cells. SARS-CoV-2 triggered an inflammatory response as evidenced by increased IL-6 expression. Of the 25 selenoproteins, SARS-CoV-2 significantly suppressed mRNA expression of ferroptosis-associated GPX4, DNA synthesis-related TXNRD3 and endoplasmic reticulum-resident SELENOF, SELENOK, SELENOM and SELENOS. Computational analysis has predicted an antisense interaction between SARS-CoV-2 and TXNRD3 mRNA, which is translated with high efficiency in the lung. Here, we confirmed the predicted SARS-CoV-2/TXNRD3 antisense interaction in vitro using DNA oligonucleotides, providing a plausible mechanism for the observed mRNA knockdown. Inhibition of TXNRD decreases DNA synthesis which is thereby likely to increase the ribonucleotide pool for RNA synthesis and, accordingly, RNA virus production. The present findings provide evidence for a direct inhibitory effect of SARS-CoV-2 replication on the expression of a specific set of selenoprotein mRNAs, which merits further investigation in the light of established evidence for correlations between dietary selenium status and the outcome of SARS-CoV-2 infection.


Subject(s)
DNA/biosynthesis , Endoplasmic Reticulum Stress/physiology , Ferroptosis/physiology , RNA, Messenger/metabolism , SARS-CoV-2/physiology , Selenoproteins/metabolism , Animals , Chlorocebus aethiops , Gene Expression Regulation/physiology , RNA, Messenger/genetics , Selenoproteins/genetics , Vero Cells
7.
Cell Stress Chaperones ; 26(5): 859-868, 2021 09.
Article in English | MEDLINE | ID: covidwho-1353732

ABSTRACT

Vaccinations are widely credited with reducing death rates from COVID-19, but the underlying host-viral mechanisms/interactions for morbidity and mortality of SARS-CoV-2 infection remain poorly understood. Acute respiratory distress syndrome (ARDS) describes the severe lung injury, which is pathologically associated with alveolar damage, inflammation, non-cardiogenic edema, and hyaline membrane formation. Because proteostatic pathways play central roles in cellular protection, immune modulation, protein degradation, and tissue repair, we examined the pathological features for the unfolded protein response (UPR) using the surrogate biomarker glucose-regulated protein 78 (GRP78) and co-receptor for SARS-CoV-2. At autopsy, immunostaining of COVID-19 lungs showed highly elevated expression of GRP78 in both pneumocytes and macrophages compared with that of non-COVID control lungs. GRP78 expression was detected in both SARS-CoV-2-infected and un-infected pneumocytes as determined by multiplexed immunostaining for nucleocapsid protein. In macrophages, immunohistochemical staining for GRP78 from deceased COVID-19 patients was increased but overlapped with GRP78 expression taken from surgical resections of non-COVID-19 controls. In contrast, the robust in situ GRP78 immunostaining of pneumocytes from COVID-19 autopsies exhibited no overlap and was independent of age, race/ethnicity, and gender compared with that from non-COVID-19 controls. Our findings bring new insights for stress-response pathways involving the proteostatic network implicated for host resilience and suggest that targeting of GRP78 expression with existing therapeutics might afford an alternative therapeutic strategy to modulate host-viral interactions during SARS-CoV-2 infections.


Subject(s)
Alveolar Epithelial Cells/metabolism , COVID-19/metabolism , Endoplasmic Reticulum Stress , Heat-Shock Proteins/analysis , Receptors, Coronavirus/analysis , SARS-CoV-2/pathogenicity , Adult , Aged , Aged, 80 and over , Alveolar Epithelial Cells/pathology , Alveolar Epithelial Cells/virology , Autopsy , COVID-19/mortality , COVID-19/pathology , COVID-19/virology , Case-Control Studies , Female , Host-Pathogen Interactions , Humans , Macrophages, Alveolar/metabolism , Macrophages, Alveolar/virology , Male , Middle Aged , Proteostasis , Up-Regulation , Young Adult
8.
IUBMB Life ; 74(1): 93-100, 2022 01.
Article in English | MEDLINE | ID: covidwho-1353459

ABSTRACT

Unfolded protein response (UPR) and endoplasmic reticulum (ER) stress are aspects of SARS-CoV-2-host cell interaction with proposed role in the cytopathic and inflammatory pathogenesis of this viral infection. The role of the NF-kB pathway in these cellular processes remains poorly characterized. When investigated in VERO-E6 cells, SARS-CoV-2 infection was found to markedly stimulate NF-kB protein expression and activity. NF-kB activation occurs early in the infection process (6 hpi) and it is associated with increased MAPK signaling and expression of the UPR inducer IRE-1α. These signal transduction processes characterize the cellular stress response to the virus promoting a pro-inflammatory environment and caspase activation in the host cell. Inhibition of viral replication by the viral protease inhibitor Nelfinavir reverts all these molecular changes also stimulating c-Jun expression, a key component of the JNK/AP-1 pathway with important role in the IRE-1α-mediated transcriptional regulation of stress response genes with anti-inflammatory and cytoprotection function. The present study demonstrates that UPR signaling and its interaction with cellular MAPKs and the NF-kB activity are important aspects of SARS-CoV-2-host cell interaction that deserve further investigation to identify more efficient therapies for this viral infection.


Subject(s)
Antiviral Agents/pharmacology , COVID-19/drug therapy , COVID-19/metabolism , Endoplasmic Reticulum Stress/drug effects , NF-kappa B/metabolism , SARS-CoV-2 , Adenosine Monophosphate/analogs & derivatives , Adenosine Monophosphate/pharmacology , Alanine/analogs & derivatives , Alanine/pharmacology , Animals , COVID-19/virology , Caspase 9/metabolism , Chlorocebus aethiops , Cytopathogenic Effect, Viral/drug effects , Humans , MAP Kinase Signaling System/drug effects , Models, Biological , Nelfinavir/pharmacology , SARS-CoV-2/drug effects , SARS-CoV-2/pathogenicity , Unfolded Protein Response/drug effects , Vero Cells
9.
Basic Res Cardiol ; 116(1): 42, 2021 07 05.
Article in English | MEDLINE | ID: covidwho-1293364

ABSTRACT

Coronavirus disease 2019 (COVID-19) spawned a global health crisis in late 2019 and is caused by the novel coronavirus SARS-CoV-2. SARS-CoV-2 infection can lead to elevated markers of endothelial dysfunction associated with higher risk of mortality. It is unclear whether endothelial dysfunction is caused by direct infection of endothelial cells or is mainly secondary to inflammation. Here, we investigate whether different types of endothelial cells are susceptible to SARS-CoV-2. Human endothelial cells from different vascular beds including umbilical vein endothelial cells, coronary artery endothelial cells (HCAEC), cardiac and lung microvascular endothelial cells, or pulmonary arterial cells were inoculated in vitro with SARS-CoV-2. Viral spike protein was only detected in HCAECs after SARS-CoV-2 infection but not in the other endothelial cells tested. Consistently, only HCAEC expressed the SARS-CoV-2 receptor angiotensin-converting enzyme 2 (ACE2), required for virus infection. Infection with the SARS-CoV-2 variants B.1.1.7, B.1.351, and P.2 resulted in significantly higher levels of viral spike protein. Despite this, no intracellular double-stranded viral RNA was detected and the supernatant did not contain infectious virus. Analysis of the cellular distribution of the spike protein revealed that it co-localized with endosomal calnexin. SARS-CoV-2 infection did induce the ER stress gene EDEM1, which is responsible for clearance of misfolded proteins from the ER. Whereas the wild type of SARS-CoV-2 did not induce cytotoxic or pro-inflammatory effects, the variant B.1.1.7 reduced the HCAEC cell number. Of the different tested endothelial cells, HCAECs showed highest viral uptake but did not promote virus replication. Effects on cell number were only observed after infection with the variant B.1.1.7, suggesting that endothelial protection may be particularly important in patients infected with this variant.


Subject(s)
Endoplasmic Reticulum/virology , Endothelial Cells/virology , SARS-CoV-2/pathogenicity , Angiotensin-Converting Enzyme 2/metabolism , Calnexin/metabolism , Cells, Cultured , Endoplasmic Reticulum/metabolism , Endoplasmic Reticulum Stress , Endothelial Cells/metabolism , Host-Pathogen Interactions , Humans , Membrane Proteins/metabolism , Receptors, Virus/metabolism , SARS-CoV-2/genetics , SARS-CoV-2/metabolism , Spike Glycoprotein, Coronavirus/metabolism
10.
Int J Mol Sci ; 22(12)2021 Jun 09.
Article in English | MEDLINE | ID: covidwho-1264471

ABSTRACT

Interstitial lung diseases (ILDs) comprise different fibrotic lung disorders characterized by cellular proliferation, interstitial inflammation, and fibrosis. The JAK/STAT molecular pathway is activated under the interaction of a broad number of profibrotic/pro-inflammatory cytokines, such as IL-6, IL-11, and IL-13, among others, which are increased in different ILDs. Similarly, several growth factors over-expressed in ILDs, such as platelet-derived growth factor (PDGF), transforming growth factor ß1 (TGF-ß1), and fibroblast growth factor (FGF) activate JAK/STAT by canonical or non-canonical pathways, which indicates a predominant role of JAK/STAT in ILDs. Between the different JAK/STAT isoforms, it appears that JAK2/STAT3 are predominant, initiating cellular changes observed in ILDs. This review analyzes the expression and distribution of different JAK/STAT isoforms in ILDs lung tissue and different cell types related to ILDs, such as lung fibroblasts and alveolar epithelial type II cells and analyzes JAK/STAT activation. The effect of JAK/STAT phosphorylation on cellular fibrotic processes, such as proliferation, senescence, autophagy, endoplasmic reticulum stress, or epithelial/fibroblast to mesenchymal transition will be described. The small molecules directed to inhibit JAK/STAT activation were assayed in vitro and in in vivo models of pulmonary fibrosis, and different JAK inhibitors are currently approved for myeloproliferative disorders. Recent evidence indicates that JAK inhibitors or monoclonal antibodies directed to block IL-6 are used as compassionate use to attenuate the excessive inflammation and lung fibrosis related to SARS-CoV-2 virus. These altogether indicate that JAK/STAT pathway is an attractive target to be proven in future clinical trials of lung fibrotic disorders.


Subject(s)
Janus Kinases/metabolism , Lung Diseases, Interstitial/pathology , STAT Transcription Factors/metabolism , Cellular Senescence , Endoplasmic Reticulum Stress , Humans , Interleukins/metabolism , Janus Kinases/antagonists & inhibitors , Janus Kinases/genetics , Lung Diseases, Interstitial/drug therapy , Lung Diseases, Interstitial/metabolism , Protein Isoforms/genetics , Protein Isoforms/metabolism , Protein Kinase Inhibitors/chemistry , Protein Kinase Inhibitors/metabolism , Protein Kinase Inhibitors/therapeutic use , STAT Transcription Factors/antagonists & inhibitors , STAT Transcription Factors/genetics , Signal Transduction
11.
Int J Mol Sci ; 22(11)2021 May 26.
Article in English | MEDLINE | ID: covidwho-1244042

ABSTRACT

Infection induces the production of proinflammatory cytokines and chemokines such as interleukin-8 (IL-8) and IL-6. Although they facilitate local antiviral immunity, their excessive release leads to life-threatening cytokine release syndrome, exemplified by the severe cases of coronavirus disease 2019 (COVID-19) caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection. In this study, we investigated the roles of the integrated stress response (ISR) and activator protein-1 (AP-1) family proteins in regulating coronavirus-induced IL-8 and IL-6 upregulation. The mRNA expression of IL-8 and IL-6 was significantly induced in cells infected with infectious bronchitis virus (IBV), a gammacoronavirus, and porcine epidemic diarrhea virus, an alphacoronavirus. Overexpression of a constitutively active phosphomimetic mutant of eukaryotic translation initiation factor 2α (eIF2α), chemical inhibition of its dephosphorylation, or overexpression of its upstream double-stranded RNA-dependent protein kinase (PKR) significantly enhanced IL-8 mRNA expression in IBV-infected cells. Overexpression of the AP-1 protein cJUN or its upstream kinase also increased the IBV-induced IL-8 mRNA expression, which was synergistically enhanced by overexpression of cFOS. Taken together, this study demonstrated the important regulatory roles of ISR and AP-1 proteins in IL-8 production during coronavirus infection, highlighting the complex interactions between cellular stress pathways and the innate immune response.


Subject(s)
Coronavirus Infections/metabolism , Endoplasmic Reticulum Stress/genetics , Eukaryotic Initiation Factor-2/metabolism , Interleukin-8/metabolism , Unfolded Protein Response/genetics , Alphacoronavirus/metabolism , Alphacoronavirus/pathogenicity , Animals , Cell Line , Chlorocebus aethiops , Coronavirus Infections/genetics , Gammacoronavirus/metabolism , Gammacoronavirus/pathogenicity , Gene Expression Regulation , Humans , Immunity, Innate , Infectious bronchitis virus/metabolism , Infectious bronchitis virus/pathogenicity , Interleukin-8/genetics , Phosphorylation , Porcine epidemic diarrhea virus/metabolism , Porcine epidemic diarrhea virus/pathogenicity , Proto-Oncogene Proteins c-fos/genetics , Proto-Oncogene Proteins c-fos/metabolism , Proto-Oncogene Proteins c-jun/genetics , Proto-Oncogene Proteins c-jun/metabolism , Signal Transduction/genetics , Transcription Factor AP-1/genetics , Transcription Factor AP-1/metabolism , Up-Regulation , Vero Cells , eIF-2 Kinase/genetics , eIF-2 Kinase/metabolism
12.
J Ovarian Res ; 14(1): 70, 2021 May 21.
Article in English | MEDLINE | ID: covidwho-1238729

ABSTRACT

Coronavirus disease 2019 (COVID-19) caused by Severe Acute Respiratory Syndrome Coronavirus 2 (SARS-CoV-2) mainly attacks the respiratory system and is characterized by pneumonia, cytokine storm, coagulation disorders and severe immune downregulation. Although public health experts predicted worst outcomes in Africa, the incidence, hospitalization and mortality rates have been lower in Africa compared to other continents. Interestingly, lower incidence and mortality rates have been observed in women from Africa compared to their cohorts from other continents. Also, in the US non-Hispanic Black females have lower COVID-19 and death rates compared to their white counterparts. It's unclear why this significant difference exists; however, the ovarian function, genetics and immunological statuses could play a major role. Women of African descent have elevated levels of estrogen compared with Caucasians hence we anticipate that estrogen might offer some protection against the SARS-CoV-2 infections. The racial differences in lifestyle, age and inaccessibility to contraceptive usage might also play a role. Here, we provide insight on how the high levels of estrogen in African women might contribute to the lower cases and fatalities in Africa. Specifically, estrogen might offer protection against COVID-19 by suppressing hyper-production of cytokines, promoting anti-inflammatory cytokines, stimulating antibody production and suppressing endoplasmic reticulum (ER) stress. This will as well provide useful information on how future pandemics could be managed using Africa as a case study.


Subject(s)
COVID-19 Testing/trends , COVID-19/epidemiology , COVID-19/etiology , Africa/epidemiology , African Americans , Angiotensin-Converting Enzyme 2/metabolism , COVID-19/drug therapy , COVID-19/mortality , COVID-19 Testing/methods , Cytokine Release Syndrome/etiology , Endoplasmic Reticulum Stress , Estrogens/metabolism , Estrogens/pharmacology , Female , Humans , Incidence , Male , Mortality , Race Factors , Sex Factors
13.
Front Cell Infect Microbiol ; 11: 668034, 2021.
Article in English | MEDLINE | ID: covidwho-1231324

ABSTRACT

The ability to sense and adequately respond to variable environmental conditions is central for cellular and organismal homeostasis. Eukaryotic cells are equipped with highly conserved stress-response mechanisms that support cellular function when homeostasis is compromised, promoting survival. Two such mechanisms - the unfolded protein response (UPR) and autophagy - are involved in the cellular response to perturbations in the endoplasmic reticulum, in calcium homeostasis, in cellular energy or redox status. Each of them operates through conserved signaling pathways to promote cellular adaptations that include re-programming transcription of genes and translation of new proteins and degradation of cellular components. In addition to their specific functions, it is becoming increasingly clear that these pathways intersect in many ways in different contexts of cellular stress. Viral infections are a major cause of cellular stress as many cellular functions are coopted to support viral replication. Both UPR and autophagy are induced upon infection with many different viruses with varying outcomes - in some instances controlling infection while in others supporting viral replication and infection. The role of UPR and autophagy in response to coronavirus infection has been a matter of debate in the last decade. It has been suggested that CoV exploit components of autophagy machinery and UPR to generate double-membrane vesicles where it establishes its replicative niche and to control the balance between cell death and survival during infection. Even though the molecular mechanisms are not fully elucidated, it is clear that UPR and autophagy are intimately associated during CoV infections. The current SARS-CoV-2 pandemic has brought renewed interest to this topic as several drugs known to modulate autophagy - including chloroquine, niclosamide, valinomycin, and spermine - were proposed as therapeutic options. Their efficacy is still debatable, highlighting the need to better understand the molecular interactions between CoV, UPR and autophagy.


Subject(s)
COVID-19 , Autophagy , Endoplasmic Reticulum/metabolism , Endoplasmic Reticulum Stress , Humans , SARS-CoV-2 , Unfolded Protein Response
14.
Sci Adv ; 7(19)2021 05.
Article in English | MEDLINE | ID: covidwho-1226702

ABSTRACT

The endoplasmic reticulum (ER) is a central eukaryotic organelle with a tubular network made of hairpin proteins linked by hydrolysis of guanosine triphosphate nucleotides. Among posttranslational modifications initiated at the ER level, glycosylation is the most common reaction. However, our understanding of the impact of glycosylation on the ER structure remains unclear. Here, we show that exostosin-1 (EXT1) glycosyltransferase, an enzyme involved in N-glycosylation, is a key regulator of ER morphology and dynamics. We have integrated multiomics and superresolution imaging to characterize the broad effect of EXT1 inactivation, including the ER shape-dynamics-function relationships in mammalian cells. We have observed that inactivating EXT1 induces cell enlargement and enhances metabolic switches such as protein secretion. In particular, suppressing EXT1 in mouse thymocytes causes developmental dysfunctions associated with the ER network extension. Last, our data illuminate the physical and functional aspects of the ER proteome-glycome-lipidome structure axis, with implications in biotechnology and medicine.


Subject(s)
Endoplasmic Reticulum Stress , Endoplasmic Reticulum , Animals , Endoplasmic Reticulum/metabolism , Glycosylation , Mammals , Mice , Protein Processing, Post-Translational , Protein Transport
15.
IUBMB Life ; 73(6): 843-854, 2021 06.
Article in English | MEDLINE | ID: covidwho-1219298

ABSTRACT

The 78 kDa glucose-regulated protein (GRP78) is an endoplasmic reticulum (ER)-resident molecular chaperone. GRP78 is a member of the 70 kDa heat shock family of proteins involved in correcting and clearing misfolded proteins in the ER. In response to cellular stress, GRP78 escapes from the ER and moves to the plasma membrane where it (a) functions as a receptor for many ligands, and (b) behaves as an autoantigen for autoantibodies that contribute to human disease and cancer. Cell surface GRP78 (csGRP78) associates with the major histocompatibility complex class I (MHC-I), and is the port of entry for several viruses, including the predictive binding of the novel SARS-CoV-2. Furthermore, csGRP78 is found in association with partners as diverse as the teratocarcinoma-derived growth factor 1 (Cripto), the melanocortin-4 receptor (MC4R) and the DnaJ-like protein MTJ-1. CsGRP78 also serves as a receptor for a large variety of ligands including activated α2 -macroglobulin (α2 M*), plasminogen kringle 5 (K5), microplasminogen, the voltage-dependent anion channel (VDAC), tissue factor (TF), and the prostate apoptosis response-4 protein (Par-4). In this review, we discuss the mechanisms involved in the translocation of GRP78 from the ER to the cell surface, and the role of secreted GRP78 and its autoantibodies in cancer and neurological disorders.


Subject(s)
Autoimmune Diseases of the Nervous System/immunology , COVID-19/transmission , Heat-Shock Proteins/physiology , Neoplasm Proteins/physiology , Nerve Tissue Proteins/physiology , Receptors, Cell Surface/physiology , Receptors, Virus/physiology , SARS-CoV-2/metabolism , Spike Glycoprotein, Coronavirus/metabolism , Autoantibodies/immunology , Autoantigens/immunology , Autoimmune Diseases of the Nervous System/metabolism , Cell Survival , Endoplasmic Reticulum Stress/physiology , Exosomes , GPI-Linked Proteins/metabolism , Heat-Shock Proteins/chemistry , Heat-Shock Proteins/immunology , Humans , Ligands , Neoplasm Invasiveness , Neoplasm Proteins/immunology , Nerve Tissue Proteins/immunology , Protein Domains , Protein Transport , Signal Transduction , Tumor Microenvironment , Unfolded Protein Response/physiology , Virus Internalization
16.
Arch Virol ; 166(8): 2109-2117, 2021 Aug.
Article in English | MEDLINE | ID: covidwho-1216221

ABSTRACT

Millions of people across the globe have been affected by coronavirus disease 2019 (COVID-19), which began in Wuhan, China, and is caused by SARS-CoV-2. COVID-19 has a variety of clinical characteristics and triggers immune responses required for the elimination of the viral agent. Currently, no effective treatment options are available for targeting SARS-CoV-2 infection. Repurposing of drugs such as chloroquine, thalidomide, and leflunomide alongside convalescent plasma is being employed as a therapeutic strategy. Clinical studies have shown that both asymptomatic and symptomatic patients can have an extremely active immune response that is largely attributable to immune system modulations. This includes cytokine storm syndrome (CSS), which affects the adaptive immune system, leading to exhaustion of natural killer (NK) cells and thrombocytopenia in some cases. This review examines the interaction of SARS-CoV-2 with the host immune system and the potential for the development of appropriate immunotherapy for the treatment of COVID-19.


Subject(s)
COVID-19/immunology , SARS-CoV-2/physiology , CD8-Positive T-Lymphocytes/immunology , COVID-19/therapy , Cytokine Release Syndrome/immunology , Endoplasmic Reticulum Stress/immunology , Humans , Immunotherapy , Inflammation , Killer Cells, Natural/immunology , Thrombocytopenia/immunology
17.
Virus Res ; 296: 198350, 2021 04 15.
Article in English | MEDLINE | ID: covidwho-1091608

ABSTRACT

The open reading frame 8 (orf8) is an accessory protein of SARS-CoV-2. It has 121 amino acids with two genotypes, orf8L and orf8S. In this study, we overexpressed the orf8L and orf8S of SARS-CoV-2 as well as the orf8b of SARS-CoV to investigate their roles in the regulation of endoplasmic reticulum (ER) stress and the inhibition of interferon beta (IFNß) production. We found that the two genotypes of SARS-CoV-2 orf8 are capable of inducing ER stress without significant difference by triggering the activating transcription factor 6 (ATF6) and inositol-requiring enzymes 1 (IRE1) branches of the ER stress pathway. However, the third branch of ER stress pathway, i.e. the protein kinase-like ER kinase (PERK), was unaffected by the overexpression of SARS-CoV-2 orf8L or orf8S. Moreover, both orf8L and orf8S of SARS-CoV-2 are capable of down regulating the production of IFNß and interferon-stimulated genes (ISG), ISG15 and ISG56 induced by polyinosinic-polycytidylic acid (poly (I:C)). Moreover, we also found decreased nuclear translocation of Interferon regulatory factor 3 (IRF3), after overexpressing orf8L and orf8S induced by poly (I:C). Our data demonstrated that SARS-CoV-2 orf8 protein could induce ER stress by activating the ATF6 and IRE1 pathways, but not the PERK pathway, and functions as an interferon antagonist to inhibit the production of IFNß. However, these functions appeared not to be affected by the genotypes of SARS-CoV-2 orf8L and orf8S.


Subject(s)
Endoplasmic Reticulum Stress/physiology , Immune Evasion , Interferon-beta/antagonists & inhibitors , Viral Proteins/physiology , Activating Transcription Factor 6/physiology , Endoribonucleases/physiology , HEK293 Cells , Humans , Interferon-beta/biosynthesis , Sequence Alignment , Signal Transduction/physiology , Unfolded Protein Response , Viral Proteins/chemistry , X-Box Binding Protein 1/physiology , eIF-2 Kinase/physiology
18.
Viruses ; 13(2)2021 02 03.
Article in English | MEDLINE | ID: covidwho-1060766

ABSTRACT

The long-term control strategy of SARS-CoV-2 and other major respiratory viruses needs to include antivirals to treat acute infections, in addition to the judicious use of effective vaccines. Whilst COVID-19 vaccines are being rolled out for mass vaccination, the modest number of antivirals in use or development for any disease bears testament to the challenges of antiviral development. We recently showed that non-cytotoxic levels of thapsigargin (TG), an inhibitor of the sarcoplasmic/endoplasmic reticulum (ER) Ca2+ ATPase pump, induces a potent host innate immune antiviral response that blocks influenza A virus replication. Here we show that TG is also highly effective in blocking the replication of respiratory syncytial virus (RSV), common cold coronavirus OC43, SARS-CoV-2 and influenza A virus in immortalized or primary human cells. TG's antiviral performance was significantly better than remdesivir and ribavirin in their respective inhibition of OC43 and RSV. Notably, TG was just as inhibitory to coronaviruses (OC43 and SARS-CoV-2) and influenza viruses (USSR H1N1 and pdm 2009 H1N1) in separate infections as in co-infections. Post-infection oral gavage of acid-stable TG protected mice against a lethal influenza virus challenge. Together with its ability to inhibit the different viruses before or during active infection, and with an antiviral duration of at least 48 h post-TG exposure, we propose that TG (or its derivatives) is a promising broad-spectrum inhibitor against SARS-CoV-2, OC43, RSV and influenza virus.


Subject(s)
Antiviral Agents/pharmacology , Betacoronavirus/drug effects , Coronavirus OC43, Human/drug effects , Influenza A Virus, H1N1 Subtype/drug effects , Respiratory Syncytial Virus, Human/drug effects , SARS-CoV-2/drug effects , Thapsigargin/pharmacology , Animals , Antiviral Agents/therapeutic use , Betacoronavirus/physiology , Cell Line , Cell Line, Tumor , Cells, Cultured , Coronavirus OC43, Human/physiology , Endoplasmic Reticulum Stress , Humans , Influenza A Virus, H1N1 Subtype/physiology , Mice , Microbial Sensitivity Tests , Orthomyxoviridae Infections/drug therapy , Orthomyxoviridae Infections/virology , Respiratory Syncytial Virus, Human/physiology , Ribavirin/pharmacology , SARS-CoV-2/physiology , Thapsigargin/therapeutic use , Virus Replication/drug effects
19.
Brief Funct Genomics ; 20(1): 28-41, 2021 03 02.
Article in English | MEDLINE | ID: covidwho-1045889

ABSTRACT

The human genome has an almost equal distribution of unique and transposable genetic elements. Although at the transcriptome level, a relatively higher contribution from transposable elements derived RNA has been reported. This is further highlighted with evidence from pervasive transcription. Of the total RNA, noncoding RNAs (ncRNAs) are significant contributors to the transcriptome pool with sizeable fraction from repetitive elements of the human genome, inclusive of Long Interspersed Nuclear Elements (LINEs) and Short Interspersed Nuclear Elements (SINEs). ncRNAs are increasingly being implicated in diverse functional roles especially during conditions of stress. These stress responses are driven through diverse mediators, inclusive of long and short ncRNAs. ncRNAs such as MALAT1, GAS5, miR-204 and miR-199a-5p have been functionally involved during oxidative stress, endoplasmic reticulum (ER) stress and unfolded protein response (UPR). Also, within SINEs, Alu RNAs derived from primate-specific Alu repeats with ~11% human genome contribution, playing a significant role. Pathogenic diseases, including the recent COVID-19, leads to differential regulation of ncRNAs. Although, limited evidence suggests the need for an inquest into the role of ncRNAs in determining the host response towards pathogen challenge.


Subject(s)
Infections/genetics , RNA, Untranslated/physiology , COVID-19/genetics , COVID-19/virology , Cytokines/physiology , Endoplasmic Reticulum Stress , Host-Pathogen Interactions , Humans , Infections/metabolism , Long Interspersed Nucleotide Elements , Oxidative Stress , RNA, Untranslated/genetics , SARS-CoV-2/isolation & purification , Short Interspersed Nucleotide Elements , Unfolded Protein Response
20.
Thorax ; 76(1): 92-99, 2021 01.
Article in English | MEDLINE | ID: covidwho-978824

ABSTRACT

The lungs are exposed to a range of environmental toxins (including cigarette smoke, air pollution, asbestos) and pathogens (bacterial, viral and fungal), and most respiratory diseases are associated with local or systemic hypoxia. All of these adverse factors can trigger endoplasmic reticulum (ER) stress. The ER is a key intracellular site for synthesis of secretory and membrane proteins, regulating their folding, assembly into complexes, transport and degradation. Accumulation of misfolded proteins within the lumen results in ER stress, which activates the unfolded protein response (UPR). Effectors of the UPR temporarily reduce protein synthesis, while enhancing degradation of misfolded proteins and increasing the folding capacity of the ER. If successful, homeostasis is restored and protein synthesis resumes, but if ER stress persists, cell death pathways are activated. ER stress and the resulting UPR occur in a range of pulmonary insults and the outcome plays an important role in many respiratory diseases. The UPR is triggered in the airway of patients with several respiratory diseases and in corresponding experimental models. ER stress has been implicated in the initiation and progression of pulmonary fibrosis, and evidence is accumulating suggesting that ER stress occurs in obstructive lung diseases (particularly in asthma), in pulmonary infections (some viral infections and in the setting of the cystic fibrosis airway) and in lung cancer. While a number of small molecule inhibitors have been used to interrogate the role of the UPR in disease models, many of these tools have complex and off-target effects, hence additional evidence (eg, from genetic manipulation) may be required to support conclusions based on the impact of such pharmacological agents. Aberrant activation of the UPR may be linked to disease pathogenesis and progression, but at present, our understanding of the context-specific and disease-specific mechanisms linking these processes is incomplete. Despite this, the ability of the UPR to defend against ER stress and influence a range of respiratory diseases is becoming increasingly evident, and the UPR is therefore attracting attention as a prospective target for therapeutic intervention strategies.


Subject(s)
Endoplasmic Reticulum Stress , Lung Diseases/metabolism , Membrane Proteins/physiology , Humans , Signal Transduction
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