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1.
Nat Commun ; 12(1): 4502, 2021 07 23.
Article in English | MEDLINE | ID: covidwho-1550282

ABSTRACT

Cells in many tissues, such as bone, muscle, and placenta, fuse into syncytia to acquire new functions and transcriptional programs. While it is known that fused cells are specialized, it is unclear whether cell-fusion itself contributes to programmatic-changes that generate the new cellular state. Here, we address this by employing a fusogen-mediated, cell-fusion system to create syncytia from undifferentiated cells. RNA-Seq analysis reveals VSV-G-induced cell fusion precedes transcriptional changes. To gain mechanistic insights, we measure the plasma membrane surface area after cell-fusion and observe it diminishes through increases in endocytosis. Consequently, glucose transporters internalize, and cytoplasmic glucose and ATP transiently decrease. This reduced energetic state activates AMPK, which inhibits YAP1, causing transcriptional-reprogramming and cell-cycle arrest. Impairing either endocytosis or AMPK activity prevents YAP1 inhibition and cell-cycle arrest after fusion. Together, these data demonstrate plasma membrane diminishment upon cell-fusion causes transient nutrient stress that may promote transcriptional-reprogramming independent from extrinsic cues.


Subject(s)
Adaptor Proteins, Signal Transducing/metabolism , Cell Membrane/metabolism , Cell Nucleus/metabolism , Membrane Glycoproteins/metabolism , Transcription Factors/metabolism , Transcription, Genetic/genetics , Viral Envelope Proteins/metabolism , AMP-Activated Protein Kinases/genetics , AMP-Activated Protein Kinases/metabolism , Adaptor Proteins, Signal Transducing/genetics , Animals , Biological Transport , Cell Fusion , Cell Line , Cell Line, Tumor , Cells, Cultured , Giant Cells/metabolism , HEK293 Cells , Humans , Membrane Glycoproteins/genetics , Mice , RNA-Seq/methods , Signal Transduction/genetics , Transcription Factors/genetics , Viral Envelope Proteins/genetics
2.
Front Endocrinol (Lausanne) ; 12: 714909, 2021.
Article in English | MEDLINE | ID: covidwho-1497067

ABSTRACT

Background: Clinically, evidence shows that uterine corpus endometrial carcinoma (UCEC) patients infected with severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) may have a higher death-rate. However, current anti-UCEC/coronavirus disease 2019 (COVID-19) treatment is lacking. Plumbagin (PLB), a pharmacologically active alkaloid, is an emerging anti-cancer inhibitor. Accordingly, the current report was designed to identify and characterize the anti-UCEC function and mechanism of PLB in the treatment of patients infected with SARS-CoV-2 via integrated in silico analysis. Methods: The clinical analyses of UCEC and COVID-19 in patients were conducted using online-accessible tools. Meanwhile, in silico methods including network pharmacology and biological molecular docking aimed to screen and characterize the anti-UCEC/COVID-19 functions, bio targets, and mechanisms of the action of PLB. Results: The bioinformatics data uncovered the clinical characteristics of UCEC patients infected with SARS-CoV-2, including specific genes, health risk, survival rate, and prognostic index. Network pharmacology findings disclosed that PLB-exerted anti-UCEC/COVID-19 effects were achieved through anti-proliferation, inducing cytotoxicity and apoptosis, anti-inflammation, immunomodulation, and modulation of some of the key molecular pathways associated with anti-inflammatory and immunomodulating actions. Following molecular docking analysis, in silico investigation helped identify the anti-UCEC/COVID-19 pharmacological bio targets of PLB, including mitogen-activated protein kinase 3 (MAPK3), tumor necrosis factor (TNF), and urokinase-type plasminogen activator (PLAU). Conclusions: Based on the present bioinformatic and in silico findings, the clinical characterization of UCEC/COVID-19 patients was revealed. The candidate, core bio targets, and molecular pathways of PLB action in the potential treatment of UCEC/COVID-19 were identified accordingly.


Subject(s)
COVID-19 , Carcinoma, Endometrioid , Endometrial Neoplasms , Host-Pathogen Interactions , Naphthoquinones/pharmacology , Adult , Aged , Aged, 80 and over , COVID-19/complications , COVID-19/diagnosis , COVID-19/drug therapy , COVID-19/genetics , Calcium-Binding Proteins/drug effects , Calcium-Binding Proteins/metabolism , Carcinoma, Endometrioid/complications , Carcinoma, Endometrioid/diagnosis , Carcinoma, Endometrioid/drug therapy , Carcinoma, Endometrioid/genetics , Computational Biology , Drug Screening Assays, Antitumor/methods , Endometrial Neoplasms/complications , Endometrial Neoplasms/diagnosis , Endometrial Neoplasms/drug therapy , Endometrial Neoplasms/genetics , Female , Gene Expression Regulation, Neoplastic/drug effects , Gene Regulatory Networks/drug effects , Genetic Association Studies , Host-Pathogen Interactions/drug effects , Host-Pathogen Interactions/genetics , Humans , Membrane Proteins/drug effects , Membrane Proteins/metabolism , Middle Aged , Mitogen-Activated Protein Kinase 3/drug effects , Mitogen-Activated Protein Kinase 3/metabolism , Molecular Docking Simulation/methods , Naphthoquinones/therapeutic use , Prognosis , SARS-CoV-2/drug effects , SARS-CoV-2/physiology , Signal Transduction/drug effects , Signal Transduction/genetics , Tumor Necrosis Factor-alpha/drug effects , Tumor Necrosis Factor-alpha/metabolism , Uterus/drug effects , Uterus/metabolism , Uterus/pathology , Uterus/virology
3.
Sci Rep ; 11(1): 18985, 2021 09 23.
Article in English | MEDLINE | ID: covidwho-1437690

ABSTRACT

The COVID-19 pandemic is raging. It revealed the importance of rapid scientific advancement towards understanding and treating new diseases. To address this challenge, we adapt an explainable artificial intelligence algorithm for data fusion and utilize it on new omics data on viral-host interactions, human protein interactions, and drugs to better understand SARS-CoV-2 infection mechanisms and predict new drug-target interactions for COVID-19. We discover that in the human interactome, the human proteins targeted by SARS-CoV-2 proteins and the genes that are differentially expressed after the infection have common neighbors central in the interactome that may be key to the disease mechanisms. We uncover 185 new drug-target interactions targeting 49 of these key genes and suggest re-purposing of 149 FDA-approved drugs, including drugs targeting VEGF and nitric oxide signaling, whose pathways coincide with the observed COVID-19 symptoms. Our integrative methodology is universal and can enable insight into this and other serious diseases.


Subject(s)
COVID-19/drug therapy , Drug Evaluation, Preclinical/methods , SARS-CoV-2/genetics , Antiviral Agents/therapeutic use , Artificial Intelligence , COVID-19/genetics , COVID-19/metabolism , Drug Repositioning/methods , Gene Regulatory Networks/genetics , Humans , Models, Theoretical , Pandemics , Pharmaceutical Preparations , SARS-CoV-2/drug effects , SARS-CoV-2/pathogenicity , Signal Transduction/genetics
4.
mBio ; 12(5): e0233521, 2021 10 26.
Article in English | MEDLINE | ID: covidwho-1430167

ABSTRACT

Newly emerged severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) caused a global pandemic with astonishing mortality and morbidity. The high replication and transmission of SARS-CoV-2 are remarkably distinct from those of previous closely related coronaviruses, and the underlying molecular mechanisms remain unclear. The innate immune defense is a physical barrier that restricts viral replication. We report here that the SARS-CoV-2 Nsp5 main protease targets RIG-I and mitochondrial antiviral signaling (MAVS) protein via two distinct mechanisms for inhibition. Specifically, Nsp5 cleaves off the 10 most-N-terminal amino acids from RIG-I and deprives it of the ability to activate MAVS, whereas Nsp5 promotes the ubiquitination and proteosome-mediated degradation of MAVS. As such, Nsp5 potently inhibits interferon (IFN) induction by double-stranded RNA (dsRNA) in an enzyme-dependent manner. A synthetic small-molecule inhibitor blunts the Nsp5-mediated destruction of cellular RIG-I and MAVS and processing of SARS-CoV-2 nonstructural proteins, thus restoring the innate immune response and impeding SARS-CoV-2 replication. This work offers new insight into the immune evasion strategy of SARS-CoV-2 and provides a potential antiviral agent to treat CoV disease 2019 (COVID-19) patients. IMPORTANCE The ongoing COVID-19 pandemic is caused by SARS-CoV-2, which is rapidly evolving with better transmissibility. Understanding the molecular basis of the SARS-CoV-2 interaction with host cells is of paramount significance, and development of antiviral agents provides new avenues to prevent and treat COVID-19 diseases. This study describes a molecular characterization of innate immune evasion mediated by the SARS-CoV-2 Nsp5 main protease and subsequent development of a small-molecule inhibitor.


Subject(s)
Adaptor Proteins, Signal Transducing/metabolism , Coronavirus 3C Proteases/metabolism , DEAD Box Protein 58/metabolism , Receptors, Immunologic/metabolism , SARS-CoV-2/immunology , SARS-CoV-2/pathogenicity , A549 Cells , Adaptor Proteins, Signal Transducing/genetics , Animals , Caco-2 Cells , Coronavirus 3C Proteases/genetics , DEAD Box Protein 58/genetics , Enzyme-Linked Immunosorbent Assay , HCT116 Cells , HEK293 Cells , Humans , Immunity, Innate/genetics , Immunity, Innate/physiology , Immunoblotting , Interferon Type I/metabolism , Mice , Receptors, Immunologic/genetics , Reverse Transcriptase Polymerase Chain Reaction , Signal Transduction/genetics , Signal Transduction/physiology , Ubiquitination , Virus Replication/genetics , Virus Replication/physiology
5.
Nat Commun ; 12(1): 652, 2021 01 28.
Article in English | MEDLINE | ID: covidwho-1397868

ABSTRACT

Injury and loss of oligodendrocytes can cause demyelinating diseases such as multiple sclerosis. To improve our understanding of human oligodendrocyte development, which could facilitate development of remyelination-based treatment strategies, here we describe time-course single-cell-transcriptomic analysis of developing human stem cell-derived oligodendrocyte-lineage-cells (hOLLCs). The study includes hOLLCs derived from both genome engineered embryonic stem cell (ESC) reporter cells containing an Identification-and-Purification tag driven by the endogenous PDGFRα promoter and from unmodified induced pluripotent (iPS) cells. Our analysis uncovers substantial transcriptional heterogeneity of PDGFRα-lineage hOLLCs. We discover sub-populations of human oligodendrocyte progenitor cells (hOPCs) including a potential cytokine-responsive hOPC subset, and identify candidate regulatory genes/networks that define the identity of these sub-populations. Pseudotime trajectory analysis defines developmental pathways of oligodendrocytes vs astrocytes from PDGFRα-expressing hOPCs and predicts differentially expressed genes between the two lineages. In addition, pathway enrichment analysis followed by pharmacological intervention of these pathways confirm that mTOR and cholesterol biosynthesis signaling pathways are involved in maturation of oligodendrocytes from hOPCs.


Subject(s)
Genetic Heterogeneity , Genetic Variation , Induced Pluripotent Stem Cells/metabolism , Oligodendrocyte Precursor Cells/metabolism , Single-Cell Analysis/methods , Transcriptome/genetics , Astrocytes/cytology , Astrocytes/metabolism , Cell Differentiation/genetics , Cell Line , Cell Lineage/genetics , Cholesterol/biosynthesis , Embryonic Stem Cells/cytology , Embryonic Stem Cells/metabolism , Gene Regulatory Networks/genetics , Humans , Induced Pluripotent Stem Cells/cytology , Oligodendrocyte Precursor Cells/cytology , Receptor, Platelet-Derived Growth Factor alpha/genetics , Receptor, Platelet-Derived Growth Factor alpha/metabolism , Signal Transduction/genetics , TOR Serine-Threonine Kinases/genetics , TOR Serine-Threonine Kinases/metabolism
6.
Int J Med Sci ; 18(12): 2561-2569, 2021.
Article in English | MEDLINE | ID: covidwho-1389722

ABSTRACT

SARS-CoV-2 infection poses a global challenge to human health. Upon viral infection, host cells initiate the innate antiviral response, which primarily involves type I interferons (I-IFNs), to enable rapid elimination of the invading virus. Previous studies revealed that SARS-CoV-2 infection limits the expression of I-IFNs in vitro and in vivo, but the underlying mechanism remains incompletely elucidated. In the present study, we performed data mining and longitudinal data analysis using SARS-CoV-2-infected normal human bronchial epithelial (NHBE) cells and ferrets, and the results confirmed the strong inhibitory effect of SARS-CoV-2 on the induction of I-IFNs. Moreover, we identified genes that are negatively correlated with IFNB1 expression in vitro and in vivo based on Pearson correlation analysis. We found that SARS-CoV-2 activates numerous intrinsic pathways, such as the circadian rhythm, phosphatidylinositol signaling system, peroxisome, and TNF signaling pathways, to inhibit I-IFNs. These intrinsic inhibitory pathways jointly facilitate the successful immune evasion of SARS-CoV-2. Our study elucidates the underlying mechanism by which SARS-CoV-2 evades the host innate antiviral response in vitro and in vivo, providing theoretical evidence for targeting these immune evasion-associated pathways to combat SARS-CoV-2 infection.


Subject(s)
COVID-19/immunology , Host-Pathogen Interactions/immunology , Interferon-gamma/metabolism , SARS-CoV-2/immunology , Animals , Bronchi/cytology , COVID-19/virology , Cell Line , Datasets as Topic , Disease Models, Animal , Epithelial Cells , Ferrets , Gene Expression Regulation/immunology , Host-Pathogen Interactions/genetics , Humans , Immunity, Innate , Interferon-gamma/immunology , RNA-Seq , Respiratory Mucosa/cytology , Signal Transduction/genetics , Signal Transduction/immunology
7.
J Phys Chem B ; 125(3): 850-873, 2021 01 28.
Article in English | MEDLINE | ID: covidwho-1387111

ABSTRACT

The rapidly growing body of structural and biochemical studies of the SARS-CoV-2 spike glycoprotein has revealed a variety of distinct functional states with radically different arrangements of the receptor-binding domain, highlighting a remarkable function-driven conformational plasticity and adaptability of the spike proteins. In this study, we examined molecular mechanisms underlying conformational and dynamic changes in the SARS-CoV-2 spike mutant trimers through the lens of dynamic analysis of allosteric interaction networks and atomistic modeling of signal transmission. Using an integrated approach that combined coarse-grained molecular simulations, protein stability analysis, and perturbation-based modeling of residue interaction networks, we examined how mutations in the regulatory regions of the SARS-CoV-2 spike protein can differentially affect dynamics and allosteric signaling in distinct functional states. The results of this study revealed key functional regions and regulatory centers that govern collective dynamics, allosteric interactions, and control signal transmission in the SARS-CoV-2 spike proteins. We found that the experimentally confirmed regulatory hotspots that dictate dynamic switching between conformational states of the SARS-CoV-2 spike protein correspond to the key hinge sites and global mediating centers of the allosteric interaction networks. The results of this study provide a novel insight into allosteric regulatory mechanisms of SARS-CoV-2 spike proteins showing that mutations at the key regulatory positions can differentially modulate distribution of states and determine topography of signal communication pathways operating through state-specific cascades of control switch points. This analysis provides a plausible strategy for allosteric probing of the conformational equilibrium and therapeutic intervention by targeting specific hotspots of allosteric interactions and communications in the SARS-CoV-2 spike proteins.


Subject(s)
Models, Biological , Mutation , SARS-CoV-2/chemistry , Spike Glycoprotein, Coronavirus/chemistry , Allosteric Regulation , Binding Sites , Cysteine/genetics , Molecular Dynamics Simulation , Protein Binding , Protein Conformation , Protein Stability , Protein Subunits , SARS-CoV-2/genetics , Signal Transduction/genetics , Spike Glycoprotein, Coronavirus/genetics
8.
Signal Transduct Target Ther ; 5(1): 221, 2020 10 06.
Article in English | MEDLINE | ID: covidwho-1387195
11.
Front Immunol ; 11: 607314, 2020.
Article in English | MEDLINE | ID: covidwho-1389171

ABSTRACT

Acute lung injury (ALI) is an important cause of morbidity and mortality after viral infections, including influenza A virus H1N1, SARS-CoV, MERS-CoV, and SARS-CoV-2. The angiotensin I converting enzyme 2 (ACE2) is a key host membrane-bound protein that modulates ALI induced by viral infection, pulmonary acid aspiration, and sepsis. However, the contributions of ACE2 sequence variants to individual differences in disease risk and severity after viral infection are not understood. In this study, we quantified H1N1 influenza-infected lung transcriptomes across a family of 41 BXD recombinant inbred strains of mice and both parents-C57BL/6J and DBA/2J. In response to infection Ace2 mRNA levels decreased significantly for both parental strains and the expression levels was associated with disease severity (body weight loss) and viral load (expression levels of viral NA segment) across the BXD family members. Pulmonary RNA-seq for 43 lines was analyzed using weighted gene co-expression network analysis (WGCNA) and Bayesian network approaches. Ace2 not only participated in virus-induced ALI by interacting with TNF, MAPK, and NOTCH signaling pathways, but was also linked with high confidence to gene products that have important functions in the pulmonary epithelium, including Rnf128, Muc5b, and Tmprss2. Comparable sets of transcripts were also highlighted in parallel studies of human SARS-CoV-infected primary human airway epithelial cells. Using conventional mapping methods, we determined that weight loss at two and three days after viral infection maps to chromosome X-the location of Ace2. This finding motivated the hierarchical Bayesian network analysis, which defined molecular endophenotypes of lung infection linked to Ace2 expression and to a key disease outcome. Core members of this Bayesian network include Ace2, Atf4, Csf2, Cxcl2, Lif, Maml3, Muc5b, Reg3g, Ripk3, and Traf3. Collectively, these findings define a causally-rooted Ace2 modulatory network relevant to host response to viral infection and identify potential therapeutic targets for virus-induced respiratory diseases, including those caused by influenza and coronaviruses.


Subject(s)
Angiotensin-Converting Enzyme 2/genetics , Lung/virology , Virus Diseases/genetics , Animals , Bayes Theorem , Epithelial Cells/virology , Female , Humans , Mice , Mice, Inbred C57BL , Mice, Inbred DBA , Respiratory Mucosa/virology , Signal Transduction/genetics
12.
Front Immunol ; 12: 700184, 2021.
Article in English | MEDLINE | ID: covidwho-1365542

ABSTRACT

Coronavirus disease 2019 (COVID-19), which has high incidence rates with rapid rate of transmission, is a pandemic that spread across the world, resulting in more than 3,000,000 deaths globally. Currently, several drugs have been used for the clinical treatment of COVID-19, such as antivirals (radecivir, baritinib), monoclonal antibodies (tocilizumab), and glucocorticoids (dexamethasone). Accumulating evidence indicates that long noncoding RNAs (lncRNAs) are essential regulators of virus infections and antiviral immune responses including biological processes that are involved in the regulation of COVID-19 and subsequent disease states. Upon viral infections, cellular lncRNAs directly regulate viral genes and influence viral replication and pathology through virus-mediated changes in the host transcriptome. Additionally, several host lncRNAs could help the occurrence of viral immune escape by inhibiting type I interferons (IFN-1), while others could up-regulate IFN-1 production to play an antiviral role. Consequently, understanding the expression and function of lncRNAs during severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2) infection will provide insights into the development of lncRNA-based methods. In this review, we summarized the current findings of lncRNAs in the regulation of the strong inflammatory response, immune dysfunction and thrombosis induced by SARS-CoV-2 infection, discussed the underlying mechanisms, and highlighted the therapeutic challenges of COVID-19 treatment and its future research directions.


Subject(s)
COVID-19/immunology , Host Microbial Interactions/genetics , Immunity, Innate/genetics , RNA, Long Noncoding/metabolism , Thrombosis/immunology , Antiviral Agents/pharmacology , Antiviral Agents/therapeutic use , Biomarkers/analysis , COVID-19/complications , COVID-19/drug therapy , COVID-19/genetics , COVID-19 Testing/methods , Cytokines/genetics , Cytokines/metabolism , Gene Expression Regulation, Viral/drug effects , Gene Expression Regulation, Viral/immunology , Host Microbial Interactions/drug effects , Host Microbial Interactions/immunology , Humans , Immune Evasion/genetics , Pandemics/prevention & control , RNA, Long Noncoding/analysis , RNA, Long Noncoding/antagonists & inhibitors , SARS-CoV-2/drug effects , SARS-CoV-2/genetics , SARS-CoV-2/immunology , SARS-CoV-2/pathogenicity , Signal Transduction/genetics , Signal Transduction/immunology , Thrombosis/genetics , Thrombosis/virology , Virus Replication/drug effects , Virus Replication/genetics , Virus Replication/immunology
13.
J Med Virol ; 93(9): 5376-5389, 2021 09.
Article in English | MEDLINE | ID: covidwho-1363676

ABSTRACT

The suppression of types I and III interferon (IFN) responses by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) contributes to the pathogenesis of coronavirus disease 2019 (COVID-19). The strategy used by SARS-CoV-2 to evade antiviral immunity needs further investigation. Here, we reported that SARS-CoV-2 ORF9b inhibited types I and III IFN production by targeting multiple molecules of innate antiviral signaling pathways. SARS-CoV-2 ORF9b impaired the induction of types I and III IFNs by Sendai virus and poly (I:C). SARS-CoV-2 ORF9b inhibited the activation of types I and III IFNs induced by the components of cytosolic dsRNA-sensing pathways of RIG-I/MDA5-MAVS signaling, including RIG-I, MDA-5, MAVS, TBK1, and IKKε, rather than IRF3-5D, which is the active form of IRF3. SARS-CoV-2 ORF9b also suppressed the induction of types I and III IFNs by TRIF and STING, which are the adaptor protein of the endosome RNA-sensing pathway of TLR3-TRIF signaling and the adaptor protein of the cytosolic DNA-sensing pathway of cGAS-STING signaling, respectively. A mechanistic analysis revealed that the SARS-CoV-2 ORF9b protein interacted with RIG-I, MDA-5, MAVS, TRIF, STING, and TBK1 and impeded the phosphorylation and nuclear translocation of IRF3. In addition, SARS-CoV-2 ORF9b facilitated the replication of the vesicular stomatitis virus. Therefore, the results showed that SARS-CoV-2 ORF9b negatively regulates antiviral immunity and thus facilitates viral replication. This study contributes to our understanding of the molecular mechanism through which SARS-CoV-2 impairs antiviral immunity and provides an essential clue to the pathogenesis of COVID-19.


Subject(s)
DEAD Box Protein 58/immunology , Immune Evasion/genetics , Interferons/immunology , Nucleotidyltransferases/immunology , Receptors, Immunologic/immunology , SARS-CoV-2/immunology , Toll-Like Receptor 3/immunology , Adaptor Proteins, Signal Transducing/genetics , Adaptor Proteins, Signal Transducing/immunology , Adaptor Proteins, Vesicular Transport/genetics , Adaptor Proteins, Vesicular Transport/immunology , Animals , Chlorocebus aethiops , Coronavirus Nucleocapsid Proteins/genetics , Coronavirus Nucleocapsid Proteins/immunology , DEAD Box Protein 58/genetics , Gene Expression Regulation , HEK293 Cells , HeLa Cells , Humans , I-kappa B Kinase/genetics , I-kappa B Kinase/immunology , Immunity, Innate , Interferon Regulatory Factor-3/genetics , Interferon Regulatory Factor-3/immunology , Interferon-Induced Helicase, IFIH1/genetics , Interferon-Induced Helicase, IFIH1/immunology , Interferons/genetics , Membrane Proteins/genetics , Membrane Proteins/immunology , Nucleotidyltransferases/genetics , Phosphoproteins/genetics , Phosphoproteins/immunology , Plasmids/chemistry , Plasmids/metabolism , /immunology , Receptors, Immunologic/genetics , SARS-CoV-2/genetics , SARS-CoV-2/pathogenicity , Signal Transduction/genetics , Signal Transduction/immunology , Toll-Like Receptor 3/genetics , Transfection , Vero Cells , Virus Replication/immunology
14.
Viruses ; 13(8)2021 08 12.
Article in English | MEDLINE | ID: covidwho-1355052

ABSTRACT

Severe Acute Respiratory Syndrome Coronavirus 2 (SARS-CoV-2) is the causative agent of coronavirus disease 2019 (COVID-19), a global pandemic characterized by an exaggerated immune response and respiratory illness. Age (>60 years) is a significant risk factor for developing severe COVID-19. To better understand the host response of the aged airway epithelium to SARS-CoV-2 infection, we performed an in vitro study using primary human bronchial epithelial cells from donors >67 years of age differentiated on an air-liquid interface culture. We demonstrate that SARS-CoV-2 infection leads to early induction of a proinflammatory response and a delayed interferon response. In addition, we observed changes in the genes and pathways associated with cell death and senescence throughout infection. In summary, our study provides new and important insights into the temporal kinetics of the airway epithelial innate immune response to SARS-CoV-2 in older individuals.


Subject(s)
Bronchi/immunology , Bronchi/virology , Immunity, Innate , Respiratory Mucosa/immunology , Respiratory Mucosa/virology , SARS-CoV-2/immunology , Aged , Aging/immunology , Bronchi/cytology , Bronchi/metabolism , COVID-19/immunology , Cell Death/genetics , Cells, Cultured , Cellular Senescence/genetics , Cytokines/biosynthesis , Cytokines/genetics , Epithelial Cells/immunology , Epithelial Cells/metabolism , Epithelial Cells/virology , Female , Humans , Inflammation , Interferons/biosynthesis , Interferons/genetics , Male , RNA-Seq , Respiratory Mucosa/cytology , Respiratory Mucosa/metabolism , SARS-CoV-2/physiology , Signal Transduction/genetics
15.
Int J Mol Sci ; 22(15)2021 Jul 28.
Article in English | MEDLINE | ID: covidwho-1335097

ABSTRACT

Different mechanisms were proposed as responsible for COVID-19 neurological symptoms but a clear one has not been established yet. In this work we aimed to study SARS-CoV-2 capacity to infect pediatric human cortical neuronal HCN-2 cells, studying the changes in the transcriptomic profile by next generation sequencing. SARS-CoV-2 was able to replicate in HCN-2 cells, that did not express ACE2, confirmed also with Western blot, and TMPRSS2. Looking for pattern recognition receptor expression, we found the deregulation of scavenger receptors, such as SR-B1, and the downregulation of genes encoding for Nod-like receptors. On the other hand, TLR1, TLR4 and TLR6 encoding for Toll-like receptors (TLRs) were upregulated. We also found the upregulation of genes encoding for ERK, JNK, NF-κB and Caspase 8 in our transcriptomic analysis. Regarding the expression of known receptors for viral RNA, only RIG-1 showed an increased expression; downstream RIG-1, the genes encoding for TRAF3, IKKε and IRF3 were downregulated. We also found the upregulation of genes encoding for chemokines and accordingly we found an increase in cytokine/chemokine levels in the medium. According to our results, it is possible to speculate that additionally to ACE2 and TMPRSS2, also other receptors may interact with SARS-CoV-2 proteins and mediate its entry or pathogenesis in pediatric cortical neurons infected with SARS-CoV-2. In particular, TLRs signaling could be crucial for the neurological involvement related to SARS-CoV-2 infection.


Subject(s)
COVID-19/metabolism , Cerebral Cortex/metabolism , Neurons/virology , SARS-CoV-2/pathogenicity , Toll-Like Receptors/metabolism , COVID-19/genetics , COVID-19/immunology , Child , Cytokines/metabolism , Gene Expression Profiling , Gene Expression Regulation , Humans , Neurons/immunology , SARS-CoV-2/immunology , SARS-CoV-2/metabolism , Signal Transduction/genetics , Toll-Like Receptors/genetics , Virus Replication
16.
Respir Res ; 21(1): 154, 2020 Jun 18.
Article in English | MEDLINE | ID: covidwho-1331943

ABSTRACT

Electronic cigarette (e-cig) vaping is increasing rapidly in the United States, as e-cigs are considered less harmful than combustible cigarettes. However, limited research has been conducted to understand the possible mechanisms that mediate toxicity and pulmonary health effects of e-cigs. We hypothesized that sub-chronic e-cig exposure induces inflammatory response and dysregulated repair/extracellular matrix (ECM) remodeling, which occur through the α7 nicotinic acetylcholine receptor (nAChRα7). Adult wild-type (WT), nAChRα7 knockout (KO), and lung epithelial cell-specific KO (nAChRα7 CreCC10) mice were exposed to e-cig aerosol containing propylene glycol (PG) with or without nicotine. Bronchoalveolar lavage fluids (BALF) and lung tissues were collected to determine e-cig induced inflammatory response and ECM remodeling, respectively. Sub-chronic e-cig exposure with nicotine increased inflammatory cellular influx of macrophages and T-lymphocytes including increased pro-inflammatory cytokines in BALF and increased SARS-Cov-2 Covid-19 ACE2 receptor, whereas nAChRα7 KO mice show reduced inflammatory responses associated with decreased ACE2 receptor. Interestingly, matrix metalloproteinases (MMPs), such as MMP2, MMP8 and MMP9, were altered both at the protein and mRNA transcript levels in female and male KO mice, but WT mice exposed to PG alone showed a sex-dependent phenotype. Moreover, MMP12 was increased significantly in male mice exposed to PG with or without nicotine in a nAChRα7-dependent manner. Additionally, sub-chronic e-cig exposure with or without nicotine altered the abundance of ECM proteins, such as collagen and fibronectin, significantly in a sex-dependent manner, but without the direct role of nAChRα7 gene. Overall, sub-chronic e-cig exposure with or without nicotine affected lung inflammation and repair responses/ECM remodeling, which were mediated by nAChRα7 in a sex-dependent manner.


Subject(s)
Coronavirus Infections/epidemiology , Electronic Nicotine Delivery Systems , Peptidyl-Dipeptidase A/metabolism , Pneumonia, Viral/epidemiology , Pneumonia/metabolism , Vaping/adverse effects , alpha7 Nicotinic Acetylcholine Receptor/genetics , Angiotensin-Converting Enzyme 2 , Animals , Blood Gas Analysis , Blotting, Western , Bronchoalveolar Lavage Fluid , COVID-19 , Cytokines/analysis , Disease Models, Animal , Female , Humans , Male , Mice , Mice, Inbred C57BL , Mice, Knockout , Pandemics , Pneumonia/physiopathology , Random Allocation , Reference Values , Role , Severe Acute Respiratory Syndrome/epidemiology , Signal Transduction/genetics
17.
Sci Rep ; 11(1): 15107, 2021 07 23.
Article in English | MEDLINE | ID: covidwho-1322506

ABSTRACT

COVID-19 outbreak brings intense pressure on healthcare systems, with an urgent demand for effective diagnostic, prognostic and therapeutic procedures. Here, we employed Automated Machine Learning (AutoML) to analyze three publicly available high throughput COVID-19 datasets, including proteomic, metabolomic and transcriptomic measurements. Pathway analysis of the selected features was also performed. Analysis of a combined proteomic and metabolomic dataset led to 10 equivalent signatures of two features each, with AUC 0.840 (CI 0.723-0.941) in discriminating severe from non-severe COVID-19 patients. A transcriptomic dataset led to two equivalent signatures of eight features each, with AUC 0.914 (CI 0.865-0.955) in identifying COVID-19 patients from those with a different acute respiratory illness. Another transcriptomic dataset led to two equivalent signatures of nine features each, with AUC 0.967 (CI 0.899-0.996) in identifying COVID-19 patients from virus-free individuals. Signature predictive performance remained high upon validation. Multiple new features emerged and pathway analysis revealed biological relevance by implication in Viral mRNA Translation, Interferon gamma signaling and Innate Immune System pathways. In conclusion, AutoML analysis led to multiple biosignatures of high predictive performance, with reduced features and large choice of alternative predictors. These favorable characteristics are eminent for development of cost-effective assays to contribute to better disease management.


Subject(s)
COVID-19/diagnosis , COVID-19/metabolism , Immunity, Innate/immunology , Machine Learning , SARS-CoV-2/metabolism , Biomarkers/blood , COVID-19/genetics , COVID-19/pathology , Computer Simulation , Databases, Factual , Databases, Genetic , Databases, Protein , Gene Expression Profiling , Humans , Immunity, Innate/genetics , Interferon-gamma/blood , Metabolomics , Prognosis , Proteomics , ROC Curve , SARS-CoV-2/genetics , Severity of Illness Index , Signal Transduction/genetics , Signal Transduction/immunology , Software
18.
PLoS One ; 16(6): e0253089, 2021.
Article in English | MEDLINE | ID: covidwho-1282298

ABSTRACT

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) has caused a devastating global pandemic, infecting over 43 million people and claiming over 1 million lives, with these numbers increasing daily. Therefore, there is urgent need to understand the molecular mechanisms governing SARS-CoV-2 pathogenesis, immune evasion, and disease progression. Here, we show that SARS-CoV-2 can block IRF3 and NF-κB activation early during virus infection. We also identify that the SARS-CoV-2 viral proteins NSP1 and NSP13 can block interferon activation via distinct mechanisms. NSP1 antagonizes interferon signaling by suppressing host mRNA translation, while NSP13 downregulates interferon and NF-κB promoter signaling by limiting TBK1 and IRF3 activation, as phospho-TBK1 and phospho-IRF3 protein levels are reduced with increasing levels of NSP13 protein expression. NSP13 can also reduce NF-κB activation by both limiting NF-κB phosphorylation and nuclear translocation. Last, we also show that NSP13 binds to TBK1 and downregulates IFIT1 protein expression. Collectively, these data illustrate that SARS-CoV-2 bypasses multiple innate immune activation pathways through distinct mechanisms.


Subject(s)
Adaptor Proteins, Signal Transducing/immunology , COVID-19/immunology , Cell Nucleus/immunology , Interferon Regulatory Factor-3/immunology , RNA-Binding Proteins/immunology , SARS-CoV-2/immunology , Signal Transduction/immunology , Viral Nonstructural Proteins/immunology , Active Transport, Cell Nucleus/genetics , Active Transport, Cell Nucleus/immunology , Adaptor Proteins, Signal Transducing/genetics , COVID-19/genetics , Cell Nucleus/genetics , HeLa Cells , Humans , Interferon Regulatory Factor-3/genetics , NF-kappa B/genetics , NF-kappa B/immunology , Phosphorylation/genetics , Phosphorylation/immunology , /immunology , RNA-Binding Proteins/genetics , SARS-CoV-2/genetics , Signal Transduction/genetics , Viral Nonstructural Proteins/genetics
19.
Sci Rep ; 11(1): 12948, 2021 06 21.
Article in English | MEDLINE | ID: covidwho-1279894

ABSTRACT

COVID 19 disease has become a global catastrophe over the past year that has claimed the lives of over two million people around the world. Despite the introduction of vaccines against the disease, there is still a long way to completely eradicate it. There are concerns about the complications following infection with SARS-CoV-2. This research aimed to evaluate the possible correlation between infection with SARS-CoV viruses and cancer in an in-silico study model. To do this, the relevent dataset was selected from GEO database. Identification of differentially expressed genes among defined groups including SARS-CoV, SARS-dORF6, SARS-BatSRBD, and H1N1 were screened where the |Log FC| ≥ 1and p < 0.05 were considered statistically significant. Later, the pathway enrichment analysis and gene ontology (GO) were used by Enrichr and Shiny GO databases. Evaluation with STRING online was applied to predict the functional interactions of proteins, followed by Cytoscape analysis to identify the master genes. Finally, analysis with GEPIA2 server was carried out to reveal the possible correlation between candidate genes and cancer development. The results showed that the main molecular function of up- and down-regulated genes was "double-stranded RNA binding" and actin-binding, respectively. STRING and Cytoscape analysis presented four genes, PTEN, CREB1, CASP3, and SMAD3 as the key genes involved in cancer development. According to TCGA database results, these four genes were up-regulated notably in pancreatic adenocarcinoma. Our findings suggest that pancreatic adenocarcinoma is the most probably malignancy happening after infection with SARS-CoV family.


Subject(s)
Adenocarcinoma/etiology , COVID-19/complications , Carcinogenesis/genetics , Influenza A Virus, H1N1 Subtype , Influenza, Human/complications , Pancreatic Neoplasms/etiology , SARS Virus , SARS-CoV-2 , Severe Acute Respiratory Syndrome/complications , COVID-19/genetics , COVID-19/metabolism , COVID-19/virology , Caspase 3/genetics , Cyclic AMP Response Element-Binding Protein/genetics , Gene Expression Regulation , Gene Ontology , Humans , Influenza, Human/genetics , Influenza, Human/metabolism , Influenza, Human/virology , PTEN Phosphohydrolase/genetics , Protein Interaction Maps , Risk , Severe Acute Respiratory Syndrome/genetics , Severe Acute Respiratory Syndrome/metabolism , Severe Acute Respiratory Syndrome/virology , Signal Transduction/genetics , Smad3 Protein/genetics , Up-Regulation/genetics
20.
Sci Immunol ; 6(60)2021 06 18.
Article in English | MEDLINE | ID: covidwho-1276879

ABSTRACT

The nutrient-sensing mammalian target of rapamycin (mTOR) is integral to cell fate decisions after T cell activation. Sustained mTORC1 activity favors the generation of terminally differentiated effector T cells instead of follicular helper and memory T cells. This is particularly pertinent for T cell responses of older adults who have sustained mTORC1 activation despite dysfunctional lysosomes. Here, we show that lysosome-deficient T cells rely on late endosomes rather than lysosomes as an mTORC1 activation platform, where mTORC1 is activated by sensing cytosolic amino acids. T cells from older adults have an increased expression of the plasma membrane leucine transporter SLC7A5 to provide a cytosolic amino acid source. Hence, SLC7A5 and VPS39 deficiency (a member of the HOPS complex promoting early to late endosome conversion) substantially reduced mTORC1 activities in T cells from older but not young individuals. Late endosomal mTORC1 is independent of the negative-feedback loop involving mTORC1-induced inactivation of the transcription factor TFEB that controls expression of lysosomal genes. The resulting sustained mTORC1 activation impaired lysosome function and prevented lysosomal degradation of PD-1 in CD4+ T cells from older adults, thereby inhibiting their proliferative responses. VPS39 silencing of human T cells improved their expansion to pertussis and to SARS-CoV-2 peptides in vitro. Furthermore, adoptive transfer of CD4+ Vps39-deficient LCMV-specific SMARTA cells improved germinal center responses, CD8+ memory T cell generation, and recall responses to infection. Thus, curtailing late endosomal mTORC1 activity is a promising strategy to enhance T cell immunity.


Subject(s)
CD4-Positive T-Lymphocytes/immunology , CD8-Positive T-Lymphocytes/immunology , COVID-19/immunology , Endosomes/metabolism , Mechanistic Target of Rapamycin Complex 1/metabolism , SARS-CoV-2/metabolism , Signal Transduction/genetics , Adoptive Transfer/methods , Adult , Aged , Aged, 80 and over , Animals , Autophagy-Related Proteins/deficiency , Autophagy-Related Proteins/genetics , Basic Helix-Loop-Helix Leucine Zipper Transcription Factors/genetics , Basic Helix-Loop-Helix Leucine Zipper Transcription Factors/metabolism , COVID-19/virology , Cells, Cultured , Female , Forkhead Box Protein O1/deficiency , Forkhead Box Protein O1/genetics , Healthy Volunteers , Humans , Large Neutral Amino Acid-Transporter 1/metabolism , Lysosomes/metabolism , Male , Mice , Mice, Inbred C57BL , Mice, Transgenic , Signal Transduction/immunology , Transfection , Vesicular Transport Proteins/deficiency , Vesicular Transport Proteins/genetics , Young Adult
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