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1.
Proc Natl Acad Sci U S A ; 118(19)2021 05 11.
Article in English | MEDLINE | ID: covidwho-1203481

ABSTRACT

Binding of the spike protein of SARS-CoV-2 to the human angiotensin-converting enzyme 2 (ACE2) receptor triggers translocation of the virus into cells. Both the ACE2 receptor and the spike protein are heavily glycosylated, including at sites near their binding interface. We built fully glycosylated models of the ACE2 receptor bound to the receptor binding domain (RBD) of the SARS-CoV-2 spike protein. Using atomistic molecular dynamics (MD) simulations, we found that the glycosylation of the human ACE2 receptor contributes substantially to the binding of the virus. Interestingly, the glycans at two glycosylation sites, N90 and N322, have opposite effects on spike protein binding. The glycan at the N90 site partly covers the binding interface of the spike RBD. Therefore, this glycan can interfere with the binding of the spike protein and protect against docking of the virus to the cell. By contrast, the glycan at the N322 site interacts tightly with the RBD of the ACE2-bound spike protein and strengthens the complex. Remarkably, the N322 glycan binds to a conserved region of the spike protein identified previously as a cryptic epitope for a neutralizing antibody. By mapping the glycan binding sites, our MD simulations aid in the targeted development of neutralizing antibodies and SARS-CoV-2 fusion inhibitors.


Subject(s)
/metabolism , Spike Glycoprotein, Coronavirus/metabolism , Glycosylation , Humans , Molecular Dynamics Simulation , Protein Binding , Virus Internalization
2.
Nat Commun ; 12(1): 2417, 2021 04 23.
Article in English | MEDLINE | ID: covidwho-1203428

ABSTRACT

SARS-CoV-2 uses ACE2, an inhibitor of the Renin-Angiotensin-Aldosterone System (RAAS), for cellular entry. Studies indicate that RAAS imbalance worsens the prognosis in COVID-19. We present a consecutive retrospective COVID-19 cohort with findings of frequent pulmonary thromboembolism (17%), high pulmonary artery pressure (60%) and lung MRI perfusion disturbances. We demonstrate, in swine, that infusing angiotensin II or blocking ACE2 induces increased pulmonary artery pressure, reduces blood oxygenation, increases coagulation, disturbs lung perfusion, induces diffuse alveolar damage, and acute tubular necrosis compared to control animals. We further demonstrate that this imbalanced state can be ameliorated by infusion of an angiotensin receptor blocker and low-molecular-weight heparin. In this work, we show that a pathophysiological state in swine induced by RAAS imbalance shares several features with the clinical COVID-19 presentation. Therefore, we propose that severe COVID-19 could partially be driven by a RAAS imbalance.


Subject(s)
/physiopathology , Lung/physiopathology , Renin-Angiotensin System/physiology , /isolation & purification , Angiotensin II/administration & dosage , Angiotensin II/metabolism , Angiotensin Receptor Antagonists/administration & dosage , /metabolism , Animals , /virology , Female , Humans , Lung/diagnostic imaging , Lung/virology , Magnetic Resonance Imaging/methods , Protein Binding/drug effects , Retrospective Studies , /physiology , Spike Glycoprotein, Coronavirus/metabolism , Swine , Virus Internalization/drug effects
3.
Int J Mol Sci ; 22(9)2021 Apr 25.
Article in English | MEDLINE | ID: covidwho-1202187

ABSTRACT

SARS-CoV-2, the causative agent of COVID-19, infects host cells using the angiotensin I converting enzyme 2 (ACE2) as its receptor after priming by host proteases, including TMPRSS2. COVID-19 affects multiple organ systems, and male patients suffer increased severity and mortality. Polycystic Ovary Syndrome (PCOS) is the most common endocrine disorder in reproductive-age women and is characterized by hyperandrogenism, ovulatory dysfunction, and polycystic ovarian morphology. PCOS is associated with obesity and cardiometabolic comorbidities, both being risk factors associated with severe COVID-19 pathology. We hypothesize that elevated androgens in PCOS regulate SARS-CoV-2 entry proteins in multiple tissues increasing the risk for this population. Female mice were treated with dihydrotestosterone (DHT) for 90 days. Body composition was measured by EchoMRI. Fasting glucose was determined by an enzymatic method. mRNA and protein levels of ACE2, Tmprss2, Cathepsin L, Furin, Tmprss4, and Adam17 were quantified by RT-qPCR, Western-blot, or ELISA in tissues, serum, and urine. DHT treatment increased body weight, fat and lean mass, and fasting glucose. Ace2 mRNA was upregulated in the lung, cecum, heart, and kidney, while downregulated in the brain by DHT. ACE2 protein was upregulated by DHT in the small intestine, heart, and kidney. The SARS-CoV-2 priming proteases Tmprss2, Cathepsin L, and Furin mRNA were upregulated by DHT in the kidney. ACE2 sheddase Adam17 mRNA was upregulated by DHT in the kidney, which corresponded with increased urinary ACE2 in DHT treated mice. Our results highlight the potential for increased cardiac, renal, and gastrointestinal dysfunction in PCOS women with COVID-19.


Subject(s)
/pathology , Hyperandrogenism/pathology , Polycystic Ovary Syndrome/pathology , /metabolism , /blood , /metabolism , Animals , Blood Glucose/analysis , Body Weight/drug effects , /virology , Cathepsin L/genetics , Cathepsin L/metabolism , Dihydrotestosterone/pharmacology , Female , Humans , Kidney/metabolism , Mice , Mice, Inbred C57BL , Polycystic Ovary Syndrome/complications , Serine Endopeptidases/genetics , Serine Endopeptidases/metabolism , Up-Regulation/drug effects , Virus Internalization
4.
Int J Mol Sci ; 22(9)2021 Apr 26.
Article in English | MEDLINE | ID: covidwho-1201444

ABSTRACT

SARS-CoV-2 impairs the renin-angiotensin-aledosterone system via binding ACE2 enzyme. ACE2 plays a key role in the biosynthesis of angiotensin (1-7), catalyzing the conversion of angiotensin 2 into angiotensin (1-7) and the reaction of angiotensin synthesis (1-9), from which angiotensin is (1-7) produced under the influence of ACE (Angiotensin-Converting Enzyme). Angiotensin 2 is a potent vasoconstrictor and atherogenic molecule converted by ACE2 to reducing inflammation and vasodilating in action angiotensin (1-7). Angiotensin (1-9), that is a product of angiotensin 1 metabolism and precursor of angiotensin (1-7), also exerts cell protective properties. Balance between angiotensin 2 and angiotensin (1-7) regulates blood pressure and ACE2 plays a critical role in this balance. ACE2, unlike ACE, is not inhibited by ACE inhibitors at the doses used in humans during the treatment of arterial hypertension. Membrane ACE2 is one of the receptors that allows SARS-CoV-2 to enter the host cells. ACE2 after SARS-CoV-2 binding is internalized and degraded. Hence ACE2 activity on the cell surface is reduced leading to increase the concentration of angiotensin 2 and decrease the concentration of angiotensin (1-7). Disturbed angiotensins metabolism, changes in ratio between angiotensins with distinct biological activities leading to domination of atherogenic angiotensin 2 can increase the damage to the lungs.


Subject(s)
/pathology , Renin-Angiotensin System/physiology , /metabolism , /metabolism , Angiotensins/metabolism , Humans , Peptidyl-Dipeptidase A/metabolism , Virus Internalization
5.
Sci Rep ; 11(1): 8692, 2021 04 22.
Article in English | MEDLINE | ID: covidwho-1199310

ABSTRACT

A metal nanoparticle composite, namely TPNT1, which contains Au-NP (1 ppm), Ag-NP (5 ppm), ZnO-NP (60 ppm) and ClO2 (42.5 ppm) in aqueous solution was prepared and characterized by spectroscopy, transmission electron microscopy, dynamic light scattering analysis and potentiometric titration. Based on the in vitro cell-based assay, TPNT1 inhibited six major clades of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) with effective concentration within the range to be used as food additives. TPNT1 was shown to block viral entry by inhibiting the binding of SARS-CoV-2 spike proteins to the angiotensin-converting enzyme 2 (ACE2) receptor and to interfere with the syncytium formation. In addition, TPNT1 also effectively reduced the cytopathic effects induced by human (H1N1) and avian (H5N1) influenza viruses, including the wild-type and oseltamivir-resistant virus isolates. Together with previously demonstrated efficacy as antimicrobials, TPNT1 can block viral entry and inhibit or prevent viral infection to provide prophylactic effects against both SARS-CoV-2 and opportunistic infections.


Subject(s)
Gold/pharmacology , Influenza A Virus, H1N1 Subtype/physiology , Influenza A Virus, H5N1 Subtype/physiology , Silver/pharmacology , Zinc Oxide/pharmacology , /metabolism , Antiviral Agents/chemistry , Antiviral Agents/pharmacology , Drug Resistance, Viral/drug effects , Food Additives/pharmacology , Gold/chemistry , HEK293 Cells , Humans , Influenza A Virus, H1N1 Subtype/drug effects , Influenza A Virus, H5N1 Subtype/drug effects , Metal Nanoparticles/chemistry , Nanocomposites/chemistry , Oseltamivir/pharmacology , Particle Size , Protein Binding/drug effects , Silver/chemistry , Spike Glycoprotein, Coronavirus/metabolism , Virus Internalization/drug effects , Zinc Oxide/chemistry
6.
PLoS Pathog ; 17(4): e1009500, 2021 04.
Article in English | MEDLINE | ID: covidwho-1197396

ABSTRACT

The high transmissibility of SARS-CoV-2 is related to abundant replication in the upper airways, which is not observed for the other highly pathogenic coronaviruses SARS-CoV and MERS-CoV. We here reveal features of the coronavirus spike (S) protein, which optimize the virus towards the human respiratory tract. First, the S proteins exhibit an intrinsic temperature preference, corresponding with the temperature of the upper or lower airways. Pseudoviruses bearing the SARS-CoV-2 spike (SARS-2-S) were more infectious when produced at 33°C instead of 37°C, a property shared with the S protein of HCoV-229E, a common cold coronavirus. In contrast, the S proteins of SARS-CoV and MERS-CoV favored 37°C, in accordance with virus preference for the lower airways. Next, SARS-2-S-driven entry was efficiently activated by not only TMPRSS2, but also the TMPRSS13 protease, thus broadening the cell tropism of SARS-CoV-2. Both proteases proved relevant in the context of authentic virus replication. TMPRSS13 appeared an effective spike activator for the virulent coronaviruses but not the low pathogenic HCoV-229E virus. Activation of SARS-2-S by these surface proteases requires processing of the S1/S2 cleavage loop, in which both the furin recognition motif and extended loop length proved critical. Conversely, entry of loop deletion mutants is significantly increased in cathepsin-rich cells. Finally, we demonstrate that the D614G mutation increases SARS-CoV-2 stability, particularly at 37°C, and, enhances its use of the cathepsin L pathway. This indicates a link between S protein stability and usage of this alternative route for virus entry. Since these spike properties may promote virus spread, they potentially explain why the spike-G614 variant has replaced the early D614 variant to become globally predominant. Collectively, our findings reveal adaptive mechanisms whereby the coronavirus spike protein is adjusted to match the temperature and protease conditions of the airways, to enhance virus transmission and pathology.


Subject(s)
/metabolism , Respiratory System/metabolism , Respiratory System/virology , Spike Glycoprotein, Coronavirus/metabolism , /transmission , Coronavirus 229E, Human/metabolism , Furin/metabolism , Humans , Membrane Proteins/metabolism , Middle East Respiratory Syndrome Coronavirus/metabolism , Peptide Hydrolases/metabolism , Serine Endopeptidases/metabolism , Spike Glycoprotein, Coronavirus/genetics , Temperature , Virus Internalization , Virus Replication/physiology
7.
Top Curr Chem (Cham) ; 379(3): 23, 2021 Apr 22.
Article in English | MEDLINE | ID: covidwho-1196651

ABSTRACT

Coronavirus disease 2019, caused by the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), is still a pandemic around the world. Currently, specific antiviral drugs to control the epidemic remain deficient. Understanding the details of SARS-CoV-2 structural biology is extremely important for development of antiviral agents that will enable regulation of its life cycle. This review focuses on the structural biology and medicinal chemistry of various key proteins (Spike, ACE2, TMPRSS2, RdRp and Mpro) in the life cycle of SARS-CoV-2, as well as their inhibitors/drug candidates. Representative broad-spectrum antiviral drugs, especially those against the homologous virus SARS-CoV, are summarized with the expectation they will drive the development of effective, broad-spectrum inhibitors against coronaviruses. We are hopeful that this review will be a useful aid for discovery of novel, potent anti-SARS-CoV-2 drugs with excellent therapeutic results in the near future.


Subject(s)
Antiviral Agents/chemistry , Spike Glycoprotein, Coronavirus/chemistry , Viral Matrix Proteins/chemistry , /antagonists & inhibitors , Antiviral Agents/metabolism , Antiviral Agents/pharmacology , Antiviral Agents/therapeutic use , /pathology , Drug Repositioning , Humans , Serine Endopeptidases/chemistry , Serine Endopeptidases/metabolism , Spike Glycoprotein, Coronavirus/metabolism , Viral Matrix Proteins/metabolism , Virus Internalization/drug effects
8.
Gac Med Mex ; 156(6): 570-575, 2020.
Article in English | MEDLINE | ID: covidwho-1194843

ABSTRACT

SARS-CoV-2 virus has been identified as the causative agent of the COVID-19 pandemic. Even when no standard treatment is available, antivirals such as remdesivir and other drugs such as chloroquine and ivermectin, which interfere with viral replication, have been assayed. Some strategies aimed at reducing immune mechanisms, such as the use of tocilizumab and natural antioxidants, have also been tested. The use of drugs related to the renin-angiotensin system has been controversial. Pathogenicity mechanisms, as well as controlled treatments, still have to be studied in detail in order to propose a viable therapeutic option that prevents the entry and replication of the virus or enhances the host immune system.El virus SARS-CoV-2 ha sido identificado como el agente patológico causante de la pandemia de COVID-19. Aun cuando no se cuenta con un tratamiento estándar, se han probado antivirales como remdesivir y otros fármacos como cloroquina e ivermectina, que interfieren con la replicación del virus. También se han intentado algunas estrategias encaminadas a disminuir los mecanismos inmunitarios, como el uso de tocilizumab y antioxidantes naturales. Los fármacos relacionados con el sistema renina-angiotensina han resultado controversiales. Aún se debe estudiar con detalle los mecanismos de patogenicidad, así como los tratamientos controlados para proponer alguna opción terapéutica viable que evite la entrada y replicación del virus o que aumente los sistemas inmunitarios del huésped.


Subject(s)
Antiviral Agents/administration & dosage , /drug therapy , Animals , Antiviral Agents/pharmacology , Humans , /isolation & purification , Virus Internalization/drug effects , Virus Replication/drug effects
9.
Sci Adv ; 7(16)2021 04.
Article in English | MEDLINE | ID: covidwho-1189804

ABSTRACT

The COVID-19 (coronavirus disease 2019) pandemic underwent a rapid transition with the emergence of a dominant viral variant (from the "D-form" to the "G-form") that carried an amino acid substitution D614G in its "Spike" protein. The G-form is more infectious in vitro and is associated with increased viral loads in the upper airways. To gain insight into the molecular-level underpinnings of these characteristics, we used microsecond all-atom simulations. We show that changes in the protein energetics favor a higher population of infection-capable states in the G-form through release of asymmetry present in the D-form inter-protomer interactions. Thus, the increased infectivity of the G-form is likely due to a higher rate of profitable binding encounters with the host receptor. It is also predicted to be more neutralization sensitive owing to enhanced exposure of the receptor binding domain, a key target region for neutralizing antibodies. These results are critical for vaccine design.


Subject(s)
/genetics , Spike Glycoprotein, Coronavirus/chemistry , Amino Acid Sequence , /metabolism , Antibodies, Neutralizing/immunology , /virology , Glycosylation , Humans , Hydrogen Bonding , Molecular Dynamics Simulation , Mutation , Protein Binding , Protein Structure, Quaternary , Protein Subunits/chemistry , Protein Subunits/immunology , /isolation & purification , Spike Glycoprotein, Coronavirus/metabolism , Virus Internalization
10.
Circ Res ; 128(8): 1214-1236, 2021 04 16.
Article in English | MEDLINE | ID: covidwho-1186415

ABSTRACT

A pandemic of historic impact, coronavirus disease 2019 (COVID-19) has potential consequences on the cardiovascular health of millions of people who survive infection worldwide. Severe acute respiratory syndrome-coronavirus 2 (SARS-CoV-2), the etiologic agent of COVID-19, can infect the heart, vascular tissues, and circulating cells through ACE2 (angiotensin-converting enzyme 2), the host cell receptor for the viral spike protein. Acute cardiac injury is a common extrapulmonary manifestation of COVID-19 with potential chronic consequences. This update provides a review of the clinical manifestations of cardiovascular involvement, potential direct SARS-CoV-2 and indirect immune response mechanisms impacting the cardiovascular system, and implications for the management of patients after recovery from acute COVID-19 infection.


Subject(s)
/metabolism , Cardiovascular Diseases/virology , Myocytes, Cardiac/virology , Virus Internalization , Biomarkers/metabolism , /epidemiology , Cardiomyopathies/virology , Gene Expression , Humans , Immune System/physiology , Myocardium/enzymology , Myocytes, Cardiac/enzymology , Neuropilin-1/metabolism , Platelet Activation , RNA, Messenger/metabolism , Renin-Angiotensin System/physiology , Return to Sport , Risk Factors , Spike Glycoprotein, Coronavirus/metabolism , Troponin/metabolism , Ventricular Remodeling , Virus Attachment , Virus Internalization/drug effects
11.
mSphere ; 6(2)2021 04 14.
Article in English | MEDLINE | ID: covidwho-1186210

ABSTRACT

The coronavirus disease 2019 (COVID-19) pandemic, caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), has had a massive impact on human lives worldwide. While the airborne SARS-CoV-2 primarily affects the lungs, viremia is not uncommon. As placental trophoblasts are directly bathed in maternal blood, they are vulnerable to SARS-CoV-2. Intriguingly, the human fetus is largely spared from SARS-CoV-2 infection. We tested whether the human placenta expresses the main SARS-CoV-2 entry factors angiotensin-converting enzyme 2 (ACE2), transmembrane protease serine 2 (TMPRSS2), and furin and showed that ACE2 and TMPRSS2 are expressed in the trophoblast rather than in other placental villous cells. While furin is expressed in the main placental villous cell types, we surveyed, trophoblasts exhibit the highest expression. In line with the expression of these entry factors, we demonstrated that a SARS-CoV-2 pseudovirus could enter primary human trophoblasts. Mechanisms underlying placental defense against SARS-CoV-2 infection likely involve postentry processing, which may be germane for mitigating interventions against SARS-CoV-2.IMPORTANCE Pregnant women worldwide have been affected by COVID-19. As the virus is commonly spread to various organs via the bloodstream and because human placental trophoblasts are directly bathed in maternal blood, feto-placental infection by SARS-CoV-2 seems likely. However, despite the heightened risk to pregnant women, thus far the transmission risk of COVID-19 to the feto-placental unit seems extremely low. This has been recently attributed to a negligible expression of SARS-CoV-2 entry factors in the human placenta. We therefore sought to explore the expression of the entry factors ACE2 and TMPRSS2 in the different cell types of human placental villi. Using a combination of transcriptome sequencing (RNA-seq), real-time quantitative PCR (RT-qPCR), in situ hybridization, and immunofluorescence, we found that trophoblasts, but not the other main villous cell types, express ACE2 and TMPRSS2, with a broad expression of furin. Correspondingly, we also showed that primary human trophoblasts are permissive to entry of SARS-CoV-2 pseudovirus particles.


Subject(s)
/metabolism , Furin/metabolism , Receptors, Virus/metabolism , Serine Endopeptidases/metabolism , Trophoblasts/metabolism , Cells, Cultured , Female , Fetus/virology , Humans , Pregnancy , Pregnancy Complications, Infectious/virology , Virus Internalization
12.
Sci Signal ; 14(675)2021 03 23.
Article in English | MEDLINE | ID: covidwho-1186203

ABSTRACT

Nicotinic acid adenine dinucleotide phosphate (NAADP) is a second messenger that releases Ca2+ from acidic organelles through the activation of two-pore channels (TPCs) to regulate endolysosomal trafficking events. NAADP action is mediated by NAADP-binding protein(s) of unknown identity that confer NAADP sensitivity to TPCs. Here, we used a "clickable" NAADP-based photoprobe to isolate human NAADP-binding proteins and identified Jupiter microtubule-associated homolog 2 (JPT2) as a TPC accessory protein required for endogenous NAADP-evoked Ca2+ signaling. JPT2 was also required for the translocation of a severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) pseudovirus through the endolysosomal system. Thus, JPT2 is a component of the NAADP receptor complex that is essential for TPC-dependent Ca2+ signaling and control of coronaviral entry.


Subject(s)
/metabolism , Calcium Signaling/physiology , Microtubule-Associated Proteins/metabolism , NADP/analogs & derivatives , /physiology , Affinity Labels , Animals , Calcium Channels/metabolism , Carrier Proteins/metabolism , Click Chemistry/methods , Gene Knockdown Techniques , HEK293 Cells , Humans , Microtubule-Associated Proteins/antagonists & inhibitors , Microtubule-Associated Proteins/genetics , NADP/metabolism , Recombinant Proteins/genetics , Recombinant Proteins/metabolism , Second Messenger Systems/physiology , Transcriptome , Virus Internalization
13.
J Gen Virol ; 102(4)2021 04.
Article in English | MEDLINE | ID: covidwho-1186013

ABSTRACT

SARS-CoV-2 is thought to have originated in the human population from a zoonotic spillover event. Infection in humans results in a variety of outcomes ranging from asymptomatic cases to the disease COVID-19, which can have significant morbidity and mortality, with over two million confirmed deaths worldwide as of January 2021. Over a year into the pandemic, sequencing analysis has shown that variants of SARS-CoV-2 are being selected as the virus continues to circulate widely within the human population. The predominant drivers of genetic variation within SARS-CoV-2 are single nucleotide polymorphisms (SNPs) caused by polymerase error, potential host factor driven RNA modification, and insertion/deletions (indels) resulting from the discontinuous nature of viral RNA synthesis. While many mutations represent neutral 'genetic drift' or have quickly died out, a subset may be affecting viral traits such as transmissibility, pathogenicity, host range, and antigenicity of the virus. In this review, we summarise the current extent of genetic change in SARS-CoV-2, particularly recently emerging variants of concern, and consider the phenotypic consequences of this viral evolution that may impact the future trajectory of the pandemic.


Subject(s)
Adaptation, Physiological/genetics , /genetics , Animals , Antigens, Viral/genetics , Antigens, Viral/immunology , /immunology , Humans , Immune Evasion/genetics , Mutation , RNA, Viral/biosynthesis , RNA, Viral/genetics , /pathogenicity , Spike Glycoprotein, Coronavirus/genetics , Spike Glycoprotein, Coronavirus/metabolism , Viral Proteins/genetics , Viral Proteins/metabolism , Virus Internalization , Virus Replication , Zoonoses/transmission , Zoonoses/virology
14.
mBio ; 12(2)2021 04 13.
Article in English | MEDLINE | ID: covidwho-1183286

ABSTRACT

Mammalian cells detect microbial molecules known as pathogen-associated molecular patterns (PAMPs) as indicators of potential infection. Upon PAMP detection, diverse defensive responses are induced by the host, including those that promote inflammation and cell-intrinsic antimicrobial activities. Host-encoded molecules released from dying or damaged cells, known as damage-associated molecular patterns (DAMPs), also induce defensive responses. Both DAMPs and PAMPs are recognized for their inflammatory potential, but only the latter are well established to stimulate cell-intrinsic host defense. Here, we report a class of DAMPs that engender an antiviral state in human epithelial cells. These DAMPs include oxPAPC (oxidized 1-palmitoyl-2-arachidonoyl-sn-glycero-3-phosphocholine), PGPC (1-palmitoyl-2-glutaryl phosphatidylcholine), and POVPC [1-palmitoyl-2-(5-oxovaleroyl)-sn-glycero-3-phosphatidylcholine], oxidized lipids that are naturally released from dead or dying cells. Exposing cells to these DAMPs prior to vesicular stomatitis virus (VSV) infection limits viral replication. Mechanistically, these DAMPs prevent viral entry, thereby limiting the percentage of cells that are productively infected and consequently restricting viral load. We found that the antiviral actions of oxidized lipids are distinct from those mediated by the PAMP Poly I:C, in that the former induces a more rapid antiviral response without the induction of the interferon response. These data support a model whereby interferon-independent defensive activities can be induced by DAMPs, which may limit viral replication before PAMP-mediated interferon responses are induced. This antiviral activity may impact viruses that disrupt interferon responses in the oxygenated environment of the lung, such as influenza virus and SARS-CoV-2.IMPORTANCE In this work, we explored how a class of oxidized lipids, spontaneously created during tissue damage and unprogrammed cell lysis, block the earliest events in RNA virus infection in the human epithelium. This gives us novel insight into the ways that we view infection models, unveiling a built-in mechanism to slow viral growth that neither engages the interferon response nor is subject to known viral antagonism. These oxidized phospholipids act prior to infection, allowing time for other, better-known innate immune mechanisms to take effect. This discovery broadens our understanding of host defenses, introducing a soluble factor that alters the cellular environment to protect from RNA virus infection.


Subject(s)
Alarmins/pharmacology , Antiviral Agents/pharmacology , RNA Viruses/drug effects , Virus Internalization/drug effects , Virus Replication/drug effects , A549 Cells , Cell Death/drug effects , Humans , Immunity, Innate , Interferons/genetics , Interferons/metabolism , Kinetics , Pathogen-Associated Molecular Pattern Molecules/pharmacology , Phosphatidylcholines/pharmacology , RNA Viruses/physiology , /physiology , Vesiculovirus/drug effects , Vesiculovirus/physiology , Viral Load
15.
Cell Rep ; 35(3): 109020, 2021 04 20.
Article in English | MEDLINE | ID: covidwho-1182447

ABSTRACT

COVID-19, caused by the novel coronavirus SARS-CoV-2, is a global health issue with more than 2 million fatalities to date. Viral replication is shaped by the cellular microenvironment, and one important factor to consider is oxygen tension, in which hypoxia inducible factor (HIF) regulates transcriptional responses to hypoxia. SARS-CoV-2 primarily infects cells of the respiratory tract, entering via its spike glycoprotein binding to angiotensin-converting enzyme 2 (ACE2). We demonstrate that hypoxia and the HIF prolyl hydroxylase inhibitor Roxadustat reduce ACE2 expression and inhibit SARS-CoV-2 entry and replication in lung epithelial cells via an HIF-1α-dependent pathway. Hypoxia and Roxadustat inhibit SARS-CoV-2 RNA replication, showing that post-entry steps in the viral life cycle are oxygen sensitive. This study highlights the importance of HIF signaling in regulating multiple aspects of SARS-CoV-2 infection and raises the potential use of HIF prolyl hydroxylase inhibitors in the prevention or treatment of COVID-19.


Subject(s)
/metabolism , Epithelial Cells/metabolism , Glycine/analogs & derivatives , Hypoxia-Inducible Factor 1, alpha Subunit/metabolism , Isoquinolines/pharmacology , Lung/metabolism , Virus Internalization/drug effects , Virus Replication/drug effects , A549 Cells , Animals , /pathology , Caco-2 Cells , Cell Hypoxia/drug effects , Chlorocebus aethiops , Epithelial Cells/virology , Glycine/pharmacology , Humans , Lung/virology , Mice , Vero Cells
16.
Emerg Microbes Infect ; 10(1): 810-821, 2021 Dec.
Article in English | MEDLINE | ID: covidwho-1180458

ABSTRACT

EK1 peptide is a membrane fusion inhibitor with broad-spectrum activity against human coronaviruses (CoVs). In the outbreak of COVID-19, we generated a lipopeptide EK1V1 by modifying EK1 with cholesterol, which exhibited significantly improved antiviral activity. In this study, we surprisingly found that EK1V1 also displayed potent cross-inhibitory activities against divergent HIV-1, HIV-2, and simian immunodeficiency virus (SIV) isolates. Consistently, the recently reported EK1 derivative EK1C4 and SARS-CoV-2 derived fusion inhibitor lipopeptides (IPB02 ∼ IPB09) also inhibited HIV-1 Env-mediated cell-cell fusion and infection efficiently. In the inhibition of a panel of HIV-1 mutants resistant to HIV-1 fusion inhibitors, EK1V1 and IPB02-based inhibitors exhibited significantly decreased or increased activities, suggesting the heptad repeat-1 region (HR1) of HIV-1 gp41 being their target. Furthermore, the sequence alignment and molecular docking analyses verified the target site and revealed the mechanism underlying the resistance. Combined, we conclude that this serendipitous discovery provides a proof-of-concept for a common mechanism of viral fusion and critical information for the development of broad-spectrum antivirals.


Subject(s)
Antiviral Agents/pharmacology , Coronavirus/drug effects , HIV-1/drug effects , HIV-2/drug effects , Simian Immunodeficiency Virus/drug effects , Virus Internalization/drug effects , Amino Acid Sequence , Animals , Antiviral Agents/isolation & purification , Dose-Response Relationship, Drug , HIV Fusion Inhibitors/isolation & purification , HIV Fusion Inhibitors/pharmacology , Humans , Lipopeptides/isolation & purification , Lipopeptides/pharmacology , Molecular Docking Simulation , Molecular Dynamics Simulation , Peptide Fragments/isolation & purification , Peptide Fragments/pharmacology , Structure-Activity Relationship , Virus Replication/drug effects
17.
Commun Biol ; 4(1): 475, 2021 04 12.
Article in English | MEDLINE | ID: covidwho-1180282

ABSTRACT

COVID-19 is a respiratory illness caused by a novel coronavirus called SARS-CoV-2. The viral spike (S) protein engages the human angiotensin-converting enzyme 2 (ACE2) receptor to invade host cells with ~10-15-fold higher affinity compared to SARS-CoV S-protein, making it highly infectious. Here, we assessed if ACE2 polymorphisms can alter host susceptibility to SARS-CoV-2 by affecting this interaction. We analyzed over 290,000 samples representing >400 population groups from public genomic datasets and identified multiple ACE2 protein-altering variants. Using reported structural data, we identified natural ACE2 variants that could potentially affect virus-host interaction and thereby alter host susceptibility. These include variants S19P, I21V, E23K, K26R, T27A, N64K, T92I, Q102P and H378R that were predicted to increase susceptibility, while variants K31R, N33I, H34R, E35K, E37K, D38V, Y50F, N51S, M62V, K68E, F72V, Y83H, G326E, G352V, D355N, Q388L and D509Y were predicted to be protective variants that show decreased binding to S-protein. Using biochemical assays, we confirmed that K31R and E37K had decreased affinity, and K26R and T92I variants showed increased affinity for S-protein when compared to wildtype ACE2. Consistent with this, soluble ACE2 K26R and T92I were more effective in blocking entry of S-protein pseudotyped virus suggesting that ACE2 variants can modulate susceptibility to SARS-CoV-2.


Subject(s)
/genetics , Genetic Predisposition to Disease/genetics , Mutation, Missense/genetics , Polymorphism, Genetic , Receptors, Virus/genetics , Amino Acid Sequence , /metabolism , /virology , Host-Pathogen Interactions , Humans , Models, Molecular , Protein Binding , Protein Domains , Receptors, Virus/chemistry , Receptors, Virus/metabolism , /physiology , Sequence Homology, Amino Acid , Spike Glycoprotein, Coronavirus/chemistry , Spike Glycoprotein, Coronavirus/metabolism , Virus Internalization
19.
Clin Sci (Lond) ; 134(21): 2851-2871, 2020 11 13.
Article in English | MEDLINE | ID: covidwho-1177131

ABSTRACT

Angiotensin converting enzyme (ACE) is well-known for its role in blood pressure regulation via the renin-angiotensin aldosterone system (RAAS) but also functions in fertility, immunity, haematopoiesis and diseases such as obesity, fibrosis and Alzheimer's dementia. Like ACE, the human homologue ACE2 is also involved in blood pressure regulation and cleaves a range of substrates involved in different physiological processes. Importantly, it is the functional receptor for severe acute respiratory syndrome (SARS)-coronavirus (CoV)-2 responsible for the 2020, coronavirus infectious disease 2019 (COVID-19) pandemic. Understanding the interaction between SARS-CoV-2 and ACE2 is crucial for the design of therapies to combat this disease. This review provides a comparative analysis of methodologies and findings to describe how structural biology techniques like X-ray crystallography and cryo-electron microscopy have enabled remarkable discoveries into the structure-function relationship of ACE and ACE2. This, in turn, has enabled the development of ACE inhibitors for the treatment of cardiovascular disease and candidate therapies for the treatment of COVID-19. However, despite these advances the function of ACE homologues in non-human organisms is not yet fully understood. ACE homologues have been discovered in the tissues, body fluids and venom of species from diverse lineages and are known to have important functions in fertility, envenoming and insect-host defence mechanisms. We, therefore, further highlight the need for structural insight into insect and venom ACE homologues for the potential development of novel anti-venoms and insecticides.


Subject(s)
Betacoronavirus/pathogenicity , Coronavirus Infections/enzymology , Peptidyl-Dipeptidase A/metabolism , Pneumonia, Viral/enzymology , Receptors, Virus/metabolism , Virus Internalization , Angiotensin-Converting Enzyme Inhibitors/therapeutic use , Animals , Antiviral Agents/therapeutic use , Betacoronavirus/drug effects , Coronavirus Infections/drug therapy , Coronavirus Infections/virology , Host-Pathogen Interactions , Humans , Pandemics , Peptidyl-Dipeptidase A/chemistry , Pneumonia, Viral/drug therapy , Pneumonia, Viral/virology , Protein Conformation , Receptors, Virus/chemistry , Structure-Activity Relationship
20.
Front Immunol ; 12: 613045, 2021.
Article in English | MEDLINE | ID: covidwho-1177974

ABSTRACT

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) initiates infection by attachment of the surface-exposed spike glycoprotein to the host cell receptors. The spike glycoprotein (S) is a promising target for inducing immune responses and providing protection; thus the ongoing efforts for the SARS-CoV-2 vaccine and therapeutic developments are mostly spiraling around S glycoprotein. The matured functional spike glycoprotein is presented on the virion surface as trimers, which contain two subunits, such as S1 (virus attachment) and S2 (virus fusion). The S1 subunit harbors the N-terminal domain (NTD) and the receptor-binding domain (RBD). The RBD is responsible for binding to host-cellular receptor angiotensin-converting enzyme 2 (ACE2). The NTD and RBD of S1, and the S2 of S glycoprotein are the major structural moieties to design and develop spike-based vaccine candidates and therapeutics. Here, we have identified three novel epitopes (20-amino acid peptides) in the regions NTD, RBD, and S2 domains, respectively, by structural and immunoinformatic analysis. We have shown as a proof of principle in the murine model, the potential role of these novel epitopes in-inducing humoral and cellular immune responses. Further analysis has shown that RBD and S2 directed epitopes were able to efficiently inhibit the replication of SARS-CoV-2 wild-type virus in vitro suggesting their role as virus entry inhibitors. Structural analysis revealed that S2-epitope is a part of the heptad repeat 2 (HR2) domain which might have plausible inhibitory effects on virus fusion. Taken together, this study discovered novel epitopes that might have important implications in the development of potential SARS-CoV-2 spike-based vaccine and therapeutics.


Subject(s)
Epitopes/immunology , Spike Glycoprotein, Coronavirus/immunology , Virus Replication/immunology , Animals , Female , Humans , Mice , Mice, Inbred BALB C , Protein Domains , Virus Internalization
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