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1.
bioRxiv ; 2024 Jul 10.
Artigo em Inglês | MEDLINE | ID: mdl-39026801

RESUMO

Defining the subset of cellular factors governing SARS-CoV-2 replication can provide critical insights into viral pathogenesis and identify targets for host-directed antiviral therapies. While a number of genetic screens have previously reported SARS-CoV-2 host dependency factors, these approaches relied on utilizing pooled genome-scale CRISPR libraries, which are biased towards the discovery of host proteins impacting early stages of viral replication. To identify host factors involved throughout the SARS-CoV-2 infectious cycle, we conducted an arrayed genome-scale siRNA screen. Resulting data were integrated with published datasets to reveal pathways supported by orthogonal datasets, including transcriptional regulation, epigenetic modifications, and MAPK signalling. The identified proviral host factors were mapped into the SARS-CoV-2 infectious cycle, including 27 proteins that were determined to impact assembly and release. Additionally, a subset of proteins were tested across other coronaviruses revealing 17 potential pan-coronavirus targets. Further studies illuminated a role for the heparan sulfate proteoglycan perlecan in SARS-CoV-2 viral entry, and found that inhibition of the non-canonical NF-kB pathway through targeting of BIRC2 restricts SARS-CoV-2 replication both in vitro and in vivo. These studies provide critical insight into the landscape of virus-host interactions driving SARS-CoV-2 replication as well as valuable targets for host-directed antivirals.

3.
Nat Commun ; 15(1): 1853, 2024 Feb 29.
Artigo em Inglês | MEDLINE | ID: mdl-38424040

RESUMO

Many machine learning applications in bioinformatics currently rely on matching gene identities when analyzing input gene signatures and fail to take advantage of preexisting knowledge about gene functions. To further enable comparative analysis of OMICS datasets, including target deconvolution and mechanism of action studies, we develop an approach that represents gene signatures projected onto their biological functions, instead of their identities, similar to how the word2vec technique works in natural language processing. We develop the Functional Representation of Gene Signatures (FRoGS) approach by training a deep learning model and demonstrate that its application to the Broad Institute's L1000 datasets results in more effective compound-target predictions than models based on gene identities alone. By integrating additional pharmacological activity data sources, FRoGS significantly increases the number of high-quality compound-target predictions relative to existing approaches, many of which are supported by in silico and/or experimental evidence. These results underscore the general utility of FRoGS in machine learning-based bioinformatics applications. Prediction networks pre-equipped with the knowledge of gene functions may help uncover new relationships among gene signatures acquired by large-scale OMICs studies on compounds, cell types, disease models, and patient cohorts.


Assuntos
Aprendizado Profundo , Humanos , Aprendizado de Máquina , Biologia Computacional , Desenvolvimento de Medicamentos
4.
J Mol Biol ; 436(4): 168409, 2024 02 15.
Artigo em Inglês | MEDLINE | ID: mdl-38128824

RESUMO

Human immunodeficiency virus type 1 (HIV-1) stimulates innate immune responses upon infection, including cyclic GMP-AMP synthase (cGAS) signaling that results in type I interferon production. HIV-1-induced activation of cGAS requires the host cell factor polyglutamine binding protein 1 (PQBP1), an intrinsically disordered protein that bridges capsid recognition and cGAS recruitment. However, the molecular details of PQBP1 interactions with the HIV-1 capsid and their functional implications remain poorly understood. Here, we show that PQBP1 binds to HIV-1 capsids through charge complementing contacts between acidic residues in the N-terminal region of PQBP1 and an arginine ring in the central channel of the HIV-1 CA hexamer that makes up the viral capsid. These studies reveal the molecular details of PQBP1's primary interaction with the HIV-1 capsid and suggest that additional elements are likely to contribute to stable capsid binding.


Assuntos
Capsídeo , Proteínas de Ligação a DNA , HIV-1 , Humanos , Capsídeo/química , Proteínas do Capsídeo/química , Proteínas de Ligação a DNA/química , HIV-1/química , Imunidade Inata , Nucleotidiltransferases/química , Ligação Proteica , Conformação Proteica
5.
Pathogens ; 12(12)2023 Nov 30.
Artigo em Inglês | MEDLINE | ID: mdl-38133292

RESUMO

As part of the non-clinical safety package characterizing bamlanivimab (SARS-CoV-2 neutralizing monoclonal antibody), the risk profile for antibody-dependent enhancement of infection (ADE) was evaluated in vitro and in an African green monkey (AGM) model of COVID-19. In vitro ADE assays in primary human macrophage, Raji, or THP-1 cells were used to evaluate enhancement of viral infection. Bamlanivimab binding to C1q, FcR, and cell-based effector activity was also assessed. In AGMs, the impact of bamlanivimab pretreatment on viral loads and clinical and histological pathology was assessed to evaluate enhanced SARS-CoV-2 replication or pathology. Bamlanivimab did not increase viral replication in vitro, despite a demonstrated effector function. In vivo, no significant differences were found among the AGM groups for weight, temperature, or food intake. Treatment with bamlanivimab reduced viral loads in nasal and oral swabs and BAL fluid relative to control groups. Viral antigen was not detected in lung tissue from animals treated with the highest dose of bamlanivimab. Bamlanivimab did not induce ADE of SARS-CoV-2 infection in vitro or in an AGM model of infection at any dose evaluated. The findings suggest that high-affinity monoclonal antibodies pose a low risk of mediating ADE in patients and support their safety profile as a treatment of COVID-19 disease.

6.
Viruses ; 15(12)2023 12 15.
Artigo em Inglês | MEDLINE | ID: mdl-38140676

RESUMO

Antiretroviral therapy (ART) has brought the HIV/AIDS epidemic under control, but a curative strategy for viral eradication is still needed. The cessation of ART results in rapid viral rebound from latently infected CD4+ T cells, showing that control of viral replication alone does not fully restore immune function, nor does it eradicate viral reservoirs. With a better understanding of factors and mechanisms that promote viral latency, current approaches are primarily focused on the permanent silencing of latently infected cells ("block and lock") or reactivating HIV-1 gene expression in latently infected cells, in combination with immune restoration strategies to eliminate HIV infected cells from the host ("shock and kill"). In this review, we provide a summary of the current, most promising approaches for HIV-1 cure strategies, including an analysis of both latency-promoting agents (LPA) and latency-reversing agents (LRA) that have shown promise in vitro, ex vivo, and in human clinical trials to reduce the HIV-1 reservoir.


Assuntos
Infecções por HIV , Soropositividade para HIV , HIV-1 , Humanos , Latência Viral , Replicação Viral , HIV-1/fisiologia , Linfócitos T CD4-Positivos , Ativação Viral
7.
Nat Commun ; 14(1): 6030, 2023 09 27.
Artigo em Inglês | MEDLINE | ID: mdl-37758692

RESUMO

Influenza A Virus (IAV) is a recurring respiratory virus with limited availability of antiviral therapies. Understanding host proteins essential for IAV infection can identify targets for alternative host-directed therapies (HDTs). Using affinity purification-mass spectrometry and global phosphoproteomic and protein abundance analyses using three IAV strains (pH1N1, H3N2, H5N1) in three human cell types (A549, NHBE, THP-1), we map 332 IAV-human protein-protein interactions and identify 13 IAV-modulated kinases. Whole exome sequencing of patients who experienced severe influenza reveals several genes, including scaffold protein AHNAK, with predicted loss-of-function variants that are also identified in our proteomic analyses. Of our identified host factors, 54 significantly alter IAV infection upon siRNA knockdown, and two factors, AHNAK and coatomer subunit COPB1, are also essential for productive infection by SARS-CoV-2. Finally, 16 compounds targeting our identified host factors suppress IAV replication, with two targeting CDK2 and FLT3 showing pan-antiviral activity across influenza and coronavirus families. This study provides a comprehensive network model of IAV infection in human cells, identifying functional host targets for pan-viral HDT.


Assuntos
COVID-19 , Virus da Influenza A Subtipo H5N1 , Vírus da Influenza A , Influenza Humana , Humanos , Vírus da Influenza A/genética , Influenza Humana/genética , Virus da Influenza A Subtipo H5N1/genética , Vírus da Influenza A Subtipo H3N2/metabolismo , Proteômica , Replicação Viral/genética , SARS-CoV-2 , Antivirais/metabolismo , Interações Hospedeiro-Patógeno/genética
8.
Res Sq ; 2023 Sep 08.
Artigo em Inglês | MEDLINE | ID: mdl-37720034

RESUMO

SARS-CoV-2 initiates infection in the conducting airways, which rely on mucocilliary clearance (MCC) to minimize pathogen penetration. However, it is unclear how MCC impacts SARS-CoV-2 spread after infection is established. To understand viral spread at this site, we performed live imaging of SARS-CoV-2 infected differentiated primary human bronchial epithelium cultures for up to 9 days. Fluorescent markers for cilia and mucus allowed longitudinal monitoring of MCC, ciliary motion, and infection. The number of infected cells peaked at 4 days post-infection in characteristic foci that followed mucus movement. Inhibition of MCC using physical and genetic perturbations limited foci. Later in infection, MCC was diminished despite relatively subtle ciliary function defects. Resumption of MCC and infection spread after mucus removal suggests that mucus secretion mediates this effect. We show that MCC facilitates SARS-CoV-2 spread early in infection while later decreases in MCC inhibit spread, suggesting a complex interplay between SARS-CoV-2 and MCC.

9.
Nat Commun ; 14(1): 948, 2023 02 20.
Artigo em Inglês | MEDLINE | ID: mdl-36804936

RESUMO

Small molecule inhibitors of glycosylation enzymes are valuable tools for dissecting glycan functions and potential drug candidates. Screening for inhibitors of glycosyltransferases are mainly performed by in vitro enzyme assays with difficulties moving candidates to cells and animals. Here, we circumvent this by employing a cell-based screening assay using glycoengineered cells expressing tailored reporter glycoproteins. We focused on GalNAc-type O-glycosylation and selected the GalNAc-T11 isoenzyme that selectively glycosylates endocytic low-density lipoprotein receptor (LDLR)-related proteins as targets. Our screen of a limited small molecule compound library did not identify selective inhibitors of GalNAc-T11, however, we identify two compounds that broadly inhibited Golgi-localized glycosylation processes. These compounds mediate the reversible fragmentation of the Golgi system without affecting secretion. We demonstrate how these inhibitors can be used to manipulate glycosylation in cells to induce expression of truncated O-glycans and augment binding of cancer-specific Tn-glycoprotein antibodies and to inhibit expression of heparan sulfate and binding and infection of SARS-CoV-2.


Assuntos
COVID-19 , SARS-CoV-2 , Animais , Glicosilação , SARS-CoV-2/metabolismo , Glicoproteínas/metabolismo , Polissacarídeos/metabolismo
10.
Stem Cells Dev ; 32(9-10): 225-236, 2023 05.
Artigo em Inglês | MEDLINE | ID: mdl-36803114

RESUMO

SARS-CoV-2 infection during pregnancy has been associated with poor maternal and neonatal outcomes and placental defects. The placenta, which acts as a physical and immunological barrier at the maternal-fetal interface, is not established until the end of the first trimester. Therefore, localized viral infection of the trophoblast compartment early in gestation could trigger an inflammatory response resulting in altered placental function and consequent suboptimal conditions for fetal growth and development. In this study, we investigated the effect of SARS-CoV-2 infection in early gestation placentae using placenta-derived human trophoblast stem cells (TSCs), a novel in vitro model, and their extravillous trophoblast (EVT) and syncytiotrophoblast (STB) derivatives. SARS-CoV-2 was able to productively replicate in TSC-derived STB and EVT, but not undifferentiated TSCs, which is consistent with the expression of SARS-CoV-2 entry host factors, ACE2 (angiotensin-converting enzyme 2) and TMPRSS2 (transmembrane cellular serine protease) in these cells. In addition, both TSC-derived EVT and STB infected with SARS-CoV-2 elicited an interferon-mediated innate immune response. Combined, these results suggest that placenta-derived TSCs are a robust in vitro model to investigate the effect of SARS-CoV-2 infection in the trophoblast compartment of the early placenta and that SARS-CoV-2 infection in early gestation activates the innate immune response and inflammation pathways. Therefore, placental development could be adversely affected by early SARS-CoV-2 infection by directly infecting the developing differentiated trophoblast compartment, posing a higher risk for poor pregnancy outcomes.


Assuntos
COVID-19 , SARS-CoV-2 , Recém-Nascido , Gravidez , Feminino , Humanos , COVID-19/metabolismo , Trofoblastos/metabolismo , Interferons , Placenta
11.
Front Pharmacol ; 13: 1029093, 2022.
Artigo em Inglês | MEDLINE | ID: mdl-36532738

RESUMO

The translation initiation complex 4F (eIF4F) is a rate-limiting factor in protein synthesis. Alterations in eIF4F activity are linked to several diseases, including cancer and infectious diseases. To this end, coronaviruses require eIF4F complex activity to produce proteins essential for their life cycle. Efforts to target coronaviruses by abrogating translation have been largely limited to repurposing existing eIF4F complex inhibitors. Here, we report the results of a high throughput screen to identify small molecules that disrupt eIF4F complex formation and inhibit coronavirus RNA and protein levels. Of 338,000 small molecules screened for inhibition of the eIF4F-driven, CAP-dependent translation, we identified SBI-1232 and two structurally related analogs, SBI-5844 and SBI-0498, that inhibit human coronavirus OC43 (HCoV-OC43; OC43) with minimal cell toxicity. Notably, gene expression changes after OC43 infection of Vero E6 or A549 cells were effectively reverted upon treatment with SBI-5844 or SBI-0498. Moreover, SBI-5844 or SBI-0498 treatment effectively impeded the eIF4F complex assembly, with concomitant inhibition of newly synthesized OC43 nucleocapsid protein and OC43 RNA and protein levels. Overall, we identify SBI-5844 and SBI-0498 as small molecules targeting the eIF4F complex that may limit coronavirus transcripts and proteins, thereby representing a basis for developing novel therapeutic modalities against coronaviruses.

12.
PLoS Biol ; 20(11): e3001845, 2022 11.
Artigo em Inglês | MEDLINE | ID: mdl-36327326

RESUMO

The Severe Acute Respiratory Syndrome Coronavirus 2 (SARS-CoV-2) is the causative agent of coronavirus disease 2019 (COVID-19), which was rapidly declared a pandemic by the World Health Organization (WHO). Early clinical symptomatology focused mainly on respiratory illnesses. However, a variety of neurological manifestations in both adults and newborns are now well-documented. To experimentally determine whether SARS-CoV-2 could replicate in and affect human brain cells, we infected iPSC-derived human brain organoids. Here, we show that SARS-CoV-2 can productively replicate and promote death of neural cells, including cortical neurons. This phenotype was accompanied by loss of excitatory synapses in neurons. Notably, we found that the U.S. Food and Drug Administration (FDA)-approved antiviral Sofosbuvir was able to inhibit SARS-CoV-2 replication and rescued these neuronal alterations in infected brain organoids. Given the urgent need for readily available antivirals, these results provide a cellular basis supporting repurposed antivirals as a strategic treatment to alleviate neurocytological defects that may underlie COVID-19- related neurological symptoms.


Assuntos
Tratamento Farmacológico da COVID-19 , SARS-CoV-2 , Recém-Nascido , Humanos , Sofosbuvir/farmacologia , Sofosbuvir/uso terapêutico , Organoides , Antivirais/farmacologia , Antivirais/uso terapêutico , Encéfalo , Morte Celular , Sinapses
13.
Sci Adv ; 8(40): eabm5859, 2022 10 07.
Artigo em Inglês | MEDLINE | ID: mdl-36197970

RESUMO

Molecular responses to influenza A virus (IAV) infections vary between mammalian species. To identify conserved and species-specific molecular responses, we perform a comparative study of transcriptomic data derived from blood cells, primary epithelial cells, and lung tissues collected from IAV-infected humans, ferrets, and mice. The molecular responses in the human host have unique functions such as antigen processing that are not observed in mice or ferrets. Highly conserved gene coexpression modules across the three species are enriched for IAV infection-induced pathways including cell cycle and interferon (IFN) signaling. TDRD7 is predicted as an IFN-inducible host factor that is up-regulated upon IAV infection in the three species. TDRD7 is required for antiviral IFN response, potentially modulating IFN signaling via the JAK/STAT/IRF9 pathway. Identification of the common and species-specific molecular signatures, networks, and regulators of IAV infection provides insights into host-defense mechanisms and will facilitate the development of novel therapeutic interventions against IAV infection.


Assuntos
Doenças Transmissíveis , Vírus da Influenza A , Influenza Humana , Infecções por Orthomyxoviridae , Animais , Antivirais , Furões/metabolismo , Humanos , Vírus da Influenza A/fisiologia , Influenza Humana/genética , Interferons/metabolismo , Camundongos , Infecções por Orthomyxoviridae/genética , Ribonucleoproteínas
14.
Mol Cell ; 82(15): 2871-2884.e6, 2022 08 04.
Artigo em Inglês | MEDLINE | ID: mdl-35809572

RESUMO

We have previously described polyglutamine-binding protein 1 (PQBP1) as an adapter required for the cyclic GMP-AMP synthase (cGAS)-mediated innate response to the human immunodeficiency virus 1 (HIV-1) and other lentiviruses. Cytoplasmic HIV-1 DNA is a transient and low-abundance pathogen-associated molecular pattern (PAMP), and the mechanism for its detection and verification is not fully understood. Here, we show a two-factor authentication strategy by the innate surveillance machinery to selectively respond to the low concentration of HIV-1 DNA, while distinguishing these species from extranuclear DNA molecules. We find that, upon HIV-1 infection, PQBP1 decorates the intact viral capsid, and this serves as a primary verification step for the viral nucleic acid cargo. As reverse transcription and capsid disassembly initiate, cGAS is recruited to the capsid in a PQBP1-dependent manner. This positions cGAS at the site of PAMP generation and sanctions its response to a low-abundance DNA PAMP.


Assuntos
HIV-1 , Capsídeo/metabolismo , DNA/metabolismo , Proteínas de Ligação a DNA/genética , Proteínas de Ligação a DNA/metabolismo , HIV-1/genética , Humanos , Imunidade Inata , Nucleotidiltransferases/metabolismo , Moléculas com Motivos Associados a Patógenos/metabolismo
15.
ACS Infect Dis ; 8(7): 1265-1279, 2022 07 08.
Artigo em Inglês | MEDLINE | ID: mdl-35766385

RESUMO

There is a pressing need for host-directed therapeutics that elicit broad-spectrum antiviral activities to potentially address current and future viral pandemics. Apratoxin S4 (Apra S4) is a potent Sec61 inhibitor that prevents cotranslational translocation of secretory proteins into the endoplasmic reticulum (ER), leading to anticancer and antiangiogenic activity both in vitro and in vivo. Since Sec61 has been shown to be an essential host factor for viral proteostasis, we tested Apra S4 in cellular models of viral infection, including SARS-CoV-2, influenza A virus, and flaviviruses (Zika, West Nile, and Dengue virus). Apra S4 inhibited viral replication in a concentration-dependent manner and had high potency particularly against SARS-CoV-2 and influenza A virus, with subnanomolar activity in human cells. Characterization studies focused on SARS-CoV-2 revealed that Apra S4 impacted a post-entry stage of the viral life-cycle. Transmission electron microscopy revealed that Apra S4 blocked formation of stacked double-membrane vesicles, the sites of viral replication. Apra S4 reduced dsRNA formation and prevented viral protein production and trafficking of secretory proteins, especially the spike protein. Given the potent and broad-spectrum activity of Apra S4, further preclinical evaluation of Apra S4 and other Sec61 inhibitors as antivirals is warranted.


Assuntos
Tratamento Farmacológico da COVID-19 , Vírus da Influenza A , Infecção por Zika virus , Zika virus , Antivirais/farmacologia , Antivirais/uso terapêutico , Depsipeptídeos , Humanos , Pandemias , SARS-CoV-2 , Infecção por Zika virus/tratamento farmacológico
16.
iScience ; 25(5): 104311, 2022 May 20.
Artigo em Inglês | MEDLINE | ID: mdl-35502318

RESUMO

Novel strategies are needed to identify drug targets and treatments for the COVID-19 pandemic. The altered gene expression of virus-infected host cells provides an opportunity to specifically inhibit viral propagation via targeting the synthetic lethal and synthetic dosage lethal (SL/SDL) partners of such altered host genes. Pursuing this disparate antiviral strategy, here we comprehensively analyzed multiple in vitro and in vivo bulk and single-cell RNA-sequencing datasets of SARS-CoV-2 infection to predict clinically relevant candidate antiviral targets that are SL/SDL with altered host genes. The predicted SL/SDL-based targets are highly enriched for infected cell inhibiting genes reported in four SARS-CoV-2 CRISPR-Cas9 genome-wide genetic screens. We further selected a focused subset of 26 genes that we experimentally tested in a targeted siRNA screen using human Caco-2 cells. Notably, as predicted, knocking down these targets reduced viral replication and cell viability only under the infected condition without harming noninfected healthy cells.

17.
PLoS Pathog ; 17(11): e1009409, 2021 11.
Artigo em Inglês | MEDLINE | ID: mdl-34843601

RESUMO

The HIV-1 accessory protein Vpu modulates membrane protein trafficking and degradation to provide evasion of immune surveillance. Targets of Vpu include CD4, HLAs, and BST-2. Several cellular pathways co-opted by Vpu have been identified, but the picture of Vpu's itinerary and activities within membrane systems remains incomplete. Here, we used fusion proteins of Vpu and the enzyme ascorbate peroxidase (APEX2) to compare the ultrastructural locations and the proximal proteomes of wild type Vpu and Vpu-mutants. The proximity-omes of the proteins correlated with their ultrastructural locations and placed wild type Vpu near both retromer and ESCRT-0 complexes. Hierarchical clustering of protein abundances across the mutants was essential to interpreting the data and identified Vpu degradation-targets including CD4, HLA-C, and SEC12 as well as Vpu-cofactors including HGS, STAM, clathrin, and PTPN23, an ALIX-like protein. The Vpu-directed degradation of BST-2 was supported by STAM and PTPN23 and to a much lesser extent by the retromer subunits Vps35 and SNX3. PTPN23 also supported the Vpu-directed decrease in CD4 at the cell surface. These data suggest that Vpu directs targets from sorting endosomes to degradation at multi-vesicular bodies via ESCRT-0 and PTPN23.


Assuntos
Complexos Endossomais de Distribuição Requeridos para Transporte/metabolismo , Infecções por HIV/virologia , Proteínas do Vírus da Imunodeficiência Humana/metabolismo , Proteínas Tirosina Fosfatases não Receptoras/metabolismo , Proteoma/metabolismo , Nexinas de Classificação/metabolismo , Proteínas de Transporte Vesicular/metabolismo , Proteínas Virais Reguladoras e Acessórias/metabolismo , Proteínas Viroporinas/metabolismo , Complexos Endossomais de Distribuição Requeridos para Transporte/genética , Infecções por HIV/genética , Infecções por HIV/metabolismo , HIV-1/fisiologia , Células HeLa , Proteínas do Vírus da Imunodeficiência Humana/genética , Humanos , Microscopia Eletrônica , Domínios e Motivos de Interação entre Proteínas , Multimerização Proteica , Transporte Proteico , Proteínas Tirosina Fosfatases não Receptoras/genética , Proteoma/análise , Nexinas de Classificação/química , Nexinas de Classificação/genética , Proteínas de Transporte Vesicular/química , Proteínas de Transporte Vesicular/genética , Proteínas Virais Reguladoras e Acessórias/genética , Proteínas Viroporinas/genética
18.
Mol Syst Biol ; 17(11): e10260, 2021 11.
Artigo em Inglês | MEDLINE | ID: mdl-34709707

RESUMO

Tremendous progress has been made to control the COVID-19 pandemic caused by the SARS-CoV-2 virus. However, effective therapeutic options are still rare. Drug repurposing and combination represent practical strategies to address this urgent unmet medical need. Viruses, including coronaviruses, are known to hijack host metabolism to facilitate viral proliferation, making targeting host metabolism a promising antiviral approach. Here, we describe an integrated analysis of 12 published in vitro and human patient gene expression datasets on SARS-CoV-2 infection using genome-scale metabolic modeling (GEM), revealing complicated host metabolism reprogramming during SARS-CoV-2 infection. We next applied the GEM-based metabolic transformation algorithm to predict anti-SARS-CoV-2 targets that counteract the virus-induced metabolic changes. We successfully validated these targets using published drug and genetic screen data and by performing an siRNA assay in Caco-2 cells. Further generating and analyzing RNA-sequencing data of remdesivir-treated Vero E6 cell samples, we predicted metabolic targets acting in combination with remdesivir, an approved anti-SARS-CoV-2 drug. Our study provides clinical data-supported candidate anti-SARS-CoV-2 targets for future evaluation, demonstrating host metabolism targeting as a promising antiviral strategy.


Assuntos
Monofosfato de Adenosina/análogos & derivados , Alanina/análogos & derivados , Antivirais/uso terapêutico , COVID-19/metabolismo , Redes e Vias Metabólicas/genética , Pandemias , SARS-CoV-2/fisiologia , Monofosfato de Adenosina/uso terapêutico , Alanina/uso terapêutico , Animais , COVID-19/virologia , Células CACO-2 , Chlorocebus aethiops , Conjuntos de Dados como Assunto , Desenvolvimento de Medicamentos , Reposicionamento de Medicamentos , Interações Hospedeiro-Patógeno , Humanos , RNA Interferente Pequeno , Análise de Sequência de RNA , Células Vero , Tratamento Farmacológico da COVID-19
19.
bioRxiv ; 2021 Sep 15.
Artigo em Inglês | MEDLINE | ID: mdl-34545363

RESUMO

Novel strategies are needed to identify drug targets and treatments for the COVID-19 pandemic. The altered gene expression of virus-infected host cells provides an opportunity to specifically inhibit viral propagation via targeting the synthetic lethal (SL) partners of such altered host genes. Pursuing this antiviral strategy, here we comprehensively analyzed multiple in vitro and in vivo bulk and single-cell RNA-sequencing datasets of SARS-CoV-2 infection to predict clinically relevant candidate antiviral targets that are SL with altered host genes. The predicted SL-based targets are highly enriched for infected cell inhibiting genes reported in four SARS-CoV-2 CRISPR-Cas9 genome-wide genetic screens. Integrating our predictions with the results of these screens, we further selected a focused subset of 26 genes that we experimentally tested in a targeted siRNA screen using human Caco-2 cells. Notably, as predicted, knocking down these targets reduced viral replication and cell viability only under the infected condition without harming non-infected cells. Our results are made publicly available, to facilitate their in vivo testing and further validation.

20.
Nat Microbiol ; 6(10): 1319-1333, 2021 10.
Artigo em Inglês | MEDLINE | ID: mdl-34556855

RESUMO

The fate of influenza A virus (IAV) infection in the host cell depends on the balance between cellular defence mechanisms and viral evasion strategies. To illuminate the landscape of IAV cellular restriction, we generated and integrated global genetic loss-of-function screens with transcriptomics and proteomics data. Our multi-omics analysis revealed a subset of both IFN-dependent and independent cellular defence mechanisms that inhibit IAV replication. Amongst these, the autophagy regulator TBC1 domain family member 5 (TBC1D5), which binds Rab7 to enable fusion of autophagosomes and lysosomes, was found to control IAV replication in vitro and in vivo and to promote lysosomal targeting of IAV M2 protein. Notably, IAV M2 was observed to abrogate TBC1D5-Rab7 binding through a physical interaction with TBC1D5 via its cytoplasmic tail. Our results provide evidence for the molecular mechanism utilised by IAV M2 protein to escape lysosomal degradation and traffic to the cell membrane, where it supports IAV budding and growth.


Assuntos
Autofagia , Evasão da Resposta Imune , Vírus da Influenza A/fisiologia , Antivirais/metabolismo , Proteínas Ativadoras de GTPase/genética , Proteínas Ativadoras de GTPase/metabolismo , Interações Hospedeiro-Patógeno , Humanos , Vírus da Influenza A/patogenicidade , Lisossomos/metabolismo , Ligação Proteica , Proteínas da Matriz Viral/metabolismo , Replicação Viral , Proteínas rab de Ligação ao GTP/metabolismo , proteínas de unión al GTP Rab7
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