CRISPR screens for host factors critical for infection by SARS-CoV-2 variants of concern
Ontology highlight
ABSTRACT:
PROVIDER: PRJNA812945 | ENA |
REPOSITORIES: ENA
Dataset's files
| Action | DRS | |||
|---|---|---|---|---|
| SRR18231448.fastq.gz | Fastqsanger.gz | |||
| SRR18231449.fastq.gz | Fastqsanger.gz | |||
| SRR18231450.fastq.gz | Fastqsanger.gz | |||
| SRR18231451.fastq.gz | Fastqsanger.gz | |||
| SRR18231452.fastq.gz | Fastqsanger.gz |
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Project description:The global spread of SARS-CoV-2 lead to the most challenging pandemic in this century, posing major economic and health challenges worldwide. Revealing host genes essential for infection by multiple variants of SARS-CoV-2 can provide insights into the virus pathogenesis, and facilitates the development of novel broad-spectrum host-directed therapeutics. Here, employing genome-scale CRISPR screens, we provide a comprehensive data-set of cellular factors that are exploited by WT-SARS-CoV-2 as well as two additional recently emerged variants of concerns (VOCs), Alpha and Beta. These screens identified known and novel host factors critical for SARS-CoV-2 infection, including various components belonging to the Clathrin-dependent transport pathway, ubiquitination and Heparan sulfate biogenesis. In addition, the host phosphatidylglycerol biosynthesis processes appeared to have major anti-viral functions. Comparative analysis of the different VOCs revealed the host factors KREMEN2 and SETDB1 as potential unique candidates required only to the Alpha variant, providing a possible explanation for the increased infectivity of this variant. Furthermore, the analysis identified GATA6, a zinc finger transcription factor, as an essential pro-viral gene for all variants inspected. We revealed that GATA6 directly regulates ACE2 transcription and accordingly, is critical for SARS-CoV-2 cell entry. Analysis of clinical samples collected from SARS-CoV-2 infected individuals showed an elevated level of GATA6, indicating the important role GATA6 may be playing in COVID-19 pathogenesis. Finally, pharmacological inhibition of GATA6 resulted in down-modulation of ACE2 and consequently to inhibition of the viral infectivity. Overall, we show GATA6 represents a target for the development of anti-SARS-CoV-2 therapeutic strategies and reaffirm the value of the CRISPR loss-of-function screens in providing a list of potential new targets for therapeutic interventions.
2022-03-27 | GSE197962 | GEO
Project description:A key feature of RNA viruses, including SARS-CoV-2, is their high mutation rate, which allows them to develop resistance to vaccines and antiviral drugs targeting viral proteins. To overcome this downside, a strategy would be to target host factors, i.e. cell proteins required by the virus for its replication. However, it is still unclear whether cell responses induced by different SARS-CoV-2 variants are conserved and if the same core of host factors is exploited by different variants. We compared 3 variants of concern (VOC) that emerged during the first year of the pandemic and observed that the host transcriptional response was mostly conserved, differing only in the kinetics and magnitude. By CRISPR screening we identified the host genes required for infection by each VOC. These genes were associated with interferon and JAK/STAT pathway, autophagy, mTOR pathway, mitochondrial organisation and activity. Crucially, we failed to identify genes required by only one variant. We further validated our candidates with small molecules and repurposed FDA-approved drugs, which were effective against a novel variant emerged during the course of the study. We observed that SARS-CoV-2 infection induces a rapid spike in Reactive Oxygen Species (ROS) production, which can be targeted to block viral propagation. Our study identifies a core of host genes required for infection, and lays the foundation for general therapeutic strategies aimed at minimising emergence of novel resistant variants.
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Project description:Several genome-wide CRISPR knockout screens have been conducted to identify host factors regulating SARS-CoV-2 replication, but the models used have often relied on overexpression of ACE2 receptor and didn’t express TMPRSS2 protease, known to be important for viral entry at the plasma membrane. Here, we conducted a meta-analysis of these screens and showed a high level of cell-type specificity of the identified hits, arguing for the necessity to pursue efforts to uncover the full landscape of SARS-CoV-2 regulators. We performed genome-wide, bidirectional CRISPR screens in Calu-3 lung epithelial cells, as well as knockout screens in Caco-2 intestinal cells. As well as identifying ACE2 and TMPRSS2 as top hits, our study reveals a series of so far unidentified and critical host-dependency factors, including the Adaptins AP1G1 and AP1B1 and the flippase ATP8B1. Additionally, new anti-SARS-CoV-2 proteins with potent activity, including several membrane-associated Mucins (MUC1, MUC4 and MUC21), IL6R and CD44 were identified . We further observed that these genes mostly acted at the critical step of viral entry, with the notable exception of ATP8B1, the knockout of which prevented late stages of viral replication. Exploring the pro- and anti-viral breadth of these genes using highly pathogenic MERS-CoV, seasonal HCoV-NL63 and -229E and influenza A virus, we reveal that some genes such as AP1G1 and ATP8B1 are general coronavirus cofactors. In contrast, Mucins recapitulated their known role as a general antiviral defense mechanism. These results demonstrate the value of considering multiple cell models and perturbational modalities for understanding SARS-CoV-2 replication and provide a list of potential new targets for therapeutic interventions.
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Project description:Several genome-wide CRISPR knockout screens have been conducted to identify host factors regulating SARS-CoV-2 replication, but the models used have often relied on overexpression of ACE2 receptor and didn’t express TMPRSS2 protease, known to be important for viral entry at the plasma membrane. Here, we conducted a meta-analysis of these screens and showed a high level of cell-type specificity of the identified hits, arguing for the necessity to pursue efforts to uncover the full landscape of SARS-CoV-2 regulators. We performed genome-wide, bidirectional CRISPR screens in Calu-3 lung epithelial cells, as well as knockout screens in Caco-2 intestinal cells. As well as identifying ACE2 and TMPRSS2 as top hits, our study reveals a series of so far unidentified and critical host-dependency factors, including the Adaptins AP1G1 and AP1B1 and the flippase ATP8B1. Additionally, new anti-SARS-CoV-2 proteins with potent activity, including several membrane-associated Mucins (MUC1, MUC4 and MUC21), IL6R and CD44 were identified . We further observed that these genes mostly acted at the critical step of viral entry, with the notable exception of ATP8B1, the knockout of which prevented late stages of viral replication. Exploring the pro- and anti-viral breadth of these genes using highly pathogenic MERS-CoV, seasonal HCoV-NL63 and -229E and influenza A virus, we reveal that some genes such as AP1G1 and ATP8B1 are general coronavirus cofactors. In contrast, Mucins recapitulated their known role as a general antiviral defense mechanism. These results demonstrate the value of considering multiple cell models and perturbational modalities for understanding SARS-CoV-2 replication and provide a list of potential new targets for therapeutic interventions.
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Project description:We performed genome-wide CRISPR KO screens in human Huh7.5.1 cells to select for mutations that render host cells resistant to viral infection by SARS-CoV-2, human coronavirus 229E and OC43.
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Project description:The ongoing COVID-19 pandemic represents an unprecedented global health crisis. Here, we report the identification of a synthetic nanobody (sybody) pair, Sb#15 and Sb#68, that can bind simultaneously to the SARS-CoV-2 spike-RBD and efficiently neutralize pseudotyped and live-viruses by interfering with ACE2 interaction. Cryo-EM confirms that Sb#15 and Sb#68 engage two spatially-discrete epitopes, influencing rational design of bispecific and tri-bispecific fusion constructs that exhibit up to 100- and 1000-fold increase in neutralization potency, respectively. Cryo-EM of the sybody-spike complex additionally reveals a novel up-out RBD conformation. While resistant viruses emerge rapidly in the presence of single binders, no escape variants are observed in presence of the bispecific sybody. The multivalent bispecific constructs further increase the neutralization potency against globally-circulating SARS-CoV-2 variants of concern. Our study illustrates the power of multivalency and biparatopic nanobody fusions for the potential development of therapeutic strategies that mitigate the emergence of new SARS-CoV-2 escape mutants.
| S-SCDT-EMBOR-2021-54199V1 | biostudies-other
Project description:Identification of host genes essential for severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection may reveal novel therapeutic targets and inform our understanding of coronavirus disease 2019 (COVID-19) pathogenesis. Here we performed genome-wide CRISPR screens in Vero-E6 cells with SARS-CoV-2, Middle East respiratory syndrome CoV (MERS-CoV), bat CoV HKU5 expressing the SARS-CoV-1 spike, and vesicular stomatitis virus (VSV) expressing the SARS-CoV-2 spike. We identified known SARS-CoV-2 host factors, including the receptor ACE2 and protease Cathepsin L. We additionally discovered pro-viral genes and pathways, including HMGB1 and the SWI/SNF chromatin remodeling complex, that are SARS lineage and pan-coronavirus specific, respectively. We show that HMGB1 regulates ACE2 expression and is critical for entry of SARS-CoV-2, SARS-CoV-1, and NL63. We also show that small-molecule antagonists of identified gene products inhibited SARS-CoV-2 infection in monkey and human cells, demonstrating the conserved role of these genetic hits across species. This identifies potential therapeutic targets for SARS-CoV-2 and reveals SARS lineage-specific and pan-CoV host factors that regulate susceptibility to highly pathogenic CoVs.
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Project description:During the COVID-19 pandemic, three variants of concern (VOC) of the severe acute respiratory syndrome-coronavirus-2 (SARS-CoV-2) emerged globally with increasingly greater ability to evade the immune response and enhanced transmissibility. A critical question we have faced with each new VOC is how well the vaccine based on the ancestral strain will protect. Given vaccine hesitancy, it is also important to address how prior live infections may protect against new variants. To address this later question, K18-hACE2 mice were infected with a low, partially lethal dose of WA1/2020 (WA1), the prototype strain on which the mRNA vaccines are based. On 24 days post-infection (dpi), surviving mice were challenged with WA1, Alpha or Delta. Nasal turbinates and lungs showed active replication on 1- and 3-days post-challenge (dpc) in Alpha- and Delta-, but not WA1- challenge groups. The rate of recovery of Alpha- and Delta-challenged mice lagged behind WA1 with mice showing weight loss despite a greatly dampened proinflammatory response in the lung. In contrast, nasal turbinates had a heightened proinflammatory response for Alpha- and Delta- but not WA1-challenged groups. Inflammatory responses were limited to the upper respiratory tract. Despite this, we did not observe significant difference among the groups in fold change in the neutralizing titer. In summary, infection with a low dose of SARS-CoV-2 confers protection from challenge, but mice in the heterologous challenge group differed in having greater weight loss, viral replication, and a heightened proinflammatory response in the upper respiratory tract.
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