High-throughput screening of genetic and cellular drivers of syncytium formation induced by the spike protein of SARS-CoV-2
© 2023. The Author(s)..
Mapping mutations and discovering cellular determinants that cause the spike protein of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) to induce infected cells to form syncytia would facilitate the development of strategies for blocking the formation of such cell-cell fusion. Here we describe high-throughput screening methods based on droplet microfluidics and the size-exclusion selection of syncytia, coupled with large-scale mutagenesis and genome-wide knockout screening via clustered regularly interspaced short palindromic repeats (CRISPR), for the large-scale identification of determinants of cell-cell fusion. We used the methods to perform deep mutational scans in spike-presenting cells to pinpoint mutable syncytium-enhancing substitutions in two regions of the spike protein (the fusion peptide proximal region and the furin-cleavage site). We also used a genome-wide CRISPR screen in cells expressing the receptor angiotensin-converting enzyme 2 to identify inhibitors of clathrin-mediated endocytosis that impede syncytium formation, which we validated in hamsters infected with SARS-CoV-2. Finding genetic and cellular determinants of the formation of syncytia may reveal insights into the physiological and pathological consequences of cell-cell fusion.
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| Medienart: |
E-Artikel |
| Erscheinungsjahr: |
2024 |
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| Erschienen: |
2024 |
| Enthalten in: |
Zur Gesamtaufnahme - volume:8 |
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| Enthalten in: |
Nature biomedical engineering - 8(2024), 3 vom: 23. März, Seite 291-309 |
| Sprache: |
Englisch |
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| Beteiligte Personen: |
Chan, Charles W F [VerfasserIn] |
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Journal Article |
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Date Completed 27.03.2024 Date Revised 28.03.2024 published: Print-Electronic ErratumIn: Nat Biomed Eng. 2024 Jan 24;:. - PMID 38267642 Citation Status MEDLINE |
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| doi: |
10.1038/s41551-023-01140-z |
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| 520 | |a © 2023. The Author(s). | ||
| 520 | |a Mapping mutations and discovering cellular determinants that cause the spike protein of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) to induce infected cells to form syncytia would facilitate the development of strategies for blocking the formation of such cell-cell fusion. Here we describe high-throughput screening methods based on droplet microfluidics and the size-exclusion selection of syncytia, coupled with large-scale mutagenesis and genome-wide knockout screening via clustered regularly interspaced short palindromic repeats (CRISPR), for the large-scale identification of determinants of cell-cell fusion. We used the methods to perform deep mutational scans in spike-presenting cells to pinpoint mutable syncytium-enhancing substitutions in two regions of the spike protein (the fusion peptide proximal region and the furin-cleavage site). We also used a genome-wide CRISPR screen in cells expressing the receptor angiotensin-converting enzyme 2 to identify inhibitors of clathrin-mediated endocytosis that impede syncytium formation, which we validated in hamsters infected with SARS-CoV-2. Finding genetic and cellular determinants of the formation of syncytia may reveal insights into the physiological and pathological consequences of cell-cell fusion | ||
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| 700 | 1 | |a Huang, Xiner |e verfasserin |4 aut | |
| 700 | 1 | |a Mao, Tianjiao |e verfasserin |4 aut | |
| 700 | 1 | |a Chu, Hoi Yee |e verfasserin |4 aut | |
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| 700 | 1 | |a Chu, Hin |e verfasserin |4 aut | |
| 700 | 1 | |a Choi, Gigi C G |e verfasserin |4 aut | |
| 700 | 1 | |a Shum, Ho Cheung |e verfasserin |4 aut | |
| 700 | 1 | |a Wong, Alan S L |e verfasserin |4 aut | |
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