CRISPR/Cas9 screens identify key host factors that enhance rotavirus reverse genetics efficacy and vaccine production
Abstract Rotaviruses pose a significant threat to young children. To identify novel pro- and anti-rotavirus host factors, we performed genome-wide CRISPR/Cas9 screens using rhesus rotavirus and African green monkey cells. Genetic deletion of either SERPINB1 or TMEM236, the top two antiviral factors,...
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| Main Authors: | , , , , , , , , , , |
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| Format: | Article |
| Language: | English |
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Nature Portfolio
2024-11-01
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| Series: | npj Vaccines |
| Online Access: | https://doi.org/10.1038/s41541-024-01007-7 |
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| _version_ | 1850061832502378496 |
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| author | Yinxing Zhu Meagan E. Sullender Danielle E. Campbell Leran Wang Sanghyun Lee Takahiro Kawagishi Gaopeng Hou Alen Dizdarevic Philippe H. Jais Megan T. Baldridge Siyuan Ding |
| author_facet | Yinxing Zhu Meagan E. Sullender Danielle E. Campbell Leran Wang Sanghyun Lee Takahiro Kawagishi Gaopeng Hou Alen Dizdarevic Philippe H. Jais Megan T. Baldridge Siyuan Ding |
| author_sort | Yinxing Zhu |
| collection | DOAJ |
| description | Abstract Rotaviruses pose a significant threat to young children. To identify novel pro- and anti-rotavirus host factors, we performed genome-wide CRISPR/Cas9 screens using rhesus rotavirus and African green monkey cells. Genetic deletion of either SERPINB1 or TMEM236, the top two antiviral factors, in MA104 cells increased virus titers in a rotavirus strain independent manner. Using this information, we optimized the existing rotavirus reverse genetics systems by combining SERPINB1 knockout MA104 cells with a C3P3-G3 helper plasmid. We improved the recovery efficiency and rescued several low-titer rotavirus reporter and mutant strains that prove difficult to rescue otherwise. Furthermore, we demonstrate that TMEM236 knockout in Vero cells supported higher yields of two live-attenuated rotavirus vaccine strains than the parental cell line and represents a more robust vaccine-producing cell substrate. Collectively, we developed a third-generation optimized rotavirus reverse genetics system and generated gene-edited Vero cells as a new substrate for improving rotavirus vaccine production. |
| format | Article |
| id | doaj-art-c27da01121eb448cb209bdcb794bd3ad |
| institution | DOAJ |
| issn | 2059-0105 |
| language | English |
| publishDate | 2024-11-01 |
| publisher | Nature Portfolio |
| record_format | Article |
| series | npj Vaccines |
| spelling | doaj-art-c27da01121eb448cb209bdcb794bd3ad2025-08-20T02:50:05ZengNature Portfolionpj Vaccines2059-01052024-11-019111210.1038/s41541-024-01007-7CRISPR/Cas9 screens identify key host factors that enhance rotavirus reverse genetics efficacy and vaccine productionYinxing Zhu0Meagan E. Sullender1Danielle E. Campbell2Leran Wang3Sanghyun Lee4Takahiro Kawagishi5Gaopeng Hou6Alen Dizdarevic7Philippe H. Jais8Megan T. Baldridge9Siyuan Ding10Department of Molecular Microbiology, Washington University School of MedicineDepartment of Medicine, Division of Infectious Diseases and Edison Family Center for Genome Sciences & Systems Biology, Washington University School of MedicineDepartment of Medicine, Division of Infectious Diseases and Edison Family Center for Genome Sciences & Systems Biology, Washington University School of MedicineDepartment of Medicine, Division of Infectious Diseases and Edison Family Center for Genome Sciences & Systems Biology, Washington University School of MedicineDivision of Biology and Medicine, Department of Molecular Microbiology and Immunology, Brown UniversityDepartment of Molecular Microbiology, Washington University School of MedicineDepartment of Molecular Microbiology, Washington University School of MedicineDepartment of Molecular Microbiology, Washington University School of MedicineEukarÿs SAS, Pépinière Genopole, 4 rue Pierre Fontaine, Genopole Entreprises Campus 3, 4 Rue Pierre Fontaine 91000Department of Molecular Microbiology, Washington University School of MedicineDepartment of Molecular Microbiology, Washington University School of MedicineAbstract Rotaviruses pose a significant threat to young children. To identify novel pro- and anti-rotavirus host factors, we performed genome-wide CRISPR/Cas9 screens using rhesus rotavirus and African green monkey cells. Genetic deletion of either SERPINB1 or TMEM236, the top two antiviral factors, in MA104 cells increased virus titers in a rotavirus strain independent manner. Using this information, we optimized the existing rotavirus reverse genetics systems by combining SERPINB1 knockout MA104 cells with a C3P3-G3 helper plasmid. We improved the recovery efficiency and rescued several low-titer rotavirus reporter and mutant strains that prove difficult to rescue otherwise. Furthermore, we demonstrate that TMEM236 knockout in Vero cells supported higher yields of two live-attenuated rotavirus vaccine strains than the parental cell line and represents a more robust vaccine-producing cell substrate. Collectively, we developed a third-generation optimized rotavirus reverse genetics system and generated gene-edited Vero cells as a new substrate for improving rotavirus vaccine production.https://doi.org/10.1038/s41541-024-01007-7 |
| spellingShingle | Yinxing Zhu Meagan E. Sullender Danielle E. Campbell Leran Wang Sanghyun Lee Takahiro Kawagishi Gaopeng Hou Alen Dizdarevic Philippe H. Jais Megan T. Baldridge Siyuan Ding CRISPR/Cas9 screens identify key host factors that enhance rotavirus reverse genetics efficacy and vaccine production npj Vaccines |
| title | CRISPR/Cas9 screens identify key host factors that enhance rotavirus reverse genetics efficacy and vaccine production |
| title_full | CRISPR/Cas9 screens identify key host factors that enhance rotavirus reverse genetics efficacy and vaccine production |
| title_fullStr | CRISPR/Cas9 screens identify key host factors that enhance rotavirus reverse genetics efficacy and vaccine production |
| title_full_unstemmed | CRISPR/Cas9 screens identify key host factors that enhance rotavirus reverse genetics efficacy and vaccine production |
| title_short | CRISPR/Cas9 screens identify key host factors that enhance rotavirus reverse genetics efficacy and vaccine production |
| title_sort | crispr cas9 screens identify key host factors that enhance rotavirus reverse genetics efficacy and vaccine production |
| url | https://doi.org/10.1038/s41541-024-01007-7 |
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