PB2 and PA mutations contribute to the pathogenicity of mouse-adapted pdmH1N1-Venus reporter influenza A virus in a mammalian model
Influenza A viruses have been a threat to human health for the past 100 years. Understanding the dynamics and pathogenicity of the influenza viruses in vivo is of great value in controlling the influenza pandemic. Fluorescent protein-carrying recombinant influenza virus is a substantially useful too...
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Frontiers Media S.A.
2025-01-01
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| Series: | Frontiers in Microbiology |
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| Online Access: | https://www.frontiersin.org/articles/10.3389/fmicb.2024.1532304/full |
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| author | Shixiang Wu Ruonan Yi Yingying Tao Huimin Wu Li Wu Jiasheng Song Xin Zhang Beibei Yang Xing Wu Yulong He Jianhong Shu Huapeng Feng |
| author_facet | Shixiang Wu Ruonan Yi Yingying Tao Huimin Wu Li Wu Jiasheng Song Xin Zhang Beibei Yang Xing Wu Yulong He Jianhong Shu Huapeng Feng |
| author_sort | Shixiang Wu |
| collection | DOAJ |
| description | Influenza A viruses have been a threat to human health for the past 100 years. Understanding the dynamics and pathogenicity of the influenza viruses in vivo is of great value in controlling the influenza pandemic. Fluorescent protein-carrying recombinant influenza virus is a substantially useful tool for studying viral characteristics in vivo and high-throughput screening in vitro. In this study, we generated a recombinant pdmH1N1 CA04 influenza virus carrying a Venus reporter gene in the non-structural (NS) segment using reverse genetics. After passaging the recombinant influenza virus carrying Venus from lung to lung in mice, we found that virulence of the passaged pdmH1N1 CA04-Venus significantly increased and was lethal to the mice. We finally isolated one mouse-adapted pdmH1N1 CA04-Venus with bigger plaques expressing the amount of Venus proteins by using the ninth passage lung homogenate with plague purification. We found three different amino acids (PB2 T340K, PA I21M, and F175L) between WT-CA04-Venus and MA-CA04-Venus using whole-genome sequencing. Interestingly, the polymerase activity of MA-CA04-Venus was significantly lower than that of WT-CA04-Venus in a minigenome assay. Further investigation demonstrates that PA I21M and PA I21M + PB2 T340K significantly enhanced the polymerase activity of WT-CA04-Venus; however, PA F175L + PB2 T340K significantly decreased the polymerase activity of MA-CA04-Venus. Therefore, PA I21M mutation may determine the increased virulence in mice, and PA F175L + PB2 T340K may be involved in the stability of Venus insertion. Above all, we generated a mouse-adapted pdmH1N1 CA04-Venus virus with high virulence and stable green fluorescent Venus protein. It is a useful tool for high-throughput screening of antiviral drugs and for investigating the interaction between the influenza virus and host in vivo. |
| format | Article |
| id | doaj-art-055b1b3f0a9d40fb8be78cdf13475c4a |
| institution | Kabale University |
| issn | 1664-302X |
| language | English |
| publishDate | 2025-01-01 |
| publisher | Frontiers Media S.A. |
| record_format | Article |
| series | Frontiers in Microbiology |
| spelling | doaj-art-055b1b3f0a9d40fb8be78cdf13475c4a2025-01-07T06:44:34ZengFrontiers Media S.A.Frontiers in Microbiology1664-302X2025-01-011510.3389/fmicb.2024.15323041532304PB2 and PA mutations contribute to the pathogenicity of mouse-adapted pdmH1N1-Venus reporter influenza A virus in a mammalian modelShixiang Wu0Ruonan Yi1Yingying Tao2Huimin Wu3Li Wu4Jiasheng Song5Xin Zhang6Beibei Yang7Xing Wu8Yulong He9Jianhong Shu10Huapeng Feng11Department of Biopharmacy, College of Life Sciences and Medicine, Zhejiang Sci-Tech University, Hangzhou, ChinaDepartment of Biopharmacy, College of Life Sciences and Medicine, Zhejiang Sci-Tech University, Hangzhou, ChinaDepartment of Biopharmacy, College of Life Sciences and Medicine, Zhejiang Sci-Tech University, Hangzhou, ChinaDepartment of Biopharmacy, College of Life Sciences and Medicine, Zhejiang Sci-Tech University, Hangzhou, ChinaDepartment of Biology, College of Life Sciences, China Jiliang University, Hangzhou, ChinaZhejiang Difference Biotechnology Co., Ltd, Hangzhou, ChinaDepartment of Biopharmacy, College of Life Sciences and Medicine, Zhejiang Sci-Tech University, Hangzhou, ChinaDepartment of Biopharmacy, College of Life Sciences and Medicine, Zhejiang Sci-Tech University, Hangzhou, ChinaDepartment of Biopharmacy, College of Life Sciences and Medicine, Zhejiang Sci-Tech University, Hangzhou, ChinaDepartment of Biopharmacy, College of Life Sciences and Medicine, Zhejiang Sci-Tech University, Hangzhou, ChinaDepartment of Biopharmacy, College of Life Sciences and Medicine, Zhejiang Sci-Tech University, Hangzhou, ChinaDepartment of Biopharmacy, College of Life Sciences and Medicine, Zhejiang Sci-Tech University, Hangzhou, ChinaInfluenza A viruses have been a threat to human health for the past 100 years. Understanding the dynamics and pathogenicity of the influenza viruses in vivo is of great value in controlling the influenza pandemic. Fluorescent protein-carrying recombinant influenza virus is a substantially useful tool for studying viral characteristics in vivo and high-throughput screening in vitro. In this study, we generated a recombinant pdmH1N1 CA04 influenza virus carrying a Venus reporter gene in the non-structural (NS) segment using reverse genetics. After passaging the recombinant influenza virus carrying Venus from lung to lung in mice, we found that virulence of the passaged pdmH1N1 CA04-Venus significantly increased and was lethal to the mice. We finally isolated one mouse-adapted pdmH1N1 CA04-Venus with bigger plaques expressing the amount of Venus proteins by using the ninth passage lung homogenate with plague purification. We found three different amino acids (PB2 T340K, PA I21M, and F175L) between WT-CA04-Venus and MA-CA04-Venus using whole-genome sequencing. Interestingly, the polymerase activity of MA-CA04-Venus was significantly lower than that of WT-CA04-Venus in a minigenome assay. Further investigation demonstrates that PA I21M and PA I21M + PB2 T340K significantly enhanced the polymerase activity of WT-CA04-Venus; however, PA F175L + PB2 T340K significantly decreased the polymerase activity of MA-CA04-Venus. Therefore, PA I21M mutation may determine the increased virulence in mice, and PA F175L + PB2 T340K may be involved in the stability of Venus insertion. Above all, we generated a mouse-adapted pdmH1N1 CA04-Venus virus with high virulence and stable green fluorescent Venus protein. It is a useful tool for high-throughput screening of antiviral drugs and for investigating the interaction between the influenza virus and host in vivo.https://www.frontiersin.org/articles/10.3389/fmicb.2024.1532304/fullreporter influenza virusmouse-adaptedpdmH1N1 CA04VenusstabilityPA and PB2 |
| spellingShingle | Shixiang Wu Ruonan Yi Yingying Tao Huimin Wu Li Wu Jiasheng Song Xin Zhang Beibei Yang Xing Wu Yulong He Jianhong Shu Huapeng Feng PB2 and PA mutations contribute to the pathogenicity of mouse-adapted pdmH1N1-Venus reporter influenza A virus in a mammalian model Frontiers in Microbiology reporter influenza virus mouse-adapted pdmH1N1 CA04 Venus stability PA and PB2 |
| title | PB2 and PA mutations contribute to the pathogenicity of mouse-adapted pdmH1N1-Venus reporter influenza A virus in a mammalian model |
| title_full | PB2 and PA mutations contribute to the pathogenicity of mouse-adapted pdmH1N1-Venus reporter influenza A virus in a mammalian model |
| title_fullStr | PB2 and PA mutations contribute to the pathogenicity of mouse-adapted pdmH1N1-Venus reporter influenza A virus in a mammalian model |
| title_full_unstemmed | PB2 and PA mutations contribute to the pathogenicity of mouse-adapted pdmH1N1-Venus reporter influenza A virus in a mammalian model |
| title_short | PB2 and PA mutations contribute to the pathogenicity of mouse-adapted pdmH1N1-Venus reporter influenza A virus in a mammalian model |
| title_sort | pb2 and pa mutations contribute to the pathogenicity of mouse adapted pdmh1n1 venus reporter influenza a virus in a mammalian model |
| topic | reporter influenza virus mouse-adapted pdmH1N1 CA04 Venus stability PA and PB2 |
| url | https://www.frontiersin.org/articles/10.3389/fmicb.2024.1532304/full |
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