Characterization of a Human Sapovirus Genotype GII.3 Strain Generated by a Reverse Genetics System: VP2 Is a Minor Structural Protein of the Virion
We devised a reverse genetics system to generate an infectious human sapovirus (HuSaV) GII.3 virus. Capped/uncapped full-length RNAs derived from HuSaV GII.3 AK11 strain generated by in vitro transcription were used to transfect HuTu80 human duodenum carcinoma cells; infectious viruses were recovere...
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MDPI AG
2022-07-01
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| Series: | Viruses |
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| Online Access: | https://www.mdpi.com/1999-4915/14/8/1649 |
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| author | Tian-Cheng Li Michiyo Kataoka Yen Hai Doan Hiroyuki Saito Hirotaka Takagi Masamichi Muramatsu Tomoichiro Oka |
| author_facet | Tian-Cheng Li Michiyo Kataoka Yen Hai Doan Hiroyuki Saito Hirotaka Takagi Masamichi Muramatsu Tomoichiro Oka |
| author_sort | Tian-Cheng Li |
| collection | DOAJ |
| description | We devised a reverse genetics system to generate an infectious human sapovirus (HuSaV) GII.3 virus. Capped/uncapped full-length RNAs derived from HuSaV GII.3 AK11 strain generated by in vitro transcription were used to transfect HuTu80 human duodenum carcinoma cells; infectious viruses were recovered from the capped RNA-transfected cells and passaged in the cells. Genome-wide analyses indicated no nucleotide sequence change in the virus genomes in the cell-culture supernatants recovered from the transfection or those from the subsequent infection. No virus growth was detected in the uncapped RNA-transfected cells, suggesting that the 5′-cap structure is essential for the virus’ generation and replication. Two types of virus particles were purified from the cell-culture supernatant. The complete particles were 39.2-nm-dia., at 1.350 g/cm<sup>3</sup> density; the empty particles were 42.2-nm-dia. at 1.286 g/cm<sup>3</sup>. Two proteins (58-kDa p58 and 17-kDa p17) were detected from the purified particles; their molecular weight were similar to those of VP1 (~60-kDa) and VP2 (~16-kDa) of AK11 strain deduced from their amino acids (aa) sequences. Protein p58 interacted with HuSaV GII.3-VP1-specific antiserum, suggesting that p58 is HuSaV VP1. A total of 94 (57%) aa of p17 were identified by mass spectrometry; the sequences were identical to those of VP2, indicating that the p17 is the VP2 of AK11. Our new method produced infectious HuSaVs and demonstrated that VP2 is the minor protein of the virion, suggested to be involved in the HuSaV assembly. |
| first_indexed | 2024-03-09T03:42:07Z |
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| institution | Directory Open Access Journal |
| issn | 1999-4915 |
| language | English |
| last_indexed | 2024-03-09T03:42:07Z |
| publishDate | 2022-07-01 |
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| series | Viruses |
| spelling | doaj.art-0f883a1e0c564e3eab824377e5f2258f2023-12-03T14:38:49ZengMDPI AGViruses1999-49152022-07-01148164910.3390/v14081649Characterization of a Human Sapovirus Genotype GII.3 Strain Generated by a Reverse Genetics System: VP2 Is a Minor Structural Protein of the VirionTian-Cheng Li0Michiyo Kataoka1Yen Hai Doan2Hiroyuki Saito3Hirotaka Takagi4Masamichi Muramatsu5Tomoichiro Oka6Department of Virology II, National Institute of Infectious Diseases, Tokyo 208-0011, JapanDepartment of Pathology, National Institute of Infectious Diseases, Tokyo 208-0011, JapanCenter for Emergency Preparedness and Response, National Institute of Infectious Diseases, Tokyo 208-0011, JapanAkita Prefectural Research Center for Public Health and Environment, Akita 010-0874, JapanManagement Department of Biosafety, Laboratory Animal and Pathogen Bank, National Institute of Infectious Diseases, Tokyo 208-0011, JapanDepartment of Virology II, National Institute of Infectious Diseases, Tokyo 208-0011, JapanDepartment of Virology II, National Institute of Infectious Diseases, Tokyo 208-0011, JapanWe devised a reverse genetics system to generate an infectious human sapovirus (HuSaV) GII.3 virus. Capped/uncapped full-length RNAs derived from HuSaV GII.3 AK11 strain generated by in vitro transcription were used to transfect HuTu80 human duodenum carcinoma cells; infectious viruses were recovered from the capped RNA-transfected cells and passaged in the cells. Genome-wide analyses indicated no nucleotide sequence change in the virus genomes in the cell-culture supernatants recovered from the transfection or those from the subsequent infection. No virus growth was detected in the uncapped RNA-transfected cells, suggesting that the 5′-cap structure is essential for the virus’ generation and replication. Two types of virus particles were purified from the cell-culture supernatant. The complete particles were 39.2-nm-dia., at 1.350 g/cm<sup>3</sup> density; the empty particles were 42.2-nm-dia. at 1.286 g/cm<sup>3</sup>. Two proteins (58-kDa p58 and 17-kDa p17) were detected from the purified particles; their molecular weight were similar to those of VP1 (~60-kDa) and VP2 (~16-kDa) of AK11 strain deduced from their amino acids (aa) sequences. Protein p58 interacted with HuSaV GII.3-VP1-specific antiserum, suggesting that p58 is HuSaV VP1. A total of 94 (57%) aa of p17 were identified by mass spectrometry; the sequences were identical to those of VP2, indicating that the p17 is the VP2 of AK11. Our new method produced infectious HuSaVs and demonstrated that VP2 is the minor protein of the virion, suggested to be involved in the HuSaV assembly.https://www.mdpi.com/1999-4915/14/8/1649human sapovirusHuSaVgenogroup GII.3virus particlereverse genetics systemVP1 |
| spellingShingle | Tian-Cheng Li Michiyo Kataoka Yen Hai Doan Hiroyuki Saito Hirotaka Takagi Masamichi Muramatsu Tomoichiro Oka Characterization of a Human Sapovirus Genotype GII.3 Strain Generated by a Reverse Genetics System: VP2 Is a Minor Structural Protein of the Virion Viruses human sapovirus HuSaV genogroup GII.3 virus particle reverse genetics system VP1 |
| title | Characterization of a Human Sapovirus Genotype GII.3 Strain Generated by a Reverse Genetics System: VP2 Is a Minor Structural Protein of the Virion |
| title_full | Characterization of a Human Sapovirus Genotype GII.3 Strain Generated by a Reverse Genetics System: VP2 Is a Minor Structural Protein of the Virion |
| title_fullStr | Characterization of a Human Sapovirus Genotype GII.3 Strain Generated by a Reverse Genetics System: VP2 Is a Minor Structural Protein of the Virion |
| title_full_unstemmed | Characterization of a Human Sapovirus Genotype GII.3 Strain Generated by a Reverse Genetics System: VP2 Is a Minor Structural Protein of the Virion |
| title_short | Characterization of a Human Sapovirus Genotype GII.3 Strain Generated by a Reverse Genetics System: VP2 Is a Minor Structural Protein of the Virion |
| title_sort | characterization of a human sapovirus genotype gii 3 strain generated by a reverse genetics system vp2 is a minor structural protein of the virion |
| topic | human sapovirus HuSaV genogroup GII.3 virus particle reverse genetics system VP1 |
| url | https://www.mdpi.com/1999-4915/14/8/1649 |
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