蝙蝠来源的重症急性呼吸综合征样冠状病毒WIV1刺突蛋白利用浣熊狗血管紧张素转换酶Ⅱ受体侵入细胞能力的研究 : = Ability of bat severe acute respiratory syndromes-like coronavirus(WIV1) to utilize raccoon dog angiotensin-converting enzyme Ⅱ as receptor for cellular entry
目的研究蝙蝠来源的重症急性呼吸综合征(SARS)样冠状病毒WIV1刺突蛋白利用浣熊狗血管紧张素转换酶Ⅱ(ACE2)受体侵入细胞的能力,并探究SARS样冠状病毒WIV1潜在的跨宿主传播风险。方法构建来自浣熊狗、果子狸、中华菊头蝠和人等不同动物来源的ACE2表达质粒,转染至293T细胞并利用免疫印迹方法检测其在293T细胞中的表达水平;建立SARS冠状病毒(Tor2株系)及蝙蝠SARS样冠状病毒(WIV1株系)假病毒感染系统,并进行假病毒感染实验;利用假病毒感染系统及荧光素酶报告基因检测WIV1刺突蛋白对浣熊狗等不同动物来源ACE2受体利用能力;构建跨膜丝氨酸蛋白酶2(TMPRSS2)质粒,并转染至T Rex 293细胞,借助假病毒感染系统检测TMPRSS2对WIV1侵入能力的影响。结果本研究所获赠及构建的不同动物来源的ACE2质粒可经瞬时转染至细胞进行表达;与pc DNA3.1载体相比,浣熊狗、中华菊头蝠、果子狸和人的ACE2均可使蝙蝠SARS样冠状病毒WIV1侵入细胞的能力增加上万倍,上述ACE2组与载体组的荧光素酶活性差异均具有统计学意义(t=27.744、P <0.001,t=18.740、P <0.001,t=32.297、P <0.001,t=15.902、P <0.001);与TMPRSS2阴性组相比,TMPRSS2在靶细胞的表达可使WIV1假病毒的感染能力增加10倍以上,两组荧光素酶活性差异具有统计学意义(t=29.460、P <0.001)。结论蝙蝠SARS样冠状病毒WIV1的刺突蛋白除了利用人类、果子狸及中华菊头蝠的ACE2受体感染细胞,还可利用浣熊狗的ACE2受体侵入细胞,且TMPRSS2可显著促进其侵入能力,提示WIV1可能存在多种跨宿主传播的风险。.
Objective To investigate the ability of bat severe acute respiratory syndromes(SARS)-like coronavirus(WIV1 strain) to utilize angiotensin-converting enzyme Ⅱ(ACE2) from raccoon dog as a receptor for cellular entry, and to explore the potential cross-species transmissibility of WIV1. Methods Plasmids encoding ACE2 molecules from different animals including human(Homo sapiens), civet cat(Paguma larvata), Chinese horseshoe bat(Rhinolophus sinicus) and raccoon dog(Nyctereutes procyonoides) were constructed, and then transfected into 293 T cells. The expressions of ACE2 proteins in 293 T were detected by Western blot with anti-C9 antibody. The pseudoviral infection systems of SARS coronavirus(Tor2 strain) and bat SARS-like coronavirus(WIV1 strain) were established to detect receptor activity of different animals' ACE2 for WIV1 entry. The plasmid expressing transmembrane serine protease 2(TMPRSS2) was constructed, and then transfected into T Rex 293 cells. The effect of TMPRSS2 on WIV1 pp infectivity was examined by luciferase assay. Results ACE2 from raccoon dog, Chinese horseshoe bat, civet cat and human expressed well in 293 T cells and supported cellular entry mediated by WIV1 S protein. Compared with pcDNA3.1 vector, ACE2 molecules from raccoon dog, Chinese horseshoe bat, civet cat and human increased the WIV1 pp infection by thousands of times, all with significant differences in luciferase activity(t = 27.744, P < 0.001; t = 18.740, P < 0.001; t = 32.297, P < 0.001; t = 15.902, P < 0.001). Compared with the group without TMPRSS2, the expression of TMPRSS2 in target cells increased the infection mediated by WIV1-S protein by more than 10 folds, with significant difference in luciferase activity(t = 29.460, P < 0.001). Conclusions The ACE2 molecules from raccoon dog served as a functional receptor for cellular entry mediated by WIV1 spike protein, which was activated by the TMPRSS2 protease. Our findings herein suggest that there may exist a risk of multiple cross-species transmission of WIV1 among human, bat and raccoon dog..
| Medienart: |
E-Artikel |
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| Erscheinungsjahr: |
2019-10-15 2019 |
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| Erschienen: |
2019-10-15 |
| Enthalten in: |
Zur Gesamtaufnahme - year:2019 |
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| Enthalten in: |
Zhong hua shi yan he lin chuang gan ran bing za zhi - (2019), 05 vom: 15. Okt., Seite 370-376 Original Letters: Enthalten in 中华实验和临床感染病杂志 (DE-600)2991028-6 (DE-600)2991028-6 北京市 |
| Reihe: |
China Academic Journals (CAJ), E, 医药卫生科技 = Medicine & Public Health China Academic Journals (CAJ), A, 理工A(数学物理力学天地生) = Mathematics/ Physics/ Mechanics/ Astronomy |
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| Sprache: |
Chinesisch |
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| Weiterer Titel: |
Ability of bat severe acute respiratory syndromes-like coronavirus(WIV1) to utilize raccoon dog angiotensin-converting enzyme Ⅱ as receptor for cellular entry |
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| Beteiligte Personen: |
郑梅 [VerfasserIn] |
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| Links: |
oversea.cnki.net [lizenzpflichtig] |
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| Anmerkungen: |
Author info:Zheng Mei;Zheng Shuangli;Chen Danying;Jiang Dong;Zeng Hui;Zhao Xuesen;Institute of Infectious Diseases, Beijing Ditan Hospital, Capital Medical University |
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| Förderinstitution / Projekttitel: |
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| PPN (Katalog-ID): |
CAJ645207683 |
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| 245 | 1 | 0 | |a 蝙蝠来源的重症急性呼吸综合征样冠状病毒WIV1刺突蛋白利用浣熊狗血管紧张素转换酶Ⅱ受体侵入细胞能力的研究 |b = Ability of bat severe acute respiratory syndromes-like coronavirus(WIV1) to utilize raccoon dog angiotensin-converting enzyme Ⅱ as receptor for cellular entry |
| 246 | 3 | 1 | |a Ability of bat severe acute respiratory syndromes-like coronavirus(WIV1) to utilize raccoon dog angiotensin-converting enzyme Ⅱ as receptor for cellular entry |
| 264 | 1 | |c 2019-10-15 | |
| 336 | |a Text |b txt |2 rdacontent | ||
| 337 | |a Computermedien |b c |2 rdamedia | ||
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| 490 | 0 | |a China Academic Journals (CAJ) |a E |a 医药卫生科技 = Medicine & Public Health | |
| 490 | 0 | |a China Academic Journals (CAJ) |a A |a 理工A(数学物理力学天地生) = Mathematics/ Physics/ Mechanics/ Astronomy | |
| 500 | |a Author info:Zheng Mei;Zheng Shuangli;Chen Danying;Jiang Dong;Zeng Hui;Zhao Xuesen;Institute of Infectious Diseases, Beijing Ditan Hospital, Capital Medical University | ||
| 520 | |a 目的研究蝙蝠来源的重症急性呼吸综合征(SARS)样冠状病毒WIV1刺突蛋白利用浣熊狗血管紧张素转换酶Ⅱ(ACE2)受体侵入细胞的能力,并探究SARS样冠状病毒WIV1潜在的跨宿主传播风险。方法构建来自浣熊狗、果子狸、中华菊头蝠和人等不同动物来源的ACE2表达质粒,转染至293T细胞并利用免疫印迹方法检测其在293T细胞中的表达水平;建立SARS冠状病毒(Tor2株系)及蝙蝠SARS样冠状病毒(WIV1株系)假病毒感染系统,并进行假病毒感染实验;利用假病毒感染系统及荧光素酶报告基因检测WIV1刺突蛋白对浣熊狗等不同动物来源ACE2受体利用能力;构建跨膜丝氨酸蛋白酶2(TMPRSS2)质粒,并转染至T Rex 293细胞,借助假病毒感染系统检测TMPRSS2对WIV1侵入能力的影响。结果本研究所获赠及构建的不同动物来源的ACE2质粒可经瞬时转染至细胞进行表达;与pc DNA3.1载体相比,浣熊狗、中华菊头蝠、果子狸和人的ACE2均可使蝙蝠SARS样冠状病毒WIV1侵入细胞的能力增加上万倍,上述ACE2组与载体组的荧光素酶活性差异均具有统计学意义(t=27.744、P <0.001,t=18.740、P <0.001,t=32.297、P <0.001,t=15.902、P <0.001);与TMPRSS2阴性组相比,TMPRSS2在靶细胞的表达可使WIV1假病毒的感染能力增加10倍以上,两组荧光素酶活性差异具有统计学意义(t=29.460、P <0.001)。结论蝙蝠SARS样冠状病毒WIV1的刺突蛋白除了利用人类、果子狸及中华菊头蝠的ACE2受体感染细胞,还可利用浣熊狗的ACE2受体侵入细胞,且TMPRSS2可显著促进其侵入能力,提示WIV1可能存在多种跨宿主传播的风险。 | ||
| 520 | |a Objective To investigate the ability of bat severe acute respiratory syndromes(SARS)-like coronavirus(WIV1 strain) to utilize angiotensin-converting enzyme Ⅱ(ACE2) from raccoon dog as a receptor for cellular entry, and to explore the potential cross-species transmissibility of WIV1. Methods Plasmids encoding ACE2 molecules from different animals including human(Homo sapiens), civet cat(Paguma larvata), Chinese horseshoe bat(Rhinolophus sinicus) and raccoon dog(Nyctereutes procyonoides) were constructed, and then transfected into 293 T cells. The expressions of ACE2 proteins in 293 T were detected by Western blot with anti-C9 antibody. The pseudoviral infection systems of SARS coronavirus(Tor2 strain) and bat SARS-like coronavirus(WIV1 strain) were established to detect receptor activity of different animals' ACE2 for WIV1 entry. The plasmid expressing transmembrane serine protease 2(TMPRSS2) was constructed, and then transfected into T Rex 293 cells. The effect of TMPRSS2 on WIV1 pp infectivity was examined by luciferase assay. Results ACE2 from raccoon dog, Chinese horseshoe bat, civet cat and human expressed well in 293 T cells and supported cellular entry mediated by WIV1 S protein. Compared with pcDNA3.1 vector, ACE2 molecules from raccoon dog, Chinese horseshoe bat, civet cat and human increased the WIV1 pp infection by thousands of times, all with significant differences in luciferase activity(t = 27.744, P < 0.001; t = 18.740, P < 0.001; t = 32.297, P < 0.001; t = 15.902, P < 0.001). Compared with the group without TMPRSS2, the expression of TMPRSS2 in target cells increased the infection mediated by WIV1-S protein by more than 10 folds, with significant difference in luciferase activity(t = 29.460, P < 0.001). Conclusions The ACE2 molecules from raccoon dog served as a functional receptor for cellular entry mediated by WIV1 spike protein, which was activated by the TMPRSS2 protease. Our findings herein suggest that there may exist a risk of multiple cross-species transmission of WIV1 among human, bat and raccoon dog. | ||
| 610 | 2 | 4 | |a 首都医科大学附属北京地坛医院传染病研究所 |
| 650 | 4 | |a 医学微生物学（病原细菌学、病原微生物学） | |
| 650 | 4 | |a 基础医学 | |
| 650 | 4 | |a 医药、卫生 | |
| 650 | 4 | |a Biology | |
| 650 | 4 | |a Fundamental Medicine | |
| 650 | 4 | |a 医药卫生科技 | |
| 650 | 4 | |a Medicine & Public Health | |
| 650 | 4 | |a 理工A(数学物理力学天地生) | |
| 650 | 4 | |a Mathematics/ Physics/ Mechanics/ Astronomy | |
| 650 | 4 | |a 蝙蝠,重症急性呼吸综合征样冠状病毒 | |
| 650 | 4 | |a WIV1 | |
| 650 | 4 | |a 浣熊狗 | |
| 650 | 4 | |a 人类血管紧张素转换酶Ⅱ | |
| 650 | 4 | |a 跨膜丝氨酸蛋白酶2 | |
| 650 | 4 | |a Bat severe acute respiratory syndromes-like coronavirus | |
| 650 | 4 | |a Raccoon dog | |
| 650 | 4 | |a Angiotensin-converting enzyme Ⅱ | |
| 650 | 4 | |a Transmembrane protease serine 2 | |
| 700 | 0 | |a 郑双丽 |4 oth | |
| 700 | 0 | |a 陈丹瑛 |4 oth | |
| 700 | 0 | |a 蒋栋 |4 oth | |
| 700 | 0 | |a 曾辉 |4 oth | |
| 700 | 0 | |a 赵学森 |4 oth | |
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