Preparation of the RIPK3 Polyclonal Antibody and Its Application in Immunoassays of Nephropathogenic Infectious Bronchitis Virus-Infected Chickens
Receptor interacting protein kinase 3 (RIPK3) is a vital serine/threonine kinase in regulating the programmed destruction of infected cells to defend against RNA viruses. Although the role of RIPK3 in viruses in mice is well characterized, it remains unclear where in nephropathogenic infectious bron...
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| Format: | Article |
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MDPI AG
2022-08-01
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| Series: | Viruses |
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| Online Access: | https://www.mdpi.com/1999-4915/14/8/1747 |
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| author | Guanming Tian Yan Shi Xianhong Cao Wei Chen Yueming Gu Ning Li Cheng Huang Yu Zhuang Guyue Li Ping Liu Guoliang Hu Xiaona Gao Xiaoquan Guo |
| author_facet | Guanming Tian Yan Shi Xianhong Cao Wei Chen Yueming Gu Ning Li Cheng Huang Yu Zhuang Guyue Li Ping Liu Guoliang Hu Xiaona Gao Xiaoquan Guo |
| author_sort | Guanming Tian |
| collection | DOAJ |
| description | Receptor interacting protein kinase 3 (RIPK3) is a vital serine/threonine kinase in regulating the programmed destruction of infected cells to defend against RNA viruses. Although the role of RIPK3 in viruses in mice is well characterized, it remains unclear where in nephropathogenic infectious bronchitis virus (NIBV) in chickens. Here, we use a self-prepared polyclonal antibody to clarify the abundance of RIPK3 in tissues and define the contributions of RIPK3 in tissue damage caused by NIBV infection in chickens. Western blot analyses showed that RIPK3 polyclonal antibody can specifically recognize RIPK3 in the vital tissues of Hy-Line brown chicks and RIPK3 protein is abundantly expressed in the liver and kidney. Moreover, NIBV significantly upregulated the expression levels of RIPK3 in the trachea and kidney of chicks in a time-dependent manner. In addition, the activation of necroptosis in response to NIBV infection was demonstrated by the coimmunoprecipitation (CoIP) experiments through RIPK3 in the necrosome, which phosphorylates its downstream mixed-spectrum kinase structural domain-like protein (MLKL). Our findings offered preliminary insights into the key role of RIPK3 protein in studying the underlying mechanism of organ failure caused by NIBV infection. |
| first_indexed | 2024-03-09T12:21:46Z |
| format | Article |
| id | doaj.art-600d8502df1a44939c72f68be2207912 |
| institution | Directory Open Access Journal |
| issn | 1999-4915 |
| language | English |
| last_indexed | 2024-03-09T12:21:46Z |
| publishDate | 2022-08-01 |
| publisher | MDPI AG |
| record_format | Article |
| series | Viruses |
| spelling | doaj.art-600d8502df1a44939c72f68be22079122023-11-30T22:39:18ZengMDPI AGViruses1999-49152022-08-01148174710.3390/v14081747Preparation of the RIPK3 Polyclonal Antibody and Its Application in Immunoassays of Nephropathogenic Infectious Bronchitis Virus-Infected ChickensGuanming Tian0Yan Shi1Xianhong Cao2Wei Chen3Yueming Gu4Ning Li5Cheng Huang6Yu Zhuang7Guyue Li8Ping Liu9Guoliang Hu10Xiaona Gao11Xiaoquan Guo12Jiangxi Provincial Key Laboratory for Animal Health, College of Animal Science and Technology, Jiangxi Agricultural University, Nanchang 330045, ChinaSchool of Computer and Information Engineering, Jiangxi Agricultural University, Nanchang 330045, ChinaJiangxi Provincial Key Laboratory for Animal Health, College of Animal Science and Technology, Jiangxi Agricultural University, Nanchang 330045, ChinaJiangxi Provincial Key Laboratory for Animal Health, College of Animal Science and Technology, Jiangxi Agricultural University, Nanchang 330045, ChinaJiangxi Provincial Key Laboratory for Animal Health, College of Animal Science and Technology, Jiangxi Agricultural University, Nanchang 330045, ChinaJiangxi Provincial Key Laboratory for Animal Health, College of Animal Science and Technology, Jiangxi Agricultural University, Nanchang 330045, ChinaJiangxi Provincial Key Laboratory for Animal Health, College of Animal Science and Technology, Jiangxi Agricultural University, Nanchang 330045, ChinaJiangxi Provincial Key Laboratory for Animal Health, College of Animal Science and Technology, Jiangxi Agricultural University, Nanchang 330045, ChinaJiangxi Provincial Key Laboratory for Animal Health, College of Animal Science and Technology, Jiangxi Agricultural University, Nanchang 330045, ChinaJiangxi Provincial Key Laboratory for Animal Health, College of Animal Science and Technology, Jiangxi Agricultural University, Nanchang 330045, ChinaJiangxi Provincial Key Laboratory for Animal Health, College of Animal Science and Technology, Jiangxi Agricultural University, Nanchang 330045, ChinaJiangxi Provincial Key Laboratory for Animal Health, College of Animal Science and Technology, Jiangxi Agricultural University, Nanchang 330045, ChinaJiangxi Provincial Key Laboratory for Animal Health, College of Animal Science and Technology, Jiangxi Agricultural University, Nanchang 330045, ChinaReceptor interacting protein kinase 3 (RIPK3) is a vital serine/threonine kinase in regulating the programmed destruction of infected cells to defend against RNA viruses. Although the role of RIPK3 in viruses in mice is well characterized, it remains unclear where in nephropathogenic infectious bronchitis virus (NIBV) in chickens. Here, we use a self-prepared polyclonal antibody to clarify the abundance of RIPK3 in tissues and define the contributions of RIPK3 in tissue damage caused by NIBV infection in chickens. Western blot analyses showed that RIPK3 polyclonal antibody can specifically recognize RIPK3 in the vital tissues of Hy-Line brown chicks and RIPK3 protein is abundantly expressed in the liver and kidney. Moreover, NIBV significantly upregulated the expression levels of RIPK3 in the trachea and kidney of chicks in a time-dependent manner. In addition, the activation of necroptosis in response to NIBV infection was demonstrated by the coimmunoprecipitation (CoIP) experiments through RIPK3 in the necrosome, which phosphorylates its downstream mixed-spectrum kinase structural domain-like protein (MLKL). Our findings offered preliminary insights into the key role of RIPK3 protein in studying the underlying mechanism of organ failure caused by NIBV infection.https://www.mdpi.com/1999-4915/14/8/1747RIPK3NIBVpolyclonal antibodyimmunoassaysHy-Line brown chicks |
| spellingShingle | Guanming Tian Yan Shi Xianhong Cao Wei Chen Yueming Gu Ning Li Cheng Huang Yu Zhuang Guyue Li Ping Liu Guoliang Hu Xiaona Gao Xiaoquan Guo Preparation of the RIPK3 Polyclonal Antibody and Its Application in Immunoassays of Nephropathogenic Infectious Bronchitis Virus-Infected Chickens Viruses RIPK3 NIBV polyclonal antibody immunoassays Hy-Line brown chicks |
| title | Preparation of the RIPK3 Polyclonal Antibody and Its Application in Immunoassays of Nephropathogenic Infectious Bronchitis Virus-Infected Chickens |
| title_full | Preparation of the RIPK3 Polyclonal Antibody and Its Application in Immunoassays of Nephropathogenic Infectious Bronchitis Virus-Infected Chickens |
| title_fullStr | Preparation of the RIPK3 Polyclonal Antibody and Its Application in Immunoassays of Nephropathogenic Infectious Bronchitis Virus-Infected Chickens |
| title_full_unstemmed | Preparation of the RIPK3 Polyclonal Antibody and Its Application in Immunoassays of Nephropathogenic Infectious Bronchitis Virus-Infected Chickens |
| title_short | Preparation of the RIPK3 Polyclonal Antibody and Its Application in Immunoassays of Nephropathogenic Infectious Bronchitis Virus-Infected Chickens |
| title_sort | preparation of the ripk3 polyclonal antibody and its application in immunoassays of nephropathogenic infectious bronchitis virus infected chickens |
| topic | RIPK3 NIBV polyclonal antibody immunoassays Hy-Line brown chicks |
| url | https://www.mdpi.com/1999-4915/14/8/1747 |
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