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Volume 39 Issue 3
June 2024

    Yuming Li, Yanhong Tang, Xiaoqian Wang, Airu Zhu, Dongdong Liu, Yiyun He, Hu Guo, Jie Zheng, Xinzhuo Liu, Fengyu Chi, Yanqun Wang, Zhen Zhuang, Zhaoyong Zhang, Donglan Liu, Zhao Chen, Fang Li, Wei Ran, Kuai Yu, Dong Wang, Liyan Wen, Jianfen Zhuo, Yanjun Zhang, Yin Xi, Jingxian Zhao, Jincun Zhao and Jing Sun. Characterization of humoral immune responses against SARS-CoV-2 accessory proteins in infected patients and mouse model[J]. Virologica Sinica, 2024, 39(3): 414-421. doi: 10.1016/j.virs.2024.04.005
    Citation: Yuming Li, Yanhong Tang, Xiaoqian Wang, Airu Zhu, Dongdong Liu, Yiyun He, Hu Guo, Jie Zheng, Xinzhuo Liu, Fengyu Chi, Yanqun Wang, Zhen Zhuang, Zhaoyong Zhang, Donglan Liu, Zhao Chen, Fang Li, Wei Ran, Kuai Yu, Dong Wang, Liyan Wen, Jianfen Zhuo, Yanjun Zhang, Yin Xi, Jingxian Zhao, Jincun Zhao, Jing Sun. Characterization of humoral immune responses against SARS-CoV-2 accessory proteins in infected patients and mouse model .VIROLOGICA SINICA, 2024, 39(3) : 414-421.  http://dx.doi.org/10.1016/j.virs.2024.04.005
    shu

    COVID-19患者和SARS-CoV-2小鼠模型中病毒附属蛋白体液免疫反应特征分析

    cstr: 32224.14.j.virs.2024.04.005

    • a. 山东第一医科大学, 公共卫生与健康管理学院;

    • b. 广州医科大学附属第一医院, 呼吸疾病全国重点实验室;

    • c. State Key Laboratory of Respiratory Disease, National Clinical Research Center for Respiratory Disease, National Center for Respiratory Medicine, Guangzhou Institute of Respiratory Health, The First Affiliated Hospital of Guangzhou Medical University, Guangzhou, Guangdong, 510182, China;

    • d. Hunan Provincial People's Hospital, The First Affiliated Hospital of Hunan Normal University, Hunan Normal University, Changsha, 410005, China;

    • e. Guangzhou National Laboratory, Guangzhou, Guangdong, 510005, China;

    • f. . Shanghai Institute for Advanced Immunochemical Studies, School of Life Science and Technology, Shanghai Tech University, Shanghai, 201210, China;

    • g. . Institute for Hepatology, National Clinical Research Center for Infectious Disease, Shenzhen Third People's Hospital, The Second Affiliated Hospital, School of Medicine, Southern University of Science and Technology, Shenzhen, 518005, China

    • 严重急性呼吸系统综合征冠状病毒 2（SARS-CoV-2）是 COVID-19 的病原体，它可编码多个附属蛋白，这些蛋白在调节先天性免疫反应中起着至关重要的作用。然而，它们在病毒感染细胞中的表达以及在感染人和小鼠中的免疫原性了解并不充分。在本研究中，我们采用荧光素酶免疫沉淀系统（LIPS）、免疫荧光检测（IFA）和免疫印迹（WB）等多种技术检测了 COVID-19 患者血清中的附属蛋白特异性抗体。通过LIPS方法，可以检测到针对蛋白3a、3b、7b、8和9c的特异性抗体，但IFA和WB方法只能检测到蛋白3a的抗体。在LIPS检测结果中，蛋白 3a 和 7b 的抗体只在 ICU 患者中检测到，这也许能作为预测疾病进展的一种标志物。此外，我们还对 SARS-CoV-2 感染细胞中附属蛋白的表达进行鉴定分析，并确定了蛋白 3a、6、7a、8 和 9b 的表达。我们还分析了它们在免疫小鼠体内诱导抗体的能力，发现只有蛋白 3a、6、7a、8、9b 和 9c 能够诱导产生可测量的抗体，但这些抗体缺乏病毒中和活性，且不能保护小鼠免受 SARS-CoV-2的感染。我们的研究结果验证了SARS-CoV-2附属蛋白的表达，并阐明了它们的体液免疫反应，为蛋白检测试验及其在发病机制中的作用提供了依据。

    Characterization of humoral immune responses against SARS-CoV-2 accessory proteins in infected patients and mouse model

    • a. School of Public Health, Shandong First Medical University & Shandong Academy of Medical Sciences, Ji'nan, 250117, China;

    • b. Key Laboratory of Emerging Infectious Diseases in Universities of Shandong, Shandong First Medical University & Shandong Academy of Medical Sciences, Ji'nan, 250117, China;

    • c. State Key Laboratory of Respiratory Disease, National Clinical Research Center for Respiratory Disease, National Center for Respiratory Medicine, Guangzhou Institute of Respiratory Health, The First Affiliated Hospital of Guangzhou Medical University, Guangzhou, Guangdong, 510182, China;

    • d. Hunan Provincial People's Hospital, The First Affiliated Hospital of Hunan Normal University, Hunan Normal University, Changsha, 410005, China;

    • e. Guangzhou National Laboratory, Guangzhou, Guangdong, 510005, China;

    • f. Shanghai Institute for Advanced Immunochemical Studies, School of Life Science and Technology, Shanghai Tech University, Shanghai, 201210, China;

    • g. Institute for Hepatology, National Clinical Research Center for Infectious Disease, Shenzhen Third People's Hospital, The Second Affiliated Hospital, School of Medicine, Southern University of Science and Technology, Shenzhen, 518005, China

    • Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), the causative agent of COVID-19, encodes several accessory proteins that have been shown to play crucial roles in regulating the innate immune response. However, their expressions in infected cells and immunogenicity in infected humans and mice are still not fully understood. This study utilized various techniques such as luciferase immunoprecipitation system (LIPS), immunofluorescence assay (IFA), and western blot (WB) to detect accessory protein-specific antibodies in sera of COVID-19 patients. Specific antibodies to proteins 3a, 3b, 7b, 8 and 9c can be detected by LIPS, but only protein 3a antibody was detected by IFA or WB. Antibodies against proteins 3a and 7b were only detected in ICU patients, which may serve as a marker for predicting disease progression. Further, we investigated the expression of accessory proteins in SARS-CoV-2-infected cells and identified the expressions of proteins 3a, 6, 7a, 8, and 9b. We also analyzed their ability to induce antibodies in immunized mice and found that only proteins 3a, 6, 7a, 8, 9b and 9c were able to induce measurable antibody productions, but these antibodies lacked neutralizing activities and did not protect mice from SARS-CoV-2 infection. Our findings validate the expression of SARS-CoV-2 accessory proteins and elucidate their humoral immune response, providing a basis for protein detection assays and their role in pathogenesis.
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      Characterization of humoral immune responses against SARS-CoV-2 accessory proteins in infected patients and mouse model

        Corresponding author: Yin Xi, xiyin86@163.com
        Corresponding author: Jingxian Zhao, zhaojingxian@gird.cn
        Corresponding author: Jincun Zhao, zhaojincun@gird.cn
        Corresponding author: Jing Sun, sj-ji@163.com
      • a. School of Public Health, Shandong First Medical University & Shandong Academy of Medical Sciences, Ji'nan, 250117, China;
      • b. Key Laboratory of Emerging Infectious Diseases in Universities of Shandong, Shandong First Medical University & Shandong Academy of Medical Sciences, Ji'nan, 250117, China;
      • c. State Key Laboratory of Respiratory Disease, National Clinical Research Center for Respiratory Disease, National Center for Respiratory Medicine, Guangzhou Institute of Respiratory Health, The First Affiliated Hospital of Guangzhou Medical University, Guangzhou, Guangdong, 510182, China;
      • d. Hunan Provincial People's Hospital, The First Affiliated Hospital of Hunan Normal University, Hunan Normal University, Changsha, 410005, China;
      • e. Guangzhou National Laboratory, Guangzhou, Guangdong, 510005, China;
      • f. Shanghai Institute for Advanced Immunochemical Studies, School of Life Science and Technology, Shanghai Tech University, Shanghai, 201210, China;
      • g. Institute for Hepatology, National Clinical Research Center for Infectious Disease, Shenzhen Third People's Hospital, The Second Affiliated Hospital, School of Medicine, Southern University of Science and Technology, Shenzhen, 518005, China
      • Received Date: 05 November 2023
      • Accepted Date: 19 April 2024

      Abstract: Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), the causative agent of COVID-19, encodes several accessory proteins that have been shown to play crucial roles in regulating the innate immune response. However, their expressions in infected cells and immunogenicity in infected humans and mice are still not fully understood. This study utilized various techniques such as luciferase immunoprecipitation system (LIPS), immunofluorescence assay (IFA), and western blot (WB) to detect accessory protein-specific antibodies in sera of COVID-19 patients. Specific antibodies to proteins 3a, 3b, 7b, 8 and 9c can be detected by LIPS, but only protein 3a antibody was detected by IFA or WB. Antibodies against proteins 3a and 7b were only detected in ICU patients, which may serve as a marker for predicting disease progression. Further, we investigated the expression of accessory proteins in SARS-CoV-2-infected cells and identified the expressions of proteins 3a, 6, 7a, 8, and 9b. We also analyzed their ability to induce antibodies in immunized mice and found that only proteins 3a, 6, 7a, 8, 9b and 9c were able to induce measurable antibody productions, but these antibodies lacked neutralizing activities and did not protect mice from SARS-CoV-2 infection. Our findings validate the expression of SARS-CoV-2 accessory proteins and elucidate their humoral immune response, providing a basis for protein detection assays and their role in pathogenesis.

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