Effects of N-Linked Glycan on Lassa Virus Envelope Glycoprotein Cleavage, Infectivity, and Immune Response

Xueqin Zhu; Yang Liu; Jiao Guo; Junyuan Cao; Zonglin Wang; Gengfu Xiao; Wei Wang

doi:10.1007/s12250-021-00358-y

August 2021

Xueqin Zhu, Yang Liu, Jiao Guo, Junyuan Cao, Zonglin Wang, Gengfu Xiao and Wei Wang. Effects of N-Linked Glycan on Lassa Virus Envelope Glycoprotein Cleavage, Infectivity, and Immune Response[J]. Virologica Sinica, 2021, 36(4): 774-783. doi: 10.1007/s12250-021-00358-y

Citation: Xueqin Zhu, Yang Liu, Jiao Guo, Junyuan Cao, Zonglin Wang, Gengfu Xiao, Wei Wang. Effects of N-Linked Glycan on Lassa Virus Envelope Glycoprotein Cleavage, Infectivity, and Immune Response .VIROLOGICA SINICA, 2021, 36(4) : 774-783. http://dx.doi.org/10.1007/s12250-021-00358-y

拉沙病毒包膜糖蛋白的N-糖基化修饰及其功能研究

朱雪芹 ^1,2 ,
刘洋 ¹ ,
郭娇 ^1,2 ,
曹浚垣 ^1,2 ,
王宗林 ^1,2 ,
肖庚富 ^1,2 ,
王薇 ^1,2,,

1.
中国科学院武汉病毒研究所病毒生物化学学科

通讯作者： 王薇, wangwei@wh.iov.cn, ORCID: http://orcid.org/0000-0002-8696-9773
收稿日期： 2020-11-02
录用日期： 2021-01-13
出版日期： 2021-03-10

摘要

拉沙病毒属沙粒病毒科哺乳类沙粒病毒属，人类感染沙粒病毒会引起严重的出血热疾病。拉沙病毒包膜糖蛋白GPC高度糖基化，糖链覆盖GPC表面，参与GPC成熟、介导病毒入侵、遮蔽抗原/中和表位帮助病毒实现免疫逃逸。考察不同糖基化修饰突变策略（天冬酰胺突变为丙氨酸、天冬酰胺突变为谷酰胺、丝氨酸/苏氨酸突变为丙氨酸）对GPC切割的影响，发现在第二（N89）、第五（N119）和第八（N365）糖基化位点，采用任何一种突变策略均导致GPC切割功能丧失。进一步研究发现第二（N89）和第八（N365）糖基化位点突变降低拉沙假病毒感染力。在此基础上，我们进一步考察N-糖基化修饰对免疫反应的影响。结果发现缺失第一（N79Q）、第三（N99Q）、第五（N119Q）和第六（N167Q）糖链能显著提高免疫小鼠脾脏细胞中CD4+细胞比例；而缺失第一（N79Q）、第二（N89Q）、第三（N99Q）、第四（N109Q）、第五（N119Q）、第六（N167Q）和第九（N373Q）糖链提高CD8+效应性T细胞比例；某些特定糖链缺失增强Th1型免疫反应。同时发现，用缺失GPC上任一糖链的质粒免疫小鼠，得到的血清与糖链缺失的GPC的滴度都有所增高，而对野生型拉沙假病毒的中和能力没有显著变化。说明糖链在遮蔽GPC抗原表位和中和表位中发挥重要作用。以上研究为拉沙病毒疫苗研发和基于糖链的抗体设计提供借鉴。
- 拉沙病毒
- , 包膜糖蛋白
- , N-糖基化修饰
- , 免疫反应

Effects of N-Linked Glycan on Lassa Virus Envelope Glycoprotein Cleavage, Infectivity, and Immune Response

Xueqin Zhu ^1,2 ,
Yang Liu ¹ ,
Jiao Guo ^1,2 ,
Junyuan Cao ^1,2 ,
Zonglin Wang ^1,2 ,
Gengfu Xiao ^1,2 ,
Wei Wang ^1,2,,

1.
State Key Laboratory of Virology, Wuhan Institute of Virology, Center for Biosafety Mega-Science, Chinese Academy of Sciences, Wuhan 430071, China
2.
University of the Chinese Academy of Sciences, Beijing 100049, China

Corresponding author: Wei Wang, wangwei@wh.iov.cn
ORCID: http://orcid.org/0000-0002-8696-9773
Received Date: 02 November 2020
Accepted Date: 13 January 2021
Published Date: 10 March 2021

Abstract

Lassa virus (LASV) belongs to the Mammarenavirus genus (family Arenaviridae) and causes severe hemorrhagic fever in humans. The glycoprotein complex (GPC) contains eleven N-linked glycans that play essential roles in GPC functionalities such as cleavage, transport, receptor recognition, epitope shielding, and immune response. We used three mutagenesis strategies (asparagine to glutamine, asparagine to alanine, and serine/tyrosine to alanine mutants) to abolish individual glycan chain on GPC and found that all the three strategies led to cleavage inefficiency on the 2nd (N89), 5th (N119), or 8th (N365) glycosylation motif. To evaluate N to Q mutagenesis for further research, it was found that deletion of the 2nd (N89Q) or 8th (N365Q) glycan completely inhibited the transduction efficiency of pseudotyped particles. We further investigated the role of individual glycan on GPC-mediated immune response by DNA immunization of mice. Deletion of the individual 1st (N79Q), 3rd (N99Q), 5th (N119Q), or 6th (N167Q) glycan significantly enhanced the proportion of effector CD4+ cells, whereas deletion of the 1st (N79Q), 2nd (N89Q), 3rd (N99Q), 4th (N109Q), 5th (N119Q), 6th (N167Q), or 9th (N373Q) glycan enhanced the proportion of CD8+ effector T cells. Deletion of specific glycan improves the Th1-type immune response, and abolishment of glycan on GPC generally increases the antibody titer to the glycan-deficient GPC. However, the antibodies from either the mutant or WT GPC-immunized mice show little neutralization effect on wild-type LASV. The glycan residues on GPC provide an immune shield for the virus, and thus represent a target for the design and development of a vaccine.
- Lassa virus (LASV)
- , Glycoprotein complex (GPC)
- , N-linked glycan
- , Immune response

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10.1007s12250-021-00358-y-ESM.pdf