NSMCE1 suppresses FMDV replication by promoting the ubiquitination and degradation of 3A

doi:10.1016/j.jia.2026.04.003

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Sujie Dong¹^,²^*, Xiaodong Qin¹^,²^*^#, Yongjie Liu¹^,², Taoqing Zhang¹^,², Yuanshu Wu¹^,², Zhengwang Shi¹^,², Shuaipeng Li¹^,², Xiaolan Qi¹^,², Rongzeng Hao¹^,², Haixue Zheng¹^,²^#

¹State Key Laboratory of Animal Disease Control and Prevention, College of Veterinary Medicine, Lanzhou University, Lanzhou Veterinary Research Institute, Chinese Academy of Agricultural Sciences, Lanzhou 730046, China

² Gansu Province Research Center for Basic Disciplines of Pathogen Biology, Lanzhou 730046, China

Highlights

1. The E3 ubiquitin ligase NSMCE1 is a novel host factor that suppresses FMDV replication.

2. NSMCE1 interacts with the FMDV 3A and mediates its K33-linked ubiquitination at K16, leading to proteasomal degradation of 3A.

3. Mutation of 3A K16 enhances FMDV replication capacity both in vitro and in vivo, confirming the critical role of NSMCE1-mediated ubiquitination in restricting viral infection.

Abstract
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摘要

口蹄疫病毒（FMDV）是一种高度接触性传染的小RNA病毒，可感染猪、牛、羊等多种偶蹄动物，在全球范围内给畜禽养殖业造成了巨大的经济损失，是全球重点防控的动物烈性传染病之一。FMDV非结构蛋白3A是调控病毒复制、致病性及宿主嗜性的关键功能分子。现有研究证实，3A蛋白的氨基酸突变与病毒宿主范围改变、毒力强弱变化密切相关，在病毒与宿主细胞相互作用过程中发挥核心介导作用。但是目前针对宿主细胞因子靶向调控3A蛋白、进而干预病毒复制的分子机制仍不明确。为挖掘参与调控FMDV复制的宿主关键因子并阐明其作用机制，本研究采用western blot、RT-qPCR、TCID₅₀、免疫共沉淀(Co-IP)、免疫荧光共定位、反向遗传操作等技术手段，通过一系列体内外功能验证实验，鉴定出 E3 泛素连接酶NSMCE1为新的宿主抗FMDV负调控因子。

研究结果表明，NSMCE1抑制FMDV的复制，该分子可与FMDV 3A蛋白发生相互作用，并特异性介导3A蛋白第16位赖氨酸（K16）位点发生K33型泛素化修饰。该修饰通过泛素-蛋白酶体途径促进3A蛋白降解，进而抑制FMDV的复制，发挥其抗病毒活性。通过反向遗传技术构建3A蛋白K16位点突变的重组病毒（rO-3AK16R），体内外功能验证结果表明该位点突变可逃逸NSMCE1介导的泛素化降解，显著增强FMDV在体外及体内的复制能力与致病力，进一步验证了NSMCE1介导的泛素化修饰在抑制病毒复制中的关键作用。

本研究首次揭示了NSMCE1通过靶向调控FMDV 3A蛋白抵御病毒感染的全新分子机制，明确了3A蛋白K16位点为NSMCE1介导泛素化修饰的关键靶点，阐明了泛素-蛋白酶体系统在小RNA病毒复制调控中的重要功能。研究结果不仅深化了宿主天然抗病毒免疫应答机制的理论认知，也为靶向泛素化通路研发新型抗口蹄疫药物提供了潜在靶点与理论依据。

Abstract

Foot-and-mouth disease virus (FMDV) is a highly contagious picornavirus that causes severe economic losses in livestock worldwide. The nonstructural protein 3A of FMDV is essential for viral replication and virulence, and mutations in 3A are associated with altered host tropism, highlighting its role in mediating host-virus interactions. However, the molecular mechanisms underlying the interplay between 3A and host cellular factors remain poorly understood. Here, through systematic screening and functional analyses, we identify the E3 ubiquitin ligase NSMCE1 as a novel host-encoded negative regulator of FMDV replication. NSMCE1 interacts directly with the FMDV 3A protein and mediates its K33-linked ubiquitination at lysine 16 (K16). This modification promotes the proteasomal degradation of 3A, thereby suppressing FMDV replication. Consistent with this mechanism, recombinant virus with a mutation at lysine 16 of 3A enhances the replication capacity of FMDV both in vitro and in vivo, confirming the critical role of this regulatory event. Our findings reveal a previously unrecognized role for NSMCE1 in limiting FMDV infection through targeted regulation of viral protein 3A and uncover a regulatory role of the ubiquitin-proteasome system in picornavirus replication. These insights advance our understanding of host antiviral defense mechanisms and provide a potential foundation for the development of novel antiviral therapies targeting the ubiquitin pathway.

Keywords: foot-and-mouth disease virus 3A NSMCE1 ubiquitin-proteasome system viral replication

Online: 07 April 2026

Fund:

This study was supported by the Key Project of National Natural Sciences Foundation of China (no. 32330107), The open competition program of top ten critical priorities of Agricultural Science and Technology Innovation for the 14th Five-Year Plan of Guangdong Province (no. 2024KJ14), the National Key R&D Program of China (no. 2021YFD1800300), the STI 2030-Major Projects (no. 2023ZD0404301), the Project of National Center of Technology Innovation for Pigs (no. NCTIP-XD/C03), the Innovation Program of the Chinese Academy of Agricultural Sciences(no. CAAS-CSLPDCP-2023002), the Earmarked Fund (no. CARS-35) and the Project of Gansu Province's R&D-Industry Integration Technology R&D Empowerment Program (no. 25FNNA002).

About author: #Correspondence Xiaodong Qin, E-mail: qinxiaodong@caas.cn; Haixue Zheng, E-mail: zhenghaixue@caas.cn * Sujie Dong and Xiaodong Qin contributed equally to this article.

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Sujie Dong, Xiaodong Qin, Yongjie Liu, Taoqing Zhang, Yuanshu Wu, Zhengwang Shi, Shuaipeng Li, Xiaolan Qi, Rongzeng Hao, Haixue Zheng. 2026. NSMCE1 suppresses FMDV replication by promoting the ubiquitination and degradation of 3A. Journal of Integrative Agriculture, Doi:10.1016/j.jia.2026.04.003

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