The E301R protein of African swine fever virus functions as a sliding clamp involved in viral genome replication

Author:

Li Su1ORCID,Ge Hailiang12,Li Yanhua3,Zhang Kehui1,Yu Shaoxiong1,Cao Hongwei1,Wang Yanjin1,Deng Hao1,Li Jiaqi1,Dai Jingwen1,Li Lian-Feng1,Luo Yuzi1,Sun Yuan1,Geng Zhi3,Dong Yuhui3,Zhang Heng3ORCID,Qiu Hua-Ji1ORCID

Affiliation:

1. State Key Laboratory for Animal Disease Control and Prevention, National African Swine Fever Para-Reference Laboratory, National High-Containment Facilities for Animal Disease Control and Prevention, Harbin Veterinary Research Institute, Chinese Academy of Agricultural Sciences , Harbin, China

2. College of Animal Sciences, Yangtze University , Jingzhou, China

3. Multidiscipline Research Center, Institute of High Energy Physics, Chinese Academy of Sciences , Beijing, China

Abstract

ABSTRACT African swine fever virus (ASFV) is a complex nucleocytoplasmic, large DNA virus that infects both domestic pigs and wild boar, but little is known about the process of genomic replication. The E301R protein (pE301R) from ASFV was previously predicted as a proliferating cell nuclear antigen (PCNA)-like protein through clamping DNA polymerase to the DNA duplex, but its exact structure and functions remain uncharacterized. Here, the crystal structure of pE301R revealed that it is composed of structurally similar head and tail domains and shares significant structural similarities to the DNA polymerase processivity factors, including sliding clamp and eukaryotic PCNA. More specifically, we demonstrated that pE301R exhibited multiple oligomeric states (with dimers and tetramers dominant), and the tetramers are consistent with the ring-shaped homotetramers in a head-to-tail manner generated by crystal packing. We also showed that pE301R interacted with the ASFV genome and viral DNA polymerase O174L. Furthermore, knockdown of E301R by specific small interfering RNAs (siRNAs) significantly decreased the virus genomic replication. Interestingly, the downregulation of PCNA by siRNAs significantly decreased the cell viability, whereas the inhibitory effect was reversed by pE301R overexpression. Notably, we demonstrated that overexpression of PCNA partially restored ASFV replication upon transfection of siRNAs targeting E301R . More importantly, T2 amino alcohol, a PCNA-specific inhibitor, markedly inhibited ASFV replication at the stage of viral genome replication. Taken together, we revealed that pE301R functions as a sliding clamp in ASFV genomic replication and can be used as a potential antiviral target. IMPORTANCE Sliding clamp is a highly conserved protein in the evolution of prokaryotic and eukaryotic cells. The sliding clamp is required for genomic replication as a critical co-factor of DNA polymerases. However, the sliding clamp analogs in viruses remain largely unknown. We found that the ASFV E301R protein (pE301R) exhibited a sliding clamp-like structure and similar functions during ASFV replication. Interestingly, pE301R is assembled into a unique ring-shaped homotetramer distinct from sliding clamps or proliferating cell nuclear antigens (PCNAs) from other species. Notably, the E301R gene is required for viral life cycle, but the pE301R function can be partially restored by the porcine PCNA. This study not only highlights the functional role of the ASFV pE301R as a viral sliding clamp analog, but also facilitates the dissection of the complex replication mechanism of ASFV, which provides novel clues for developing antivirals against ASF.

Funder

MOST | National Key Research and Development Program of China

MOST | National Natural Science Foundation of China

National Science Foudation of Heilongjaing Province of China

Publisher

American Society for Microbiology

Subject

Virology,Microbiology

Cited by 4 articles. 订阅此论文施引文献 订阅此论文施引文献,注册后可以免费订阅5篇论文的施引文献,订阅后可以查看论文全部施引文献

同舟云学术

1.学者识别学者识别

2.学术分析学术分析

3.人才评估人才评估

"同舟云学术"是以全球学者为主线,采集、加工和组织学术论文而形成的新型学术文献查询和分析系统,可以对全球学者进行文献检索和人才价值评估。用户可以通过关注某些学科领域的顶尖人物而持续追踪该领域的学科进展和研究前沿。经过近期的数据扩容,当前同舟云学术共收录了国内外主流学术期刊6万余种,收集的期刊论文及会议论文总量共计约1.5亿篇,并以每天添加12000余篇中外论文的速度递增。我们也可以为用户提供个性化、定制化的学者数据。欢迎来电咨询!咨询电话:010-8811{复制后删除}0370

www.globalauthorid.com

TOP

Copyright © 2019-2024 北京同舟云网络信息技术有限公司
京公网安备11010802033243号  京ICP备18003416号-3