脯氨酰寡肽酶促进口蹄疫病毒在PK15细胞中的复制

doi:10.11843/j.issn.0366-6964.2025.09.037

畜牧兽医学报 ›› 2025, Vol. 56 ›› Issue (9): 4593-4603.doi: 10.11843/j.issn.0366-6964.2025.09.037

脯氨酰寡肽酶促进口蹄疫病毒在PK15细胞中的复制

王姿逸¹^,²^,³(), 茹毅¹^,², 卢炳州¹^,², 杨洋¹^,², 赵陇和¹^,², 李亚军¹^,², 李建斌¹^,², 李明桂¹^,², 马坤¹^,², 冷非凡³, 郝荣增¹^,²^,*(), 郑海学¹^,²^,³^,*()

1. 中国农业科学院兰州兽医研究所兰州大学动物医学与生物安全学院动物疫病防控全国重点实验室, 兰州 730046
2. 甘肃省病原生物学基础学科研究中心, 兰州 730046
3. 兰州理工大学生命科学与工程学院, 兰州 730050

收稿日期:2024-12-12 出版日期:2025-09-23 发布日期:2025-09-30
通讯作者: 郝荣增,郑海学 E-mail:215391193@qq.com;haorongzeng@163.com;zhenghaixue@caas.cn
作者简介:王姿逸(1998-)，女，甘肃陇南人，硕士生，主要从事生物与医药研究，E-mail: 215391193@qq.com
基金资助:
国家自然科学基金(32372990)；甘肃省自然科学基金重点项目(23YFNA0011)；甘肃省自然科学基金(23JRRA549)

Prolyl Oligopeptidase Promotes the Replication of Foot and Mouth Disease Virus in PK15 Cells

WANG Ziyi¹^,²^,³(), RU Yi¹^,², LU Bingzhou¹^,², YANG Yang¹^,², ZHAO Longhe¹^,², LI Yajun¹^,², LI Jianbin¹^,², LI Minggui¹^,², MA Kun¹^,², LENG Feifan³, HAO Rongzeng¹^,²^,*(), ZHENG Haixue¹^,²^,³^,*()

1. State Key Laboratory for Animal Disease Control and Prevention, College of Veterinary Medicine of Lanzhou University, Lanzhou Veterinary Research Institute, Chinese Academy of Agricultural Sciences, Lanzhou 730046, China
2. Gansu Provincial Research Center of Basic Discipline of Pathogen Biology, Lanzhou 730046, China
3. School of Life Science and Engineering, Lanzhou University of Technology, Lanzhou 730050, China

Received:2024-12-12 Online:2025-09-23 Published:2025-09-30
Contact: HAO Rongzeng, ZHENG Haixue E-mail:215391193@qq.com;haorongzeng@163.com;zhenghaixue@caas.cn

摘要/Abstract

摘要：

本研究旨在探究宿主蛋白脯氨酰寡肽酶(prolyl oligopeptidase, POP)在PK15细胞中对口蹄疫病毒(foot and mouth disease virus, FMDV)复制的影响。首先利用实时荧光定量聚合酶链式反应(RT-qPCR)和蛋白免疫印迹(Western blot)分析FMDV感染PK15细胞后POP蛋白的表达变化。进一步构建POP真核表达质粒，通过病毒半数细胞感染量(TCID₅₀)、Western blot和RT-qPCR检测过表达POP对FMDV复制的影响；合成针对POP基因的特异性siRNA，利用TCID₅₀、Western blot和RT-qPCR检测siRNA对POP表达的干扰效果及POP被干扰后对FMDV复制的影响。结果表明，FMDV感染PK15细胞后对宿主细胞POP的mRNA和蛋白表达均没有显著影响，而过表达POP能显著促进FMDV在PK15细胞中复制水平，并且病毒复制水平随着POP的剂量递增呈现剂量依赖式增加；siRNA-S112显著下调内源性POP表达，进而显著抑制FMDV的复制。本研究首先在宿主细胞水平揭示了POP蛋白对FMDV复制的影响，证明POP对FMDV复制具有显著的促进作用，研究结果为进一步探究POP促进FMDV的复制及作用机制提供了基础。

关键词: 脯氨酰寡肽酶, 口蹄疫病毒, 病毒复制, PK15细胞

Abstract:

The study aims to investigate the impact of prolyl oligopeptidase (POP) on the replication of foot-and-mouth disease virus (FMDV) in PK15 cells. Firstly, the effect of FMDV infection on POP expression in PK15 cells was analyzed using Western blot and real-time quantitative PCR (RT-qPCR). Secondly, a POP eukaryotic expression plasmid was constructed, and the effect of overexpression of POP on FMDV replication was assessed through Western blot, RT-qPCR, and virus titer assays (TCID₅₀); Thirdly, specific siRNA targeting the POP gene was synthesized, and effective siRNAs sequences for interfering with endogenous POP expression were identified using RT-qPCR. The impact of downregulating endogenous POP expression on FMDV replication was further evaluated by Western blot, RT-qPCR, and virus titer assays (TCID₅₀). The results indicated that FMDV infection did not significantly affect POP expression in PK15 cells. Overexpression of POP significantly promoted FMDV replication in PK15 cells, with viral replication level increasing in a dose-dependent manner as the dosage of POP expression increased. Downregulation of endogenous POP expression using siRNA significantly inhibited FMDV replication. This study revealed that the promoting effect of POP protein on FMDV replication from the host cell level at the first time. This finding provides a foundation for further exploration of the mechanism by which POP promotes FMDV replication.

Key words: prolyl oligopeptidase (POP), foot-and-mouth disease virus, virus replication, PK15 cells

中图分类号:

S852.659.6

王姿逸, 茹毅, 卢炳州, 杨洋, 赵陇和, 李亚军, 李建斌, 李明桂, 马坤, 冷非凡, 郝荣增, 郑海学. 脯氨酰寡肽酶促进口蹄疫病毒在PK15细胞中的复制[J]. 畜牧兽医学报, 2025, 56(9): 4593-4603.

WANG Ziyi, RU Yi, LU Bingzhou, YANG Yang, ZHAO Longhe, LI Yajun, LI Jianbin, LI Minggui, MA Kun, LENG Feifan, HAO Rongzeng, ZHENG Haixue. Prolyl Oligopeptidase Promotes the Replication of Foot and Mouth Disease Virus in PK15 Cells[J]. Acta Veterinaria et Zootechnica Sinica, 2025, 56(9): 4593-4603.

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引物名称 Primer names	正向序列(5′→3′) Sense	反向序列(5′→3′) Antisense
POP	GAATTCTACCCATACGATGTTCCAGATTACGCT CTGTCCTTCCAGTACCCCGA	GGTACCACATCGACTGGATTCCGTAA
FMDV-SYBR	TGGGACCATACAGGAGAAGT	GTAGCTTGGAATCTCGAAGAGG
SusGAPDH	ACTGAGGACCAGGTTGTGT	AGGAAATGAGCTTGACGAAGTG
POPsiRNA-112	GGAUUAUCAUGGACAUAAATT	UUUAUGUCCAUGAUAAUCCTT
POPsiRNA-563	CUUGAAAGAGUCAAGUUCATT	UGAACUUGACUCUUUCAAGTT
POPsiRNA-1404	CAAAGAUCCCCAUGUUCAUTT	AUGAACAUGGGGAUCUUUGTT
NC-siRNA	UUCUCCGAACGUGUCACGUTT	ACGUGACACGUUCGGAGAATT

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脯氨酰寡肽酶促进口蹄疫病毒在PK15细胞中的复制

Prolyl Oligopeptidase Promotes the Replication of Foot and Mouth Disease Virus in PK15 Cells

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