College of Animal Science and Veterinary Medicine, Heilongjiang Bayi Agricultural University, No. 5 Xinfeng Road, Sartu District, Daqing 163319, China.
College of Animal Science and Veterinary Medicine, Heilongjiang Bayi Agricultural University, No. 5 Xinfeng Road, Sartu District, Daqing 163319, China; Key Laboratory of Applied Technology on Green-Eco-Healthy Animal Husbandry of Zhejiang Province, Zhejiang Provincial Engineering Research Center for Animal Health Diagnostics and Advanced Technology, Zhejiang International Science and Technology Cooperation Base for Veterinary Medicine and Health Management, China-Australia Joint Laboratory for Animal Health Big Data Analytics, College of Animal Science and Technology and College of Veterinary Medicine of Zhejiang A&F University, 666 Wusu Street, Lin'an District, Hangzhou, Zhejiang 311300, China.
Virus Res. 2022 Nov;321:198916. doi: 10.1016/j.virusres.2022.198916. Epub 2022 Sep 6.
Coronavirus subverts the host cell cycle to create a favorable cellular environment that enhances viral replication in host cells. Previous studies have revealed that nucleocapsid (N) protein of the coronavirus porcine epidemic diarrhea virus (PEDV) interacts with p53 to induce cell cycle arrest in S-phase and promotes viral replication. However, the mechanism by which viral replication is increased in the PEDV N protein-induced S-phase arrested cells remains unknown. In the current study, the protein expression profiles of PEDV N protein-induced S-phase arrested Vero E6 cells and thymidine-induced S-phase arrested Vero E6 cells were characterized by tandem mass tag-labeled quantitative proteomic technology. The effect of differentially expressed proteins (DEPs) on PEDV replication was investigated. The results indicated that a total of 5709 proteins, including 20,560 peptides, were identified, of which 58 and 26 DEPs were identified in the PEDV N group and thymidine group, respectively (P < 0.05; ratio ≥ 1.2 or ≤ 0.8). The unique DEPs identified in the PEDV N group were mainly involved in DNA replication, transcription, and protein synthesis, of which 60S ribosomal protein L18 (RPL18) exhibited significantly up-regulated expression in the PEDV N protein-induced S-phase arrested Vero E6/IPEC-J2 cells and PEDV-infected IPEC-J2 cells (P < 0.05). Further studies revealed that the RPL18 protein could significantly enhance PEDV replication (P < 0.05). Our findings reveal a mechanism regarding increased viral replication when the PEDV N protein-induced host cells are in S-phase arrest. These data also provide evidence that PEDV maintains its own replication by utilizing protein synthesis-associated ribosomal proteins.
冠状病毒会颠覆宿主细胞周期，创造一个有利于病毒在宿主细胞中复制的环境。先前的研究表明，冠状病毒猪流行性腹泻病毒（PEDV）的核衣壳（N）蛋白与 p53 相互作用，导致 S 期细胞周期停滞，并促进病毒复制。然而，PEDV N 蛋白诱导 S 期停滞细胞中病毒复制增加的机制尚不清楚。在本研究中，采用串联质量标签标记定量蛋白质组学技术对 PEDV N 蛋白诱导 S 期停滞 Vero E6 细胞和胸苷诱导 S 期停滞 Vero E6 细胞的蛋白质表达谱进行了分析。研究了差异表达蛋白（DEPs）对 PEDV 复制的影响。结果表明，共鉴定到 5709 种蛋白质，包括 20560 个肽段，其中 PEDV N 组和胸苷组分别鉴定到 58 种和 26 种 DEPs（P < 0.05；比值≥1.2 或≤0.8）。PEDV N 组中鉴定到的独特 DEPs 主要参与 DNA 复制、转录和蛋白质合成，其中 60S 核糖体蛋白 L18（RPL18）在 PEDV N 蛋白诱导的 S 期停滞 Vero E6/IPEC-J2 细胞和 PEDV 感染的 IPEC-J2 细胞中表达显著上调（P < 0.05）。进一步的研究表明，RPL18 蛋白可显著增强 PEDV 的复制（P < 0.05）。本研究揭示了 PEDV N 蛋白诱导的宿主细胞 S 期停滞时病毒复制增加的机制，也为 PEDV 通过利用与蛋白质合成相关的核糖体蛋白来维持自身复制提供了证据。
