Acta Veterinaria et Zootechnica Sinica ›› 2024, Vol. 55 ›› Issue (9): 4100-4109.doi: 10.11843/j.issn.0366-6964.2024.09.034

• Basic Veterinary Medicine • Previous Articles     Next Articles

Construction of Interferon Regulatory Factor Knockdown Cell Line and Its Effect on Pseudorabies Virus Proliferation

Yiqian FU1,2,3(), Dongge LIANG1,2,3, Mingyang WANG1,2,3, Jiajia PAN1,2,3, Yanbin YANG1,2,3, Lei ZENG1,2,3,*(), Xiangtao KANG4,*()   

  1. 1. College of Animal Medicine, Henan Agricultural University, Zhengzhou 450046, China
    2. Key Laboratory of Animal Biochemistry and Nutrition, Ministry of Agriculture and Rural Areas, Henan Agricultural University, Zhengzhou 450046, China
    3. Henan Key Laboratory of Animal Growth and Development Regulation, Henan Agricultural University, Zhengzhou 450046, China
    4. College of Animal Science and Technology, Henan Agricultural University, Zhengzhou 450046, China
  • Received:2023-11-14 Online:2024-09-23 Published:2024-09-27
  • Contact: Lei ZENG, Xiangtao KANG E-mail:1050223216@qq.com;zenglei2021918@163.com;xtkang2001@263.net

Abstract:

To elucidate the impact of interferon regulatory factors (IRFs) on the proliferation of porcine pseudorabies virus (PRV), this study utilized shRNA technology to establish a PK15 cell line with the targeted knockdown of IRF1-9 genes. The knockdown efficiency was verified, and the proliferation of PRV subsequent to the knockdown was assessed using flow cytometry and titration assays. Additionally, the mRNA expression levels of PRV-gB, IFN-β, ISG-15, and IL-6 in PRV-infected cells were quantified by reverse transcription quantitative polymerase chain reaction (RT-qPCR). The expression of the gB protein in PRV-infected cells was analyzed by Western blotting. The knockdown efficiency assay demonstrated a significant reduction in the expression of IRF1-9 mRNA in PK15 cells. The results from flow cytometry and titration assays indicated that the knockdown of IRF1-9 genes facilitated the proliferation of PRV. Furthermore, RT-qPCR and Western blot results revealed a significant increase in the mRNA and protein expression levels of PRV-gB following the knockdown of IRF1-9 genes. The analysis of mRNA expression levels of cellular inflammatory factors showed that the knockdown of IRF1-9 inhibited the PRV-induced expression of IFN-β, ISG-15, and IL-6 at the mRNA level. In conclusion, these findings suggest that IRF1-9 serves as a host restriction factor, limiting the replication of PRV within PK-15 cells.

Key words: shRNA, interferon regulatory factor, pseudorabies virus, inflammatory cytokines

CLC Number: