早幼粒细胞白血病 (PML) 蛋白是 PML 核体 (PML-NB) 中不可或缺的元素，在调节多种细胞功能、同时协调针对病毒入侵的先天免疫反应中发挥着关键作用。同时，许多病毒通过针对 PML-NB 来逃避免疫检测。日本脑炎病毒 (JEV) 是一种引起日本脑炎的黄病毒，日本脑炎是一种影响人类和动物的严重神经系统疾病。然而，JEV 通过 PML-NB 逃避免疫的机制却很少被研究。在本研究中，PK15细胞被JEV感染，并计数细胞内PML-NB的数量。免疫荧光结果表明，与病毒抗原阴性细胞相比，JEV抗原阳性细胞中PML-NB的数量显着减少。随后，10 个 JEV 蛋白被克隆并转染至 PK15 细胞中。结果显示，JEV 非结构蛋白 NS2B、NS3、NS4A、NS4B 和 NS5 显着减少了 PML-NB 的数量。用五种 JEV 蛋白和各种猪 PML 亚型进行共转染。结果表明，NS2B 与 PML4 和 PML5 共定位，NS4A 与 PML1 和 PML4 共定位，NS4B 与 PML1、PML3、PML4 和 PML5 共定位，而 NS3 和 NS5 与所有五种 PML 亚型相互作用。此外，PML亚型的异位表达证实PML1、PML3、PML4和PML5抑制JEV复制。这些发现表明，JEV 通过与 PML 同工型相互作用破坏 PML-NB 的结构，可能导致宿主抗病毒免疫反应减弱。版权所有 © 2023 Yang、Liu、Chen、Wang、Li、Zhou 和 Zhao。

Promyelocytic leukemia (PML) protein constitutes an indispensable element within PML-nuclear bodies (PML-NBs), playing a pivotal role in the regulation of multiple cellular functions while coordinating the innate immune response against viral invasions. Simultaneously, numerous viruses elude immune detection by targeting PML-NBs. Japanese encephalitis virus (JEV) is a flavivirus that causes Japanese encephalitis, a severe neurological disease that affects humans and animals. However, the mechanism through which JEV evades immunity via PML-NBs has been scarcely investigated. In the present study, PK15 cells were infected with JEV, and the quantity of intracellular PML-NBs was enumerated. The immunofluorescence results indicated that the number of PML-NBs was significantly reduced in JEV antigen-positive cells compared to viral antigen-negative cells. Subsequently, ten JEV proteins were cloned and transfected into PK15 cells. The results revealed that JEV non-structural proteins, NS2B, NS3, NS4A, NS4B, and NS5, significantly diminished the quantity of PML-NBs. Co-transfection was performed with the five JEV proteins and various porcine PML isoforms. The results demonstrated that NS2B colocalized with PML4 and PML5, NS4A colocalized with PML1 and PML4, NS4B colocalized with PML1, PML3, PML4, and PML5, while NS3 and NS5 interacted with all five PML isoforms. Furthermore, ectopic expression of PML isoforms confirmed that PML1, PML3, PML4, and PML5 inhibited JEV replication. These findings suggest that JEV disrupts the structure of PML-NBs through interaction with PML isoforms, potentially leading to the attenuation of the host's antiviral immune response.Copyright © 2023 Yang, Liu, Chen, Wang, Li, Zhou and Zhao.