Raza Sohail, Shahin Farzana, Zhai Wenjun, Li Hanxiong, Alvisi Gualtiero, Yang Kui, Chen Xi, Chen Yingyu, Chen Jianguo, Hu Changmin, Chen Huanchun, Guo Aizhen
The State Key Laboratory of Agricultural Microbiology, Huazhong Agricultural University, Wuhan 430070, China.
Department of Microbiology, University of Veterinary and Animal Sciences, Lahore 54000, Pakistan.
Microorganisms. 2020 Mar 13;8(3):409. doi: 10.3390/microorganisms8030409.
(BoHV-1) is a major bovine pathogen. Despite several vaccines being available to prevent viral infection, outbreaks are frequent and cause important economic consequences worldwide. The development of new antiviral drugs is therefore highly desirable. In this context, viral genome replication represents a potential target for therapeutic intervention. BoHV-1 genome is a dsDNA molecule whose replication takes place in the nuclei of infected cells and is mediated by a viral encoded DNA polymerase holoenzyme. Here, we studied the physical interaction and subcellular localization of BoHV-1 DNA polymerase subunits in cells for the first time. By means of co-immunoprecipitation and confocal laser scanning microscopy (CLSM) experiments, we could show that the processivity factor of the DNA polymerase pUL42 is capable of being autonomously transported into the nucleus, whereas the catalytic subunit pUL30 is not. Accordingly, a putative classic NLS (cNLS) was identified on pUL42 but not on pUL30. Importantly, both proteins could interact in the absence of other viral proteins and their co-expression resulted in accumulation of UL30 to the cell nucleus. Treatment of cells with Ivermectin, an anti-parasitic drug which has been recently identified as an inhibitor of importin α/β-dependent nuclear transport, reduced UL42 nuclear import and specifically reduced BoHV-1 replication in a dose-dependent manner, while virus attachment and entry into cells were not affected. Therefore, this study provides a new option of antiviral therapy for BoHV-1 infection with Ivermectin.
牛疱疹病毒1型（BoHV-1）是一种主要的牛病原体。尽管有几种疫苗可用于预防病毒感染，但疫情仍频繁发生，并在全球范围内造成重大经济后果。因此，非常需要开发新的抗病毒药物。在这种背景下，病毒基因组复制成为治疗干预的一个潜在靶点。BoHV-1基因组是一个双链DNA分子，其复制发生在受感染细胞的细胞核中，并由病毒编码的DNA聚合酶全酶介导。在这里，我们首次研究了BoHV-1 DNA聚合酶亚基在细胞中的物理相互作用和亚细胞定位。通过免疫共沉淀和共聚焦激光扫描显微镜（CLSM）实验，我们可以表明DNA聚合酶pUL42的持续合成因子能够自主转运到细胞核中，而催化亚基pUL30则不能。因此，在pUL42上鉴定出一个假定的经典核定位信号（cNLS），而在pUL30上未鉴定出。重要的是，这两种蛋白质在没有其他病毒蛋白的情况下可以相互作用，并且它们的共表达导致UL30在细胞核中积累。用伊维菌素处理细胞，伊维菌素是一种抗寄生虫药物，最近被确定为importinα/β依赖性核转运的抑制剂，可减少UL42的核输入，并以剂量依赖的方式特异性降低BoHV-1的复制，而病毒附着和进入细胞不受影响。因此，本研究为用伊维菌素治疗BoHV-1感染提供了一种新的抗病毒治疗选择。
