Ohkami Takehiro, Kitashin Ichika, Kawashima Riko, Yoshida Aimi, Saito Taizo, Takashima Yasuhiro, Kamitani Wataru, Nakagawa Keisuke
Laboratory of Veterinary Microbiology, Joint Department of Veterinary Medicine, Gifu University, Yanagido, Gifu 501-1193, Japan.
Joint Graduate School of Veterinary Sciences, Gifu University, Yanagido, Gifu 501-1193, Japan.
Viruses. 2025 Jul 13;17(7):978. doi: 10.3390/v17070978.
Coronavirus non-structural protein 1 (nsp1) is a pathogenic determinant of . Previous studies demonstrated that the nsp1 of various coronaviruses induces host shutoff through a variety of mechanisms; however, there is little information on the function of bovine coronavirus (BCoV) nsp1. We aimed to characterize the host gene expression suppression function of BCoV nsp1. We first confirmed that the expression of BCoV nsp1 in MAC-T cells, a bovine mammary epithelial cell line, suppressed host and reporter gene expression. Subsequently, lysine and phenylalanine at amino acid positions 232 and 233, respectively, were identified as key residues required for this suppressive effect. Expression levels of housekeeping genes are comparable in cells expressing wild-type BCoV nsp1 and a mutant with alanine substitutions at positions 232 and 233 (BCoV nsp1-KF). Wild-type BCoV nsp1 localized to both the cytoplasm and nucleus; however, BCoV nsp1-KF exhibited prominent nuclear accumulation with dot-like structures. Using confocal microscopy and co-sedimentation analysis, we identified an association between wild-type BCoV nsp1, but not BCoV nsp1-KF, and ribosomes, suggesting that ribosome binding is required for BCoV nsp1-mediated suppression of host gene expression. This is the first study of the characterization of host gene expression suppression by BCoV nsp1.
冠状病毒非结构蛋白1（nsp1）是……的致病决定因素。先前的研究表明，各种冠状病毒的nsp1通过多种机制诱导宿主关闭；然而，关于牛冠状病毒（BCoV）nsp1的功能知之甚少。我们旨在表征BCoV nsp1对宿主基因表达的抑制功能。我们首先证实，在牛乳腺上皮细胞系MAC-T细胞中BCoV nsp1的表达抑制了宿主基因和报告基因的表达。随后，分别位于氨基酸位置232和233的赖氨酸和苯丙氨酸被确定为这种抑制作用所需的关键残基。在表达野生型BCoV nsp1和在位置232和233处被丙氨酸取代的突变体（BCoV nsp1-KF）的细胞中，管家基因的表达水平相当。野生型BCoV nsp1定位于细胞质和细胞核；然而，BCoV nsp1-KF表现出明显的核积累并带有点状结构。使用共聚焦显微镜和共沉降分析，我们确定野生型BCoV nsp1（而非BCoV nsp1-KF）与核糖体之间存在关联，这表明核糖体结合是BCoV nsp1介导的宿主基因表达抑制所必需的。这是首次对BCoV nsp1抑制宿主基因表达的特性进行研究。
