Department of Veterinary Medicine, College of Animal Science, Southwest University, Chongqing, 402460, China.
Vet Res. 2019 Apr 27;50(1):28. doi: 10.1186/s13567-019-0648-9.
Transmissible gastroenteritis coronavirus (TGEV) is an enteropathogenic coronavirus that causes diarrhea in pigs and is associated with high morbidity and mortality in sucking piglets. S1 is one of two protein domains in the spike (S) glycoprotein and is responsible for enteric tropism, sialic acid recognition, and host receptor binding. Although there has been extensive research on the S1 protein of TGEV, little is known about the intracellular role of TGEV-S1. In the present study, we used yeast two-hybrid screening of a cDNA library from porcine intestinal cells to identify proteins that interact with TGEV-S1. Among 120 positive clones from the library, 12 intracellular proteins were identified after sequencing and a BLAST search. These intracellular proteins are involved in protein synthesis and degradation, biological signal transduction, and negative control of signaling pathways. Using a glutathione-S-transferase (GST) pulldown assay and Co-IP, we found that UBXN1 interacts with the S1 protein. Here, we observed that TGEV infection led to increased UBXN1 expression levels during the late phase of infection in IPEC-J2 cells. Inhibition of UBXN1 in IPEC-J2 cells via siRNA interference significantly decreased the viral titer and downregulated the expression of S1. UBXN1 overexpression significantly increased the viral copy number. Additionally, we provided data suggesting that UBXN1 negatively regulates IFN-β expression after TGEV infection. Finally, our research indicated that UBXN1 plays a vital role in the process of TGEV infection, making it a candidate target for the development of a novel antiviral method.
传染性胃肠炎冠状病毒（TGEV）是一种引起猪腹泻的肠致病性冠状病毒，与仔猪的高发病率和死亡率有关。S1 是刺突（S）糖蛋白中的两个蛋白结构域之一，负责肠嗜性、唾液酸识别和宿主受体结合。尽管已经对 TGEV 的 S1 蛋白进行了广泛的研究，但对 TGEV-S1 的细胞内作用知之甚少。在本研究中，我们使用酵母双杂交筛选来自猪肠细胞的 cDNA 文库，以鉴定与 TGEV-S1 相互作用的蛋白质。在文库的 120 个阳性克隆中，测序和 BLAST 搜索后鉴定出 12 种细胞内蛋白。这些细胞内蛋白参与蛋白质合成和降解、生物信号转导以及信号通路的负调控。通过谷胱甘肽 S-转移酶（GST）下拉测定和 Co-IP，我们发现 UBXN1 与 S1 蛋白相互作用。在这里，我们观察到在 IPEC-J2 细胞中，TGEV 感染导致晚期感染时 UBXN1 表达水平增加。通过 siRNA 干扰抑制 IPEC-J2 细胞中的 UBXN1 显著降低了病毒滴度并下调了 S1 的表达。UBXN1 的过表达显著增加了病毒拷贝数。此外，我们提供的数据表明 UBXN1 在 TGEV 感染后负调控 IFN-β 的表达。最后，我们的研究表明 UBXN1 在 TGEV 感染过程中发挥重要作用，使其成为开发新型抗病毒方法的候选靶标。
