Department of Preventive Veterinary Medicine, College of Veterinary Medicine, Northwest A&F University, Yangling, Shaanxi, China.
Department of Infection Biology and Microbiomes, Institute of Infection, Veterinary and Ecological Sciences, University of Liverpool, Liverpool, United Kingdom.
J Virol. 2024 Apr 16;98(4):e0164923. doi: 10.1128/jvi.01649-23. Epub 2024 Mar 29.
Hepatitis E virus (HEV) is the most common cause of acute viral hepatitis worldwide, responsible for approximately 20 million infections annually. Among the three open reading frames (ORFs) of the HEV genome, the ORF3 protein is involved in virus release. However, the host proteins involved in HEV release need to be clarified. In this study, a host protein, thioredoxin domain-containing protein 5 (TXNDC5), interacted with the non-palmitoylated ORF3 protein by co-immunoprecipitation analysis. We determined that the overexpression or knockdown of TXNDC5 positively regulated HEV release from the host cells. The FCL mutation of the ORF3 protein lost the ability to interact with TXNDC5. The releasing amounts of HEV with the ORF3 mutation (FCL17-19SSP) were decreased compared with wild-type HEV. The overexpression of TXNDC5 can stabilize and increase ORF3 protein amounts, but not the TXNDC5 mutant with amino acids 1-88 deletion. Meanwhile, we determined that the function of TXNDC5 on the stabilization of ORF3 protein is independent of the Trx-like domains. Knockdown of TXNDC5 could lead to the degradation of ORF3 protein by the endoplasmic reticulum (ER)-associated protein degradation-proteasome system. However, the ORF3 protein cannot be degraded in the knockout-TXNDC5 stable cells, suggesting that it may hijack other proteins for its stabilization. Subsequently, we found that the other members of protein disulfide isomerase (PDI), including PDIA1, PDIA3, PDIA4, and PDIA6, can increase ORF3 protein amounts, and PDIA3 and PDIA6 interact with ORF3 protein. Collectively, our study suggested that HEV ORF3 protein can utilize TXNDC5 for its stability in ER to facilitate viral release.
Hepatitis E virus (HEV) infection is the leading cause of acute viral hepatitis worldwide. After the synthesis and modification in the cells, the mature ORF3 protein is essential for HEV release. However, the host protein involved in this process has yet to be determined. Here, we reported a novel host protein, thioredoxin domain-containing protein 5 (TXNDC5), as a chaperone, contributing to HEV release by facilitating ORF3 protein stability in the endoplasmic reticulum through interacting with non-palmitoylated ORF3 protein. However, we also found that in the knockout-TXNDC5 stable cell lines, the HEV ORF3 protein may hijack other proteins for its stabilization. For the first time, our study demonstrated the involvement of TXNDC5 in viral particle release. These findings provide some new insights into the process of the HEV life cycle, the interaction between HEV and host factors, and a new direction for antiviral design.
戊型肝炎病毒（HEV）是全球范围内最常见的急性病毒性肝炎病原体，每年导致约 2000 万例感染。在 HEV 基因组的三个开放阅读框（ORF）中，ORF3 蛋白参与病毒释放。然而，需要阐明参与 HEV 释放的宿主蛋白。在本研究中，一种宿主蛋白，硫氧还蛋白结构域蛋白 5（TXNDC5），通过共免疫沉淀分析与非棕榈酰化的 ORF3 蛋白相互作用。我们确定 TXNDC5 的过表达或敲低可正向调节宿主细胞中 HEV 的释放。ORF3 蛋白的 FCL 突变丧失了与 TXNDC5 相互作用的能力。与野生型 HEV 相比，具有 ORF3 突变（FCL17-19SSP）的 HEV 的释放量减少。TXNDC5 的过表达可以稳定和增加 ORF3 蛋白的量，但氨基酸 1-88 缺失的 TXNDC5 突变体则不能。同时，我们确定 TXNDC5 对 ORF3 蛋白稳定性的作用不依赖于硫氧还蛋白样结构域。TXNDC5 的敲低会导致内质网（ER）相关蛋白降解-蛋白酶体系统降解 ORF3 蛋白。然而，在敲除-TXNDC5 的稳定细胞中，ORF3 蛋白不能被降解，表明它可能劫持其他蛋白来稳定自身。随后，我们发现其他蛋白二硫键异构酶（PDI）成员，包括 PDIA1、PDIA3、PDIA4 和 PDIA6，也可以增加 ORF3 蛋白的量，并且 PDIA3 和 PDIA6 与 ORF3 蛋白相互作用。综上所述，本研究表明 HEV ORF3 蛋白可利用 TXNDC5 在 ER 中稳定自身，从而促进病毒释放。
戊型肝炎病毒（HEV）感染是全球急性病毒性肝炎的主要病因。在细胞内合成和修饰后，成熟的 ORF3 蛋白对 HEV 的释放至关重要。然而，这一过程中涉及的宿主蛋白尚未确定。本研究中，我们发现一种新的宿主蛋白，硫氧还蛋白结构域蛋白 5（TXNDC5）作为伴侣蛋白，通过与非棕榈酰化的 ORF3 蛋白相互作用，促进 ORF3 蛋白在 ER 中的稳定性，从而有助于 HEV 的释放。然而，我们也发现，在敲除-TXNDC5 的稳定细胞系中，HEV ORF3 蛋白可能劫持其他蛋白来稳定自身。本研究首次证明了 TXNDC5 参与病毒粒子的释放。这些发现为 HEV 生命周期、HEV 与宿主因子相互作用的过程提供了一些新的见解，并为抗病毒设计提供了新的方向。
