College of Veterinary Medicine, Northwest A&F University, Yangling, Shaanxi, China.
J Virol. 2023 Feb 28;97(2):e0175122. doi: 10.1128/jvi.01751-22. Epub 2023 Feb 8.
Porcine epidemic diarrhea virus (PEDV) belongs to the genus of the Coronaviridae family and can cause fatal watery diarrhea in piglets, causing significant economic losses. Heterogeneous nuclear protein U (HNRNPU) is a novel RNA sensor involved in sensing viral RNA in the nucleus and mediating antiviral immunity. However, it remains elusive whether and how cytoplasmic PEDV can be sensed by the RNA sensor HNRNPU. In this study we determined that HNRNPU was the binding partner of Nsp13 by immunoprecipitation-liquid chromatography-tandem mass spectrometry (IP/LC-MS/MS) analysis. The interaction between Nsp13 and HNRNPU was demonstrated by using coimmunoprecipitation and confocal immunofluorescence. Next, we identified that HNRNPU expression is significantly increased during PEDV infection, whereas the transcription factor hepatocyte nuclear factor 1α (HNF1A) could negatively regulate HNRNPU expression. HNRNPU was retained in the cytoplasm by interaction with PEDV Nsp13. We found that HNRNPU overexpression effectively facilitated PEDV replication, while knockdown of HNRNPU impaired viral replication, suggesting a promoting function of HNRNPU to PEDV infection. Additionally, HNRNPU was found to promote PEDV replication by affecting TRAF3 degradation at the transcriptional level to inhibit PEDV-induced beta interferon (IFN-β) production. Mechanistically, HNRNPU downregulates TRAF3 mRNA levels via the METTL3-METTL14/YTHDF2 axis and regulates immune responses through YTHDF2-dependent mRNA decay. Together, our findings reveal that HNRNPU serves as a negative regulator of innate immunity by degrading TRAF3 mRNA in a YTHDF2-dependent manner and consequently facilitating PEDV propagation. Our findings provide new insights into the immune escape of PEDV. PEDV, a highly infectious enteric coronavirus, has spread rapidly worldwide and caused severe economic losses. During virus infection, the host regulates innate immunity to inhibit virus infection. However, PEDV has evolved a variety of different strategies to suppress host IFN-mediated antiviral responses. Here, we identified that HNRNPU interacted with viral protein Nsp13. HNRNPU protein expression was upregulated, and the transcription factor HNF1A could negatively regulate HNRNPU expression during PEDV infection. HNRNPU also downregulated TRAF3 mRNA through the METTL3-METTL14/YTHDF2 axis to inhibit the production of IFN-β and downstream antiviral genes in PEDV-infected cells, thereby promoting viral replication. Our findings reveal a new mechanism with which PEDV suppresses the host antiviral response.
猪流行性腹泻病毒（PEDV）属于冠状病毒科的冠状病毒属，可导致仔猪致命性水样腹泻，造成重大经济损失。异质核核糖核蛋白 U（HNRNPU）是一种新型的 RNA 传感器，参与细胞核中病毒 RNA 的感应，并介导抗病毒免疫。然而，细胞质中的 PEDV 是否以及如何被 RNA 传感器 HNRNPU 感应仍然不清楚。在这项研究中，我们通过免疫沉淀-液相色谱-串联质谱（IP/LC-MS/MS）分析确定 HNRNPU 是 Nsp13 的结合伴侣。通过共免疫沉淀和共聚焦免疫荧光实验证实了 Nsp13 和 HNRNPU 之间的相互作用。接下来，我们发现 PEDV 感染期间 HNRNPU 的表达显著增加，而转录因子肝细胞核因子 1α（HNF1A）可负调控 HNRNPU 的表达。PEDV Nsp13 通过与 HNRNPU 的相互作用将其保留在细胞质中。我们发现 HNRNPU 的过表达可有效促进 PEDV 的复制，而 HNRNPU 的敲低则损害病毒的复制，表明 HNRNPU 对 PEDV 感染具有促进作用。此外，我们发现 HNRNPU 通过影响 TRAF3 的降解在转录水平上促进 PEDV 复制，从而抑制 PEDV 诱导的β干扰素（IFN-β）产生。机制上，HNRNPU 通过 METTL3-METTL14/YTHDF2 轴下调 TRAF3 mRNA 水平，并通过 YTHDF2 依赖性 mRNA 降解调节免疫反应。总之，我们的研究结果表明，HNRNPU 通过 YTHDF2 依赖性方式降解 TRAF3 mRNA，从而作为先天免疫的负调节剂，促进 PEDV 的传播。我们的研究结果为 PEDV 的免疫逃逸提供了新的见解。
PEDV 是一种高度传染性的肠冠状病毒，已在全球范围内迅速传播并造成严重的经济损失。在病毒感染过程中，宿主调节先天免疫以抑制病毒感染。然而，PEDV 已进化出多种不同的策略来抑制宿主 IFN 介导的抗病毒反应。在这里，我们发现 HNRNPU 与病毒蛋白 Nsp13 相互作用。在 PEDV 感染期间，HNRNPU 蛋白表达上调，转录因子 HNF1A 可负调控 HNRNPU 的表达。HNRNPU 还通过 METTL3-METTL14/YTHDF2 轴下调 TRAF3 mRNA，从而抑制 PEDV 感染细胞中 IFN-β 和下游抗病毒基因的产生，从而促进病毒复制。我们的研究结果揭示了 PEDV 抑制宿主抗病毒反应的一种新机制。
