Tight Junction Protein Occludin Is a Porcine Epidemic Diarrhea Virus Entry Factor

Luo, Xuenong; Guo, Longjun; Zhang, Jian; Xu, Yunfei; Gu, Weihong; Feng, Li; Wang, Yue

doi:10.1128/jvi.00202-17

Cited by 68 publications

(50 citation statements)

References 67 publications

(77 reference statements)

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“…PEDV crosses the porcine intestinal mucosa to cause intestinal infection, and then results in an acute viral enteric disease, which means that PEDV must gain access to the tight junctions. As noted earlier, the alteration of tight junction proteins distribution might participate in virus entry, for example, occludin internalization contributes to PEDV entry [35]. In the experiments reported here, we demonstrated that DON could aggravate occludin internalization in PEDV-infected cells.…”

Section: Discussionsupporting

confidence: 82%

Deoxynivalenol Promotes Porcine Epidemic Diarrhea Virus Infection and Aggravates Gut Barrier Injury

Wang

et al. 2019

Preprint

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23Porcine epidemic diarrhea virus (PEDV) is a highly contagious pathogenic virus that 24 causes severe diarrhea and dehydration in pigs of all ages. Deoxynivalenol (DON), the 25 most abundant trichothecene in food and feed, causes vomit and diarrhea in animals 26 and human. However, whether DON exposure could affect PEDV infection remains 27 unknown. Herein, we investigated the impacts of DON on entry and replication of 28 PEDV, morbidity situation of piglets and the mechanisms involved. In vivo, twenty-29 seven piglets infected naturally with PEDV were randomly divided into three groups, 30 receiving the basal diet containing 0, 750 and 1500 μg/kg DON, respectively. We 31 observed significant increases in the diarrhea rates, the villous injury of jejunums and 32 the PEDV proliferation of duodenum, jejunum, ileum and mesenterium of piglets in 33 experimental groups compared with control. Additionally, the autophagosome-like 34 vesicles and the autophagy-related protein expressions were also increased in 35 experimental groups. In vitro, we observed that, approximately 2 hrs post-infection, 36 0.1, 0.5 and 1.0 μM DON promoted PEDV entry (P < 0.05) in IPEC-J2s and resulted 37 in tight junction protein occludin internalization. Knockdown of occludin and 38 CRISPR-Cas9-mediated knockout of LC3B indicated a vital role of autophagy-induced 39 occludin internalization in DON-promoted PEDV entry. We also observed that, 24 hrs 40 post-infection, a significant increase in PEDV replication after 0.1, 0.5 and 1.0 μM 41 DON treatment, along with the induction of a complete autophagy. Specifically, 42 deletion of LC3B indicated a crucial role of autophagy in DON-promoted PEDV . CC-BY 4.0 International license author/funder. It is made available under aThe copyright holder for this preprint (which was not peer-reviewed) is the . https://doi.org/10.1101/852608 doi: bioRxiv preprint 3 43 replication. Pretreatment with SB202190, a p38 signaling inhibitor, abolished the 44 induction of autophagy. Furthermore, downregulation of type I interferon revealed that 45 DON contributed PEDV to escape innate immune. Mechanistically, DON-caused 46 innate immune escape was related to the upregulation of LC3B, which further inhibited 47 STING phosphorylation. Taken together, DON could promote PEDV infection by 48 inducing occludin internalization and innate immune escape via triggering p38-49 mediated autophagy.50 Author summary 53 Porcine epidemic diarrhea (PED), a devastating enteric disease, leads to catastrophic 54 economic loss to the global pig industry. Its primary pathogen is the coronavirus PED 55 virus (PEDV). Growing evidence indicates that pathogen infection is not the only factor 56 of PED outbreaks, other non-infectious factors is also related to this disease. We 57 guessed some ubiquitous substances, such as deoxynivalenol (DON), that lead to pig 58 intestinal epithelial cell stress might encourage the progress and spread of PED. In the 59 present study, the weaning piglets infected naturally with PEDV and the IPEC-J2 cell 60 lin...

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Section: Discussionsupporting

confidence: 82%

Deoxynivalenol Promotes Porcine Epidemic Diarrhea Virus Infection and Aggravates Gut Barrier Injury

Wang

et al. 2019

Preprint

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“…qPCR reaction was performed using a SYBR Premix Ex Taq II Kit (Takara, Tokyo, Japan) using a Light Cycler 480 real-time PCR system (Roche Diagnostics, Indianapolis, IN, USA). For the Western blotting analysis [36], the cells were washed with PBS and lysed in RIPA lysis buffer on ice for 30 min. After SDS-PAGE electrophoresis, proteins were transferred onto polyvinylidene fluoride (PVDF) membrane via the semidry method and immunoblotted with the corresponding antibodies.…”

Section: Qrt-pcr and Western Blottingmentioning

confidence: 99%

PEDV enters cells through clathrin-, caveolae-, and lipid raft-mediated endocytosis and traffics via the endo-/lysosome pathway

Wang

She

et al. 2020

Vet Res

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With the emergence of highly pathogenic variant strains, porcine epidemic diarrhea virus (PEDV) has led to significant economic loss in the global swine industry. Many studies have described how coronaviruses enter cells, but information on PEDV invasion strategies remains insufficient. Given that the differences in gene sequences and pathogenicity between classical and mutant strains of PEDV may lead to diverse invasion mechanisms, this study focused on the cellular entry pathways and cellular transport of the PEDV GI and GII subtype strains in Vero cells and IPEC-J2 cells. We first characterized the kinetics of PEDV entry into cells and found that the highest invasion rate of PEDV was approximately 33% in the IPEC-J2 cells and approximately 100% in the Vero cells. To clarify the specific endocytic pathways, systematic research methods were used and showed that PEDV enters cells via the clathrin-and caveolae-mediated endocytosis pathways, in which dynamin II, clathrin heavy chain, Eps15, cholesterol, and caveolin-1 were indispensably involved. In addition, lipid raft extraction assay showed that PEDV can also enter cells through lipid raft-mediated endocytosis. To investigate the trafficking of internalized PEDV, we found that PEDV entry into cells relied on low pH and internalized virions reached lysosomes through the early endosome-late endosome-lysosome pathway. The results concretely revealed the entry mechanisms of PEDV and provided an insightful theoretical basis for the further understanding of PEDV pathogenesis and guidance for new targets of antiviral drugs.© The Author(s) 2020. This article is licensed under a Creative Commons Attribution 4.0 International License, which permits use, sharing, adaptation, distribution and reproduction in any medium or format, as long as you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons licence, and indicate if changes were made. The images or other third party material in this article are included in the article' s Creative Commons licence, unless indicated otherwise in a credit line to the material. If material is not included in the article'

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“…IFA. Immunofluorescence assays (IFA) were performed as described previously with slight modification (13). Briefly, after EGFR inhibitor treatments, IPEC-J2 and HEK293 cells were infected with PEDV.…”

Section: Cells and Viruses Ipec-j2 Cells (Porcine Small Intestine Epmentioning

confidence: 99%

“…acute destruction of intestinal villous enterocytes and villous atrophy in the jejunum and ileum (10,11). We and others have reported that the integrity of the intestinal epithelium is impaired, and the structural destruction and disorganization of tightjunction proteins in PEDV-infected cells are observed (12)(13)(14). Epidermal growth factor receptor (EGFR) is expressed in a wide spectrum of epithelial cells, and EGFR-dependent signaling is involved in the disassembly of epithelial tight junctions and epithelial barrier permeability alteration (15)(16)(17)(18)(19).…”

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confidence: 97%

Porcine Epidemic Diarrhea Virus-Induced Epidermal Growth Factor Receptor Activation Impairs the Antiviral Activity of Type I Interferon

Yang

Guo

et al. 2018

J Virol

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Porcine epidemic diarrhea virus (PEDV) causes acute and devastating enteric disease in suckling piglets and results in huge economic losses in the pig industry worldwide. To establish productive infection, viruses must first circumvent the host innate immune response. In this study, we found that PEDV infection stimulated epidermal growth factor receptor (EGFR) activation, which has been linked to not only anticancer therapeutics, but also antiviral signaling. Therefore, we determined whether EGFR activation affected PEDV infection by using an activator or overexpression assay. The data showed that EGFR activation enhanced virus replication in both cases. We also found that specific inhibition of EGFR by either inhibitors or small interfering RNA (siRNA) led to a decrease in virus yields. Further analysis revealed that inhibition of EGFR produced augmentation of type I interferon genes. We next observed that the EGFR downstream cascade STAT3 was also activated upon PEDV infection. Similar to the case of EGFR, specific inhibition of STAT3 by either inhibitor or siRNA increased the antiviral activity of interferon and resulted in decreased PEDV RNA levels, and vice versa. The data on STAT3 depletion in combination with EGFR activation suggest that the attenuation of antiviral activity by EGFR activation requires activation of the STAT3 signaling pathway. Taken together, these data demonstrate that PEDV-induced EGFR activation serves as a negative regulator of the type I interferon response and provides a novel therapeutic target for virus infection. EGFR is a transmembrane tyrosine receptor that mediates various cellular events, as well as several types of human cancers. In this study, we investigated for the first time the role of EGFR in PEDV infection. We observed that PEDV infection induced EGFR activation. The role of EGFR activation is to impair the antiviral activity of type I interferon, which requires the involvement of the EGFR downstream signaling cascade STAT3. Our findings reveal a new mechanism evolved by PEDV to circumvent the host antiviral response, which might serve as a therapeutic target against virus infection.

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Tight Junction Protein Occludin Is a Porcine Epidemic Diarrhea Virus Entry Factor

Cited by 68 publications

References 67 publications

Deoxynivalenol Promotes Porcine Epidemic Diarrhea Virus Infection and Aggravates Gut Barrier Injury

Deoxynivalenol Promotes Porcine Epidemic Diarrhea Virus Infection and Aggravates Gut Barrier Injury

PEDV enters cells through clathrin-, caveolae-, and lipid raft-mediated endocytosis and traffics via the endo-/lysosome pathway

Porcine Epidemic Diarrhea Virus-Induced Epidermal Growth Factor Receptor Activation Impairs the Antiviral Activity of Type I Interferon

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