scholarly journals Porcine Epidemic Diarrhea Virus (PEDV) ORF3 Interactome Reveals Inhibition of Virus Replication by Cellular VPS36 Protein

Viruses ◽  
2019 ◽  
Vol 11 (4) ◽  
pp. 382 ◽  
Author(s):  
Challika Kaewborisuth ◽  
Yodying Yingchutrakul ◽  
Sittiruk Roytrakul ◽  
Anan Jongkaewwattana
Keyword(s):  
Virus Replication ◽  
Porcine Epidemic Diarrhea Virus ◽  
Interacting Proteins ◽  
Gene Ontology Analysis ◽  
Orf3 Gene ◽  
Orf3 Protein ◽  
Diarrhea Virus ◽  
Host Interaction ◽  
Porcine Epidemic Diarrhea ◽  
Vacuolar Protein

The accessory protein ORF3 of porcine epidemic diarrhea virus (PEDV) has been proposed to play a key role in virus replication. However, our understanding of its function regarding virus and host interaction is still limited. In this study, we employed immunoprecipitation and mass spectrometry to screen for cellular interacting partners of ORF3. Gene ontology analysis of the host interactome highlighted the involvement of ORF3 in endosomal and immune signaling pathways. Among the identified ORF3-interacting proteins, the vacuolar protein-sorting-associated protein 36 (VPS36) was assessed for its role in PEDV replication. VPS36 was found to interact with ORF3 regardless of its GLUE domain. As a result of VPS36–ORF3 interaction, PEDV replication was substantially suppressed in cells overexpressing VPS36. Interestingly, the ORF3 protein expression was diminished in VPS36-overexpressing cells, an effect that could not be restored by treatment of lysosomal inhibitors. In addition, disruption of endogenously-expressed VPS36 by siRNA could partially augment PEDV replication. Taken together, our study provides mechanistic insights into the contribution of ORF3 in PEDV replication.

scholarly journals Porcine Epidemic Diarrhea Virus (PEDV) ORF3 Enhances Viral Proliferation by Inhibiting Apoptosis of Infected Cells

Viruses ◽  
2020 ◽  
Vol 12 (2) ◽  
pp. 214 ◽  
Author(s):  
Fusheng Si ◽  
Xiaoxia Hu ◽  
Chenyang Wang ◽  
Bingqing Chen ◽  
Ruiyang Wang ◽  
...  
Keyword(s):  
Porcine Epidemic Diarrhea Virus ◽  
Underlying Mechanism ◽  
Vero Cells ◽  
Orf3 Gene ◽  
Accessory Gene ◽  
Orf3 Protein ◽  
Diarrhea Virus ◽  
Viral Virulence ◽  
Porcine Epidemic Diarrhea

The genomes of coronaviruses carry accessory genes known to be associated with viral virulence. The single accessory gene of porcine epidemic diarrhea virus (PEDV), ORF3, is dispensable for virus replication in vitro, while viral mutants carrying ORF3 truncations exhibit an attenuated phenotype of which the underlying mechanism is unknown. Here, we studied the effect of ORF3 deletion on the proliferation of PEDV in Vero cells. To this end, four recombinant porcine epidemic diarrhea viruses (PEDVs) were rescued using targeted RNA recombination, three carrying the full-length ORF3 gene from different PEDV strains, and one from which the ORF3 gene had been deleted entirely. Our results showed that PEDVs with intact or naturally truncated ORF3 replicated to significantly higher titers than PEDV without an ORF3. Further characterization revealed that the extent of apoptosis induced by PEDV infection was significantly lower with the viruses carrying an intact or C-terminally truncated ORF3 than with the virus lacking ORF3, indicating that the ORF3 protein as well as its truncated form interfered with the apoptosis process. Collectively, we conclude that PEDV ORF3 protein promotes virus proliferation by inhibiting cell apoptosis caused by virus infection. Our findings provide important insight into the role of ORF3 protein in the pathogenicity of PEDV.

scholarly journals Emergence and evolution of highly pathogenic porcine epidemic diarrhea virus by natural recombination of a low pathogenic vaccine isolate and a highly pathogenic strain in the spike gene

2020 ◽  
Vol 6 (2) ◽  
Author(s):  
Huinan Wang ◽  
Libo Zhang ◽  
Yuanbin Shang ◽  
Rongrong Tan ◽  
Mingxiang Ji ◽  
...  
Keyword(s):  
Porcine Epidemic Diarrhea Virus ◽  
Intestinal Epithelial ◽  
Orf3 Gene ◽  
Highly Pathogenic ◽  
Spike Gene ◽  
Diarrhea Virus ◽  
Attenuated Vaccine ◽  
New Variant ◽  
Porcine Epidemic Diarrhea ◽  
Natural Recombination

Abstract Outbreaks of a new variant of porcine epidemic diarrhea virus (PEDV) at the end of 2010 have raised interest in the mutation and recombination of PEDV. A PEDV strain (CN/Liaoning25/2018) isolated from a clinical outbreak of piglet diarrhea contained a 49-bp deletion in the ORF3 gene. This deletion is considered a genetic characteristic of low pathogenic attenuated vaccine strains. However, CN/Liaoning25/2018 was highly pathogenic. Complete genome sequencing, identity analysis, phylogenetic tree construction, and recombination analysis showed that this virus was a recombinant strain containing the Spike (S) gene from the highly pathogenic CN/GDZQ/2014 strain and the remaining genomic regions from the low pathogenic vaccine isolate SQ2014. Histopathology and immunohistochemistry results confirmed that this strain was highly pathogenic and indicated that intestinal epithelial cell vacuolation was positively correlated with the intensity and density of PEDV antigens. A new natural recombination model for PEDV was identified. Our results suggest that new highly pathogenic recombinant strains in the field may be generated by recombination between low pathogenic attenuated live PEDV vaccines and pathogenic circulating PEDV strains. Our findings also highlight that the 49-bp deletion of the ORF3 gene in low pathogenic attenuated vaccine strains will no longer be a reliable standard to differentiate the classical vaccine attenuated from the field strains.

scholarly journals Porcine Epidemic Diarrhea Virus ORF3 Protein Is Transported through the Exocytic Pathway

2020 ◽  
Vol 94 (17) ◽  
Author(s):  
Fusheng Si ◽  
Bingqing Chen ◽  
Xiaoxia Hu ◽  
Ruisong Yu ◽  
Shijuan Dong ◽  
...  
Keyword(s):  
Ion Channel ◽  
Porcine Epidemic Diarrhea Virus ◽  
Intracellular Trafficking ◽  
Expression Vectors ◽  
Orf3 Protein ◽  
Diarrhea Virus ◽  
Terminal Domain ◽  
Infected Cells ◽  
Porcine Epidemic Diarrhea ◽  
Exocytic Pathway

ABSTRACT Accessory genes occurring between the S and E genes of coronaviruses have been studied quite intensively during the last decades. In porcine epidemic diarrhea virus (PEDV), the only gene at this location, ORF3, encodes a 224-residue membrane protein shown to exhibit ion channel activity and to enhance virus production. However, little is known about its intracellular trafficking or about its function during PEDV infection. In this study, two recombinant PEDVs were rescued by targeted RNA recombination, one carrying the full-length ORF3 gene and one from which the gene had been deleted entirely. These viruses as well as a PEDV encoding a naturally truncated ORF3 protein were employed to study the ORF3 protein’s subcellular trafficking. In addition, ORF3 expression vectors were constructed to study the protein’s independent transport. Our results show that the ORF3 protein uses the exocytic pathway to move to and accumulate in the Golgi area of the cell similarly in infected and transfected cells. Like the S protein, but unlike the other structural proteins M and N, the ORF3 protein was additionally observed at the surface of PEDV-infected cells. In addition, the C-terminally truncated ORF3 protein entered the exocytic pathway but it was unable to leave the endoplasmic reticulum (ER) and ER-to-Golgi intermediate compartment (ERGIC). Consistently, a YxxØ motif essential for ER exit was identified in the C-terminal domain. Finally, despite the use of sensitive antibodies and assays no ORF3 protein could be detected in highly purified PEDV particles, indicating that the protein is not a structural virion component. IMPORTANCE Coronaviruses typically express several accessory proteins. They vary in number and nature, and only one is conserved among most of the coronaviruses, pointing at an important biological function for this protein. PEDV is peculiar in that it expresses just this one accessory protein, termed the ORF3 protein. While its analogs in other coronaviruses have been studied to different extents, and these studies have indicated that they share an ion channel property, little is still known about the features and functions of the PEDV ORF3 protein except for its association with virulence. In this investigation, we studied the intracellular trafficking of the ORF3 protein both in infected cells and when expressed independently. In addition, we analyzed the effects of mutations in five sorting motifs in its C-terminal domain and investigated whether the protein, found to follow the same exocytic route by which the viral structural membrane proteins travel, is also incorporated into virions.

scholarly journals Porcine epidemic diarrhea virus ORF3 gene prolongs S-phase, facilitates formation of vesicles and promotes the proliferation of attenuated PEDV

Virus Genes ◽  
2015 ◽  
Vol 51 (3) ◽  
pp. 385-392 ◽  
Author(s):  
Shiyi Ye ◽  
Zhonghua Li ◽  
Fangzhou Chen ◽  
Wentao Li ◽  
Xiaozhen Guo ◽  
...  
Keyword(s):  
Porcine Epidemic Diarrhea Virus ◽  
S Phase ◽  
Orf3 Gene ◽  
Diarrhea Virus ◽  
Porcine Epidemic Diarrhea
Sign in / Sign up

Export Citation Format

Share Document