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Open reading frame 3 of genotype 1 hepatitis E virus inhibits nuclear factor-κappa B signaling induced by tumor necrosis factor-α in human A549 lung epithelial cells.
- Source :
-
PloS one [PLoS One] 2014 Jun 24; Vol. 9 (6), pp. e100787. Date of Electronic Publication: 2014 Jun 24 (Print Publication: 2014). - Publication Year :
- 2014
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Abstract
- Hepatitis E virus (HEV) is one of the primary causative agents of acute hepatitis, and represents a major cause of severe public health problems in developing countries. The pathogenesis of HEV is not well characterized, however, primarily due to the lack of well-defined cell and animal models. Here, we investigated the effects of genotype 1 HEV open reading frame 3 (ORF3) on TNF-α-induced nucleus factor-κappa B (NF-κB) signaling. Human lung epithelial cells (A549) were transiently transfected with ORF3 containing plasmids. These cells were then stimulated with TNF-α and the nucleus translocation of the p65 NF-κB subunit was assessed using western blot and laser confocal microscopy. DNA-binding activity of p65 was also examined using electrophoretic mobility shift assay (EMSA), and the suppression of NF-κB target genes were detected using real-time RT-PCR and ELISA. These results enabled us to identify the decreased phosphorylation levels of IKBα. We focused on the gene of negative regulation of NF-κB, represented by TNF-α-induced protein 3 (TNFAIP3, also known as A20). Reducing the levels of A20 with siRNAs significantly enhances luciferase activation of NF-κB. Furthermore, HEV ORF3 regulated A20 primarily via activating transcription factor 6 (ATF6), involved in unfolded protein response (UPR), resulting in the degradation or inactivation of the receptor interacting protein 1 (RIP1), a major upstream activator of IKB kinase compounds (IKKs). Consequently, the phosphorylation of IKBα and the nucleus translocation of p65 are blocked, which contributes to diminished NF-κB DNA-binding activation and NF-κB-dependent gene expression. The findings suggest that genotype 1 HEV, through ORF3, may transiently activate NF-κB through UPR in early stage, and subsequently inhibit TNF-α-induced NF-κB signaling in late phase so as to create a favorable virus replication environment.
- Subjects :
- Cell Line
DNA-Binding Proteins metabolism
Enzyme Activation
Epithelial Cells metabolism
Epithelial Cells virology
Humans
Intracellular Signaling Peptides and Proteins metabolism
Lung
Nuclear Proteins metabolism
Phosphorylation drug effects
Tumor Necrosis Factor alpha-Induced Protein 3
Tumor Necrosis Factor-alpha pharmacology
Unfolded Protein Response
Viral Proteins genetics
Genotype
Hepatitis E metabolism
Hepatitis E virology
Hepatitis E virus genetics
NF-kappa B metabolism
Signal Transduction drug effects
Viral Proteins metabolism
Subjects
Details
- Language :
- English
- ISSN :
- 1932-6203
- Volume :
- 9
- Issue :
- 6
- Database :
- MEDLINE
- Journal :
- PloS one
- Publication Type :
- Academic Journal
- Accession number :
- 24959724
- Full Text :
- https://doi.org/10.1371/journal.pone.0100787