Abstract
Swine hepatitis E (swine HE) is a new type of zoonotic infectious disease caused by the swine hepatitis E virus (swine HEV). Open reading frame 3 (ORF3) is an important virulent protein of swine HEV, but its function still is mainly unclear. In this study, we generated adenoviruses ADV4-ORF3 and ADV4 negative control (ADV4-NC), which successfully mediated overexpression of enhanced green fluorescent protein (EGFP)-ORF3 and EGFP, respectively, in HepG2 cells. High-throughput sequencing was used to screen for differentially expressed long non-coding RNAs (lncRNAs) and messenger RNAs (mRNAs). The cis-target genes of lncRNAs were predicted, functional enrichment (Gene Ontology [GO] and Kyoto Encyclopedia of Genes and Genomes [KEGG]) was performed, and 12 lncRNAs with statistically significant different expressions (p ≤ 0.05 and q ≤ 1) were selected for further quantitative real-time reverse transcription (qRT-PCR) validation. In HepG2 cells, we identified 62 significantly differentially expressed genes (DEGs) (6,564 transcripts) and 319 lncRNAs (124 known lncRNAs and 195 novel lncRNAs) that were affected by ORF3, which were involved in systemic lupus erythematosus, Staphylococcus aureus infection, signaling pathways pluripotency regulation of stem cells, the peroxisome proliferator-activated receptor (PPAR) signaling pathway, and platinum drug resistance pathways. Cis-target gene prediction identified 45 lncRNAs corresponding to candidate mRNAs, among which eight were validated by qRT-PCR: LINC02476 (two transcripts), RAP2C-AS1, AC016526, AL139099, and ZNF337-AS1 (3 transcripts). Our results revealed that the lncRNA profile in host cells affected by ORF3, swine HEV ORF3, might affect the pentose and glucuronate interconversions and mediate the formation of obstructive jaundice by influencing bile secretion, which will help to determine the function of ORF3 and the infection mechanism and treatment of swine HE.
Highlights
Hepatitis E (HE) caused by the HE virus (HEV) is a new type of zoonotic infectious disease
HepG2 cells were infected with the high-titer recombinant adenovirus of ADV4 negative control (ADV4-NC) and ADV4-Open reading frame 3 (ORF3) for 24 h
The results showed that enhanced green fluorescent protein (EGFP) and the EGFP-ORF3 fusion protein were successfully expressed in HepG2 cells (Figure 1A). Quantitative reverse transcription PCR (qRT-PCR) was used to detect the relative expression of the ORF3 RNA, with GAPDH used as the internal control
Summary
Hepatitis E (HE) caused by the HE virus (HEV) is a new type of zoonotic infectious disease. HEV is mainly transmitted through the fecal–oral route, can cause acute hepatitis in humans, and is prevalent worldwide [1,2,3]. About 20 million people are infected with HEV every year [4]. In addition to the clinical symptoms of hepatitis, it causes nerve, blood, and kidney diseases; neuralgia muscular atrophy; encephalitis; myelitis; and other diseases [5]. Outbreaks of swine HE have appeared globally and have become a public health issue in various countries [7, 8]. The host and route of transmission of different genotypes are completely different, and the symptoms after infection are different. Its genome comprises about 7.6 kb, with a poly A tail structure at the 3′ end and three overlapping open reading frames (ORFs): ORF1, ORF2, and ORF3 [10, 11]
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