title: RNA-seq based transcriptome analysis of hepatitis E virus (HEV) and hepatitis B virus (HBV) replicon transfected Huh-7 cells.
abstract: Pathogenesis of hepatitis B virus (HBV) and hepatitis E virus (HEV) infection is as varied as they appear similar; while HBV causes an acute and/or chronic liver disease and hepatocellular carcinoma, HEV mostly causes an acute self-limiting disease. In both infections, host responses are crucial in disease establishment and/or virus clearance. In the wake of worsening prognosis described during HEV super-infection over chronic HBV hepatitis, we investigated the host responses by studying alterations in gene expression in liver cells (Huh-7 cell line) by transfection with HEV replicon only (HEV-only), HBV replicon only (HBV-only) and both HBV and HEV replicons (HBV+HEV). Virus replication was validated by strand-specific real-time RT-PCR for HEV and HBsAg ELISA of the culture supernatants for HBV. Indirect immunofluorescence for the respective viral proteins confirmed infection. Transcription profiling was carried out by RNA Sequencing (RNA-Seq) analysis of the poly-A enriched RNA from the transfected cells. Averages of 600 million bases within 5.6 million reads were sequenced in each sample and ∼15,800 genes were mapped with at least one or more reads. A total of 461 genes in HBV+HEV, 408 in HBV-only and 306 in HEV-only groups were differentially expressed as compared to mock transfection control by two folds (p<0.05) or more. Majority of the significant genes with altered expression clustered into immune-associated, signal transduction, and metabolic process categories. Differential gene expression of functionally important genes in these categories was also validated by real-time RT-PCR based relative gene-expression analysis. To our knowledge, this is the first report of in vitro replicon transfected RNA-Seq based transcriptome analysis to understand the host responses against HEV and HBV.
authors: Department of Pathology, All India Institute of Medical Sciences, Ansari Nagar, New Delhi, India.
authors: Neetu
authors: N
authors: Jagya
authors: Department of Pathology, All India Institute of Medical Sciences, Ansari Nagar, New Delhi, India.
authors: Satya Pavan Kumar
authors: SP
authors: Varma
authors: Department of Pathology, All India Institute of Medical Sciences, Ansari Nagar, New Delhi, India.
authors: Deepshi
authors: D
authors: Thakral
authors: Department of Pathology, All India Institute of Medical Sciences, Ansari Nagar, New Delhi, India.
authors: Prashant
authors: P
authors: Joshi
authors: Department of Pathology, All India Institute of Medical Sciences, Ansari Nagar, New Delhi, India.
authors: Hemlata
authors: H
authors: Durgapal
authors: Department of Pathology, All India Institute of Medical Sciences, Ansari Nagar, New Delhi, India.
authors: Subrat Kumar
authors: SK
authors: Panda
mesh_terms: D002460:Cell Line; D060085:Coinfection / Q000235:genetics / Q000378:metabolism* / Q000473:pathology; D020869:Gene Expression Profiling; D015967:Gene Expression Regulation, Viral; D006515:Hepatitis B virus / Q000235:genetics / Q000378:metabolism*; D019694:Hepatitis B, Chronic / Q000235:genetics / Q000378:metabolism* / Q000473:pathology; D016751:Hepatitis E / Q000235:genetics / Q000378:metabolism* / Q000473:pathology; D016752:Hepatitis E virus / Q000235:genetics / Q000378:metabolism*; D006801:Humans; D012367:RNA, Viral / Q000096:biosynthesis; D012093:Replicon; D059467:Transcriptome; D014162:Transfection
chemical_list: D012367:RNA, Viral