title: Hepatitis C virus NS5A promotes insulin resistance through IRS-1 serine phosphorylation and increased gluconeogenesis.
abstract: OBJECTIVE
  To investigate the mechanisms of insulin resistance in human hepatoma cells expressing hepatitis C virus (HCV) nonstructural protein 5A (NS5A).
  
  
  METHODS
  The human hepatoma cell lines, Huh7 and Huh7.5, were infected with HCV or transiently-transfected with a vector expressing HCV NS5A. The effect of HCV NS5A on the status of the critical players involved in insulin signaling was analyzed using real-time quantitative polymerase chain reaction and Western blot assays. Data were analyzed using Graph Pad Prism version 5.0.
  
  
  RESULTS
  To investigate the effect of insulin treatment on the players involved in insulin signaling pathway, we analyzed the status of insulin receptor substrate-1 (IRS-1) phosphorylation in HCV infected cells or Huh7.5 cells transfected with an HCV NS5A expression vector. Our results indicated that there was an increased phosphorylation of IRS-1 (Ser(307)) in HCV infected or NS5A transfected Huh7.5 cells compared to their respective controls. Furthermore, an increased phosphorylation of Akt (Ser(473)) was observed in HCV infected and NS5A transfected cells compared to their mock infected cells. In contrast, we observed decreased phosphorylation of Akt Thr308 phosphorylation in HCV NS5A transfected cells. These results suggest that Huh7.5 cells either infected with HCV or ectopically expressing HCV NS5A alone have the potential to induce insulin resistance by the phosphorylation of IRS-1 at serine residue (Ser(307)) followed by decreased phosphorylation of Akt Thr(308), Fox01 Ser(256) and GSK3β Ser(9), the downstream players of the insulin signaling pathway. Furthermore, increased expression of PECK and glucose-6-phosphatase, the molecules involved in gluconeogenesis, in HCV NS5A transfected cells was observed.
  
  
  CONCLUSIONS
  Taken together, our results suggest the role of HCV NS5A in the induction of insulin resistance by modulating various cellular targets involved in the insulin signaling pathway.
authors.affiliation: Fahed Parvaiz, Sobia Manzoor, Muhammad Imran, Atta-ur-Rahman School of Applied Biosciences, National University of Sciences and Technology, Islamabad 44000, Pakistan.
authors.forename: Fahed
authors.initials: F
authors.lastname: Parvaiz
authors.affiliation: Fahed Parvaiz, Sobia Manzoor, Muhammad Imran, Atta-ur-Rahman School of Applied Biosciences, National University of Sciences and Technology, Islamabad 44000, Pakistan.
authors.forename: Sobia
authors.initials: S
authors.lastname: Manzoor
authors.affiliation: Fahed Parvaiz, Sobia Manzoor, Muhammad Imran, Atta-ur-Rahman School of Applied Biosciences, National University of Sciences and Technology, Islamabad 44000, Pakistan.
authors.forename: Jawed
authors.initials: J
authors.lastname: Iqbal
authors.affiliation: Fahed Parvaiz, Sobia Manzoor, Muhammad Imran, Atta-ur-Rahman School of Applied Biosciences, National University of Sciences and Technology, Islamabad 44000, Pakistan.
authors.forename: Mehuli
authors.initials: M
authors.lastname: Sarkar-Dutta
authors.affiliation: Fahed Parvaiz, Sobia Manzoor, Muhammad Imran, Atta-ur-Rahman School of Applied Biosciences, National University of Sciences and Technology, Islamabad 44000, Pakistan.
authors.forename: Muhammad
authors.initials: M
authors.lastname: Imran
authors.affiliation: Fahed Parvaiz, Sobia Manzoor, Muhammad Imran, Atta-ur-Rahman School of Applied Biosciences, National University of Sciences and Technology, Islamabad 44000, Pakistan.
authors.forename: Gulam
authors.initials: G
authors.lastname: Waris
mesh_terms: D006528:Carcinoma, Hepatocellular / Q000235:genetics / Q000378:metabolism / Q000821:virology*; D045744:Cell Line, Tumor; D000071161:Forkhead Box Protein O1; D051858:Forkhead Transcription Factors / Q000378:metabolism; D005943:Gluconeogenesis; D038362:Glycogen Synthase Kinase 3 / Q000378:metabolism; D000071679:Glycogen Synthase Kinase 3 beta; D016174:Hepacivirus / Q000235:genetics / Q000378:metabolism*; D054884:Host-Pathogen Interactions; D006801:Humans; D055504:Insulin Receptor Substrate Proteins / Q000378:metabolism*; D007333:Insulin Resistance; D008113:Liver Neoplasms / Q000235:genetics / Q000378:metabolism / Q000821:virology*; D010766:Phosphorylation; D051057:Proto-Oncogene Proteins c-akt / Q000378:metabolism; D012694:Serine; D015398:Signal Transduction; D014162:Transfection; D017361:Viral Nonstructural Proteins / Q000235:genetics / Q000378:metabolism*; D012324:RNA-Dependent RNA Polymerase
chemical_list: D000071161:Forkhead Box Protein O1; D051858:Forkhead Transcription Factors; D038362:Glycogen Synthase Kinase 3; D000071679:Glycogen Synthase Kinase 3 beta; D055504:Insulin Receptor Substrate Proteins; D051057:Proto-Oncogene Proteins c-akt; D012694:Serine; D017361:Viral Nonstructural Proteins; C085126:FOXO1 protein, human; C526292:IRS1 protein, human; C087036:NS-5 protein, hepatitis C virus; D012324:RNA-Dependent RNA Polymerase
keywords: Forkhead box protein 01; Gluconeogenesis; Glycogen synthase kinase-beta; Hepatitis C virus nonstructural protein 5A; Insulin resistance