Hepatitis C virus core protein transactivates the inducible nitric oxide synthase promoter via NF-kappaB activation.
Intrahepatic levels of the inducible nitric oxide synthase (iNOS) are increased in chronic hepatitis C patients. As iNOS gene promoter contains Nuclear Factor (NF)-kappaB binding sites and hepatitis C virus (HCV) core protein activates NF-kappaB, the aim of this work was to study if HCV core protein transactivates iNOS promoter through NF-kappaB activation. iNOS mRNA and protein were determined by RT-PCR and western blot in HepG2 cells. The effect of HCV core protein on iNOS promoter was assayed by cotransfecting HepG2 cells with the core protein expression plasmid pHCV-Co and p1iNOS-CAT or p2iNOS-CAT plasmids. Formation of NF-kappaB-DNA complexes was determined by electrophoretic mobility shift assay. Transfection of HepG2 cells with pHCV-Co plasmid results in an increase in iNOS mRNA and protein levels. Cotransfection with pHCV-Co and p1iNOS-CAT or p2iNOS-CAT plasmids results in a transactivation of iNOS promoter, the presence of the proximal NF-kappaB binding site in the promoter being sufficient for the transactivation. Furthermore, the HCV core protein increases the formation of complexes between NF-kappaB and its binding sequence in the iNOS promoter. The expression of the NF-kappaB inhibitor IKB reverts the effect of the HCV core protein on the iNOS promoter. In conclusion, HCV core protein transactivates iNOS gene promoter through NF-kappaB activation.
de Lucas
Susana
S
Fundación para el Estudio de las Hepatitis Virales, C/Guzman el Bueno no. 72, 28015 Madrid, Spain.
Bartolomé
Javier
J
Amaro
María José
MJ
Carreño
Vicente
V
D045744:Cell Line, Tumor; D015500:Chloramphenicol O-Acetyltransferase / Q000378:metabolism; D017930:Genes, Reporter; D016174:Hepacivirus / Q000472:pathogenicity*; D006801:Humans; D008156:Luciferases / Q000378:metabolism; D016328:NF-kappa B / Q000378:metabolism*; D019001:Nitric Oxide Synthase / Q000235:genetics* / Q000378:metabolism; D052247:Nitric Oxide Synthase Type II; D011401:Promoter Regions, Genetic; D011485:Protein Binding; D012333:RNA, Messenger / Q000032:analysis; D015533:Transcriptional Activation; D014162:Transfection; D014758:Viral Core Proteins / Q000502:physiology*
D016328:NF-kappa B; D012333:RNA, Messenger; D014758:Viral Core Proteins; D008156:Luciferases; C496319:NOS2 protein, human; D019001:Nitric Oxide Synthase; D052247:Nitric Oxide Synthase Type II; D015500:Chloramphenicol O-Acetyltransferase