Subcellular localizations of the hepatitis C virus alternate reading frame proteins.
Alternate reading frame proteins (ARFPs) resulting either from frameshifting, from transcriptional slippage or from internal initiation in the +1 open reading frame (ORF) of hepatitis C virus (HCV) core protein coding sequence have been described in vitro. As an approach to study the roles of these proteins, we investigate the subcellular localization of ARFPs fused with the green fluorescent protein (GFP) either at their N- or C-terminus. Most GFP fusion products have a diffuse localization, as revealed by confocal microscopy. One GFP chimeric protein, arising from internal initiation at codon 26 in the +1 ORF (ARFP(26-161)), is specifically targeted to mitochondria. Mitochondrial localization was confirmed by immunoblot with an anti-ARFP antibody of a mitochondria-enriched cellular fraction. Mitochondrial targeting of ARFP(26-161) mostly involved the N-terminal portion of the protein as revealed by the cellular localization of truncated mutants. Interestingly, ARFP(26-161) from both genotypes 1a and 1b, but not the protein from the genotype 2a JFH1 infectious sequence, exhibit mitochondrial localization. These results are the first concerning the cellular localization and the role of this HCV ARFP; they may serve as a platform for further studies on its mitochondrial effects and their role in the virus life cycle and pathogenesis.
Ratinier
Maxime
M
Institut de Biologie et Chimie des Protéines, UMR 5086 CNRS-Université de Lyon, IFR 128 Biosciences, 7 passage du Vercors, 69367 Lyon cedex 07, France.
Boulant
Steeve
S
Crussard
Steve
S
McLauchlan
John
J
Lavergne
Jean-Pierre
JP
D002460:Cell Line; D015967:Gene Expression Regulation, Viral / Q000502:physiology; D049452:Green Fluorescent Proteins / Q000378:metabolism; D016174:Hepacivirus / Q000235:genetics / Q000502:physiology*; D006801:Humans; D018613:Microscopy, Confocal; D008928:Mitochondria / Q000378:metabolism* / Q000821:virology; D016366:Open Reading Frames / Q000235:genetics* / Q000502:physiology; D014162:Transfection; D014758:Viral Core Proteins / Q000502:physiology*
D014758:Viral Core Proteins; D049452:Green Fluorescent Proteins