title: Cap-dependent and hepatitis C virus internal ribosome entry site-mediated translation are modulated by phosphorylation of eIF2alpha under oxidative stress.
abstract: Chronic hepatitis C is often associated with oxidative stress. Hepatitis C virus (HCV) utilizes an internal ribosome entry site (IRES) element for translation, in contrast to cap-dependent translation of the majority of cellular proteins. To understand how virus translation is modulated under oxidative stress, HCV IRES-mediated translation was compared with cap-dependent translation using a bicistronic reporter construct and hydrogen peroxide (H2O2) as a stress inducer. In H2O2-sensitive HeLa cells, H2O2 repressed translation in a time- and dose-dependent manner, concomitant with the kinetics of eIF2alpha phosphorylation. A phosphomimetic of eIF2alpha, which mimics the structure of the phosphorylated eIF2alpha, was sufficient to repress translation in the absence of H2O2. In H2O2-resistant HepG2 cells, H2O2 activated both HCV IRES-mediated and cap-dependent translation, associated with an increased level of phospho-eIF2alpha. It was postulated that H2O2 might stimulate translation in HepG2 cells via an eIF2alpha-independent mechanism, whereas the simultaneous phosphorylation of eIF2alpha repressed part of the translational activities. Indeed, the translational repression was released in the presence of a non-phosphorylatable mutant, eIF2alpha-SA, resulting in further enhancement of both translational activities after exposure to H2O2. In HuH7 cells, which exhibited an intermediate level of sensitivity towards H2O2, both HCV IRES-mediated and cap-dependent translational activities were upregulated after treatment with various doses of H2O2, but the highest level of induction was achieved with a low level of H2O2, which may represent the physiological level of H2O2. At this level, the HCV IRES-mediated translation was preferentially upregulated compared with cap-dependent translation.
authors.affiliation: Faculty of Life Sciences, The University of Manchester, Jackson's Mill, PO Box 88, Sackville Street, Manchester M60 1QD, UK.
authors.forename: Paul R
authors.initials: PR
authors.lastname: MacCallum
authors.affiliation: Faculty of Life Sciences, The University of Manchester, Jackson's Mill, PO Box 88, Sackville Street, Manchester M60 1QD, UK.
authors.forename: Samantha C
authors.initials: SC
authors.lastname: Jack
authors.affiliation: Faculty of Life Sciences, The University of Manchester, Jackson's Mill, PO Box 88, Sackville Street, Manchester M60 1QD, UK.
authors.forename: Philip A
authors.initials: PA
authors.lastname: Egan
authors.affiliation: Faculty of Life Sciences, The University of Manchester, Jackson's Mill, PO Box 88, Sackville Street, Manchester M60 1QD, UK.
authors.forename: Benjamin T
authors.initials: BT
authors.lastname: McDermott
authors.affiliation: Centre for Biomolecular Sciences, School of Biology, University of St Andrews, North Haugh, St Andrews, Fife KY16 9ST, UK.
authors.forename: Richard M
authors.initials: RM
authors.lastname: Elliott
authors.affiliation: Manchester Royal Infirmary, Oxford Road, Manchester M13 9WL, UK.|Faculty of Life Sciences, The University of Manchester, Jackson's Mill, PO Box 88, Sackville Street, Manchester M60 1QD, UK.
authors.forename: Shiu-Wan
authors.initials: SW
authors.lastname: Chan
mesh_terms: D002460:Cell Line; D004305:Dose-Response Relationship, Drug; D006367:HeLa Cells; D016174:Hepacivirus / Q000235:genetics*; D019698:Hepatitis C, Chronic / Q000378:metabolism* / Q000821:virology*; D006801:Humans; D006861:Hydrogen Peroxide / Q000494:pharmacology; D009154:Mutation; D016877:Oxidants / Q000494:pharmacology; D018384:Oxidative Stress; D010766:Phosphorylation; D014176:Protein Biosynthesis / Q000187:drug effects; D019892:eIF-2 Kinase / Q000235:genetics / Q000378:metabolism*
chemical_list: D016877:Oxidants; D006861:Hydrogen Peroxide; D019892:eIF-2 Kinase