Translational insensitivity to potent activation of PKR by HCV IRES RNA.
Translation of hepatitis C virus (HCV) is initiated at an internal ribosome entry site (IRES) located at the 5'end of its RNA genome. The HCV IRES is highly structured and greater than 50% of its nucleotides form based-paired helices. We report here that the HCV IRES is an activator of PKR, an interferon-induced enzyme that participates in host cell defense against viral infection. Binding of HCV IRES RNA to PKR leads to a greatly increased (20-fold) rate and level (4.5-fold) of PKR autophosphorylation compared to previously studied dsRNA activators. We have mapped the domains in the IRES required for PKR activation to domains III-IV and demonstrate that the N-terminal double-stranded RNA binding domains of PKR bind to the IRES in a similar manner to other RNA activators. Addition of HCV IRES RNA inhibits cap-dependent translation in lysates via phosphorylation of PKR and eIF2alpha. However, HCV IRES-mediated translation is not inhibited by the phosphorylation of PKR and eIF2alpha. The results presented here suggest that hydrolysis of GTP by eIF2 is not an essential step in IRES-mediated translation. Thus, HCV can use structured RNAs to its advantage in translation, while avoiding the deleterious effects of PKR activation.
Shimoike
Takashi
T
Department of Virology II, National Institute of Infectious Diseases, Musashi-murayama, Tokyo 208-0011, Japan.
McKenna
Sean A
SA
Lindhout
Darrin A
DA
Puglisi
Joseph D
JD
D001483:Base Sequence; D015852:Eukaryotic Initiation Factor-2 / Q000378:metabolism; D016174:Hepacivirus / Q000276:immunology* / Q000502:physiology*; D008958:Models, Molecular; D008969:Molecular Sequence Data; D009690:Nucleic Acid Conformation; D010766:Phosphorylation; D011485:Protein Binding; D014176:Protein Biosynthesis; D012367:RNA, Viral / Q000378:metabolism*; D019892:eIF-2 Kinase / Q000378:metabolism*
D015852:Eukaryotic Initiation Factor-2; D012367:RNA, Viral; D019892:eIF-2 Kinase