Promotion of viral internal ribosomal entry site-mediated translation under amino acid starvation.
Cap-dependent and internal ribosomal entry site (IRES)-mediated translation are regulated differently within cells. Viral IRES-mediated translation often remains active when cellular cap-dependent translation is severely impaired under cellular stresses induced by virus infection. To investigate how cellular stresses influence the efficiency of viral IRES-mediated translation, we used a bicistronic luciferase reporter construct harbouring IRES elements from the following viruses: encephalomyocarditis virus (EMCV), foot-and-mouth disease virus (FMDV), hepatitis C virus (HCV) or human rhinovirus (HRV). NIH3T3 cells transfected with these bicistronic reporter constructs were subjected to different cellular stresses. Increased translation initiation was only observed under amino acid starvation when EMCV or FMDV IRES elements were present. To identify cellular mechanisms that promoted viral IRES-mediated translation, we tested the involvement of eukaryotic initiation factor 4E-binding protein (4E-BP), general control non-depressed 2 (GCN2) and eukaryotic initiation factor 2B (eIF2B), as these are known to be modulated under amino acid starvation. Knockdown of 4E-BP1 impaired the promotion of EMCV and FMDV IRES-mediated translation under amino acid starvation, whereas GCN2 and eIF2B were not involved. To further investigate how 4E-BP1 regulates translation initiated by EMCV and FMDV IRES elements, we used a phosphoinositide kinase-3 inhibitor (LY294002), an mTOR inhibitor (Torin1) or leucine starvation to mimic 4E-BP1 dephosphorylation induced by amino acid starvation. 4E-BP1 dephosphorylation induced by the treatments was not sufficient to promote viral IRES-mediated translation. These results suggest that 4E-BP1 regulates EMCV and FMDV IRES-mediated translation under amino acid starvation, but not via its dephosphorylation.
Division of Biomedical Science, Faculty of Medicine, Memorial University of Newfoundland, 300 Prince Philip Drive, St John's, NL A1B 3V6, Canada.
Maria
M
Licursi
Division of Biomedical Science, Faculty of Medicine, Memorial University of Newfoundland, 300 Prince Philip Drive, St John's, NL A1B 3V6, Canada.
Yumiko
Y
Komatsu
Division of Biomedical Science, Faculty of Medicine, Memorial University of Newfoundland, 300 Prince Philip Drive, St John's, NL A1B 3V6, Canada.
Theerawat
T
Pongnopparat
Division of Biomedical Science, Faculty of Medicine, Memorial University of Newfoundland, 300 Prince Philip Drive, St John's, NL A1B 3V6, Canada.
Kensuke
K
Hirasawa
D000596:Amino Acids / Q000378:metabolism*; D000818:Animals; D019677:Artificial Gene Fusion; D002460:Cell Line; D017930:Genes, Reporter; D008156:Luciferases / Q000032:analysis / Q000235:genetics; D051379:Mice; D014176:Protein Biosynthesis; D012328:RNA Viruses / Q000235:genetics / Q000502:physiology*
D000596:Amino Acids; D008156:Luciferases