title: Global shape mimicry of tRNA within a viral internal ribosome entry site mediates translational reading frame selection.
abstract: The dicistrovirus intergenic region internal ribosome entry site (IRES) adopts a triple-pseudoknotted RNA structure and occupies the core ribosomal E, P, and A sites to directly recruit the ribosome and initiate translation at a non-AUG codon. A subset of dicistrovirus IRESs directs translation in the 0 and +1 frames to produce the viral structural proteins and a +1 overlapping open reading frame called ORFx, respectively. Here we show that specific mutations of two unpaired adenosines located at the core of the three-helical junction of the honey bee dicistrovirus Israeli acute paralysis virus (IAPV) IRES PKI domain can uncouple 0 and +1 frame translation, suggesting that the structure adopts distinct conformations that contribute to 0 or +1 frame translation. Using a reconstituted translation system, we show that ribosomes assembled on mutant IRESs that direct exclusive 0 or +1 frame translation lack reading frame fidelity. Finally, a nuclear magnetic resonance/small-angle X-ray scattering hybrid approach reveals that the PKI domain of the IAPV IRES adopts an RNA structure that resembles a complete tRNA. The tRNA shape-mimicry enables the viral IRES to gain access to the ribosome tRNA-binding sites and form intermolecular contacts with the ribosome that are necessary for initiating IRES translation in a specific reading frame.
authors.affiliation: Department of Biochemistry and Molecular Biology, University of British Columbia, Vancouver, BC, Canada V6T 1Z3.
authors.forename: Hilda H
authors.initials: HH
authors.lastname: Au
authors.affiliation: Department of Biochemistry, University of Wisconsin-Madison, Madison, WI 53706.
authors.forename: Gabriel
authors.initials: G
authors.lastname: Cornilescu
authors.affiliation: Department of Biochemistry, University of Wisconsin-Madison, Madison, WI 53706.
authors.forename: Kathryn D
authors.initials: KD
authors.lastname: Mouzakis
authors.affiliation: Department of Biochemistry and Molecular Biology, University of British Columbia, Vancouver, BC, Canada V6T 1Z3.
authors.forename: Qian
authors.initials: Q
authors.lastname: Ren
authors.affiliation: Department of Biochemistry, University of Wisconsin-Madison, Madison, WI 53706.
authors.forename: Jordan E
authors.initials: JE
authors.lastname: Burke
authors.affiliation: Department of Biochemistry and Molecular Biology, University of British Columbia, Vancouver, BC, Canada V6T 1Z3.
authors.forename: Seonghoon
authors.initials: S
authors.lastname: Lee
authors.affiliation: Department of Biochemistry, University of Wisconsin-Madison, Madison, WI 53706 sebutcher@wisc.edu ej@mail.ubc.ca.
authors.forename: Samuel E
authors.initials: SE
authors.lastname: Butcher
authors.affiliation: Department of Biochemistry and Molecular Biology, University of British Columbia, Vancouver, BC, Canada V6T 1Z3 sebutcher@wisc.edu ej@mail.ubc.ca.
authors.forename: Eric
authors.initials: E
authors.lastname: Jan
mesh_terms: D000818:Animals; D001483:Base Sequence; D002417:Cattle; D056629:Dicistroviridae / Q000235:genetics*; D006801:Humans; D000067556:Internal Ribosome Entry Sites / Q000235:genetics*; D009682:Magnetic Resonance Spectroscopy; D008958:Models, Molecular; D008969:Molecular Sequence Data; D009154:Mutation / Q000235:genetics; D009690:Nucleic Acid Conformation; D016366:Open Reading Frames / Q000235:genetics*; D010454:Peptide Termination Factors; D014176:Protein Biosynthesis; D012343:RNA, Transfer / Q000737:chemistry* / Q000235:genetics*; D012270:Ribosomes / Q000378:metabolism; D053838:Scattering, Small Angle; D014961:X-Ray Diffraction
chemical_list: D000067556:Internal Ribosome Entry Sites; D010454:Peptide Termination Factors; D012343:RNA, Transfer
keywords: RNA; internal ribosome entry site; ribosome; translation; virus