title: Structural determinants of an internal ribosome entry site that direct translational reading frame selection.
abstract: The dicistrovirus intergenic internal ribosome entry site (IGR IRES) directly recruits the ribosome and initiates translation using a non-AUG codon. A subset of IGR IRESs initiates translation in either of two overlapping open reading frames (ORFs), resulting in expression of the 0 frame viral structural polyprotein and an overlapping +1 frame ORFx. A U-G base pair adjacent to the anticodon-like pseudoknot of the IRES directs +1 frame translation. Here, we show that the U-G base pair is not absolutely required for +1 frame translation. Extensive mutagenesis demonstrates that 0 and +1 frame translation can be uncoupled. Ribonucleic acid (RNA) structural probing analyses reveal that the mutant IRESs adopt distinct conformations. Toeprinting analysis suggests that the reading frame is selected at a step downstream of ribosome assembly. We propose a model whereby the IRES adopts conformations to occlude the 0 frame aminoacyl-tRNA thereby allowing delivery of the +1 frame aminoacyl-tRNA to the A site to initiate translation of ORFx. This study provides a new paradigm for programmed recoding mechanisms that increase the coding capacity of a viral genome.
authors.affiliation: Department of Biochemistry and Molecular Biology, University of British Columbia, Vancouver, BC V6T 1Z3, Canada.
authors.forename: Qian
authors.initials: Q
authors.lastname: Ren
authors.affiliation: Department of Biochemistry and Molecular Biology, University of British Columbia, Vancouver, BC V6T 1Z3, Canada.
authors.forename: Hilda H T
authors.initials: HH
authors.lastname: Au
authors.affiliation: Department of Biochemistry and Molecular Biology, University of British Columbia, Vancouver, BC V6T 1Z3, Canada.
authors.forename: Qing S
authors.initials: QS
authors.lastname: Wang
authors.affiliation: Department of Biochemistry and Molecular Biology, University of British Columbia, Vancouver, BC V6T 1Z3, Canada.
authors.forename: Seonghoon
authors.initials: S
authors.lastname: Lee
authors.affiliation: Department of Biochemistry and Molecular Biology, University of British Columbia, Vancouver, BC V6T 1Z3, Canada ej@mail.ubc.ca.
authors.forename: Eric
authors.initials: E
authors.lastname: Jan
mesh_terms: D020029:Base Pairing; D056629:Dicistroviridae / Q000235:genetics; D006367:HeLa Cells; D006801:Humans; D009154:Mutation; D009690:Nucleic Acid Conformation; D016366:Open Reading Frames; D010442:Peptide Chain Initiation, Translational; D012367:RNA, Viral / Q000737:chemistry*; D012270:Ribosomes / Q000378:metabolism
chemical_list: D012367:RNA, Viral