title: Ensemble cryo-EM uncovers inchworm-like translocation of a viral IRES through the ribosome.
abstract: Internal ribosome entry sites (IRESs) mediate cap-independent translation of viral mRNAs. Using electron cryo-microscopy of a single specimen, we present five ribosome structures formed with the Taura syndrome virus IRES and translocase eEF2•GTP bound with sordarin. The structures suggest a trajectory of IRES translocation, required for translation initiation, and provide an unprecedented view of eEF2 dynamics. The IRES rearranges from extended to bent to extended conformations. This inchworm-like movement is coupled with ribosomal inter-subunit rotation and 40S head swivel. eEF2, attached to the 60S subunit, slides along the rotating 40S subunit to enter the A site. Its diphthamide-bearing tip at domain IV separates the tRNA-mRNA-like pseudoknot I (PKI) of the IRES from the decoding center. This unlocks 40S domains, facilitating head swivel and biasing IRES translocation via hitherto-elusive intermediates with PKI captured between the A and P sites. The structures suggest missing links in our understanding of tRNA translocation.
authors.affiliation: Janelia Research Campus, Howard Hughes Medical Institute, Ashburn, United States.
authors.forename: Priyanka D
authors.initials: PD
authors.lastname: Abeyrathne
authors.affiliation: RNA Therapeutics Institute, Department of Biochemistry and Molecular Pharmacology, University of Massachusetts Medical School, Worcester, United States.
authors.forename: Cha San
authors.identifier: 0000-0002-1579-0362
authors.initials: CS
authors.lastname: Koh
authors.affiliation: Janelia Research Campus, Howard Hughes Medical Institute, Ashburn, United States.
authors.forename: Timothy
authors.initials: T
authors.lastname: Grant
authors.affiliation: Janelia Research Campus, Howard Hughes Medical Institute, Ashburn, United States.
authors.forename: Nikolaus
authors.initials: N
authors.lastname: Grigorieff
authors.affiliation: RNA Therapeutics Institute, Department of Biochemistry and Molecular Pharmacology, University of Massachusetts Medical School, Worcester, United States.
authors.forename: Andrei A
authors.identifier: 0000-0003-1588-717X
authors.initials: AA
authors.lastname: Korostelev
mesh_terms: D020285:Cryoelectron Microscopy / Q000379:methods; D056629:Dicistroviridae / Q000235:genetics; D006160:Guanosine Triphosphate / Q000378:metabolism; D007091:Image Processing, Computer-Assisted / Q000379:methods; D000067556:Internal Ribosome Entry Sites; D046911:Macromolecular Substances / Q000378:metabolism / Q000648:ultrastructure; D009690:Nucleic Acid Conformation; D020652:Peptide Elongation Factor 2 / Q000378:metabolism* / Q000648:ultrastructure; D014176:Protein Biosynthesis; D012333:RNA, Messenger / Q000235:genetics / Q000378:metabolism* / Q000648:ultrastructure; D012367:RNA, Viral / Q000235:genetics / Q000378:metabolism* / Q000648:ultrastructure; D012270:Ribosomes / Q000378:metabolism / Q000648:ultrastructure; D012441:Saccharomyces cerevisiae / Q000235:genetics
chemical_list: D000067556:Internal Ribosome Entry Sites; D046911:Macromolecular Substances; D020652:Peptide Elongation Factor 2; D012333:RNA, Messenger; D012367:RNA, Viral; D006160:Guanosine Triphosphate
keywords: IRES; S. cerevisiae; Taura syndrome virus; biochemistry; biophysics; elongation factor eEF2; internal ribosome entry site; ribosome; structural biology; translocation