title: Cryo-EM of ribosomal 80S complexes with termination factors reveals the translocated cricket paralysis virus IRES.
abstract: The cricket paralysis virus (CrPV) uses an internal ribosomal entry site (IRES) to hijack the ribosome. In a remarkable RNA-based mechanism involving neither initiation factor nor initiator tRNA, the CrPV IRES jumpstarts translation in the elongation phase from the ribosomal A site. Here, we present cryoelectron microscopy (cryo-EM) maps of 80S⋅CrPV-STOP ⋅ eRF1 ⋅ eRF3 ⋅ GMPPNP and 80S⋅CrPV-STOP ⋅ eRF1 complexes, revealing a previously unseen binding state of the IRES and directly rationalizing that an eEF2-dependent translocation of the IRES is required to allow the first A-site occupation. During this unusual translocation event, the IRES undergoes a pronounced conformational change to a more stretched conformation. At the same time, our structural analysis provides information about the binding modes of eRF1 ⋅ eRF3 ⋅ GMPPNP and eRF1 in a minimal system. It shows that neither eRF3 nor ABCE1 are required for the active conformation of eRF1 at the intersection between eukaryotic termination and recycling.
authors.affiliation: Institut für Medizinische Physik und Biophysik, Charite - Universitätsmedizin Berlin, Charitéplatz 1, 10117 Berlin, Germany.
authors.forename: Margarita
authors.initials: M
authors.lastname: Muhs
authors.affiliation: Institut für Medizinische Physik und Biophysik, Charite - Universitätsmedizin Berlin, Charitéplatz 1, 10117 Berlin, Germany.
authors.forename: Tarek
authors.initials: T
authors.lastname: Hilal
authors.affiliation: Institut für Medizinische Physik und Biophysik, Charite - Universitätsmedizin Berlin, Charitéplatz 1, 10117 Berlin, Germany; UltraStrukturNetzwerk, Max Planck Institute for Molecular Genetics, 14195 Berlin, Germany.
authors.forename: Thorsten
authors.initials: T
authors.lastname: Mielke
authors.affiliation: Department of Cell Biology, SUNY Downstate Medical Center, Brooklyn, NY 11203, USA.
authors.forename: Maxim A
authors.initials: MA
authors.lastname: Skabkin
authors.affiliation: Theoretical Biology and Biophysics Group, Theoretical Division, Los Alamos National Laboratory, Los Alamos, NM 87545, USA; New Mexico Consortium, 4200 West Jemez Road, Suite 301, Los Alamos, NM 87544, USA.
authors.forename: Karissa Y
authors.initials: KY
authors.lastname: Sanbonmatsu
authors.affiliation: Department of Cell Biology, SUNY Downstate Medical Center, Brooklyn, NY 11203, USA.
authors.forename: Tatyana V
authors.initials: TV
authors.lastname: Pestova
authors.affiliation: Institut für Medizinische Physik und Biophysik, Charite - Universitätsmedizin Berlin, Charitéplatz 1, 10117 Berlin, Germany. Electronic address: christian.spahn@charite.de.
authors.forename: Christian M T
authors.initials: CM
authors.lastname: Spahn
mesh_terms: D018528:ATP-Binding Cassette Transporters / Q000378:metabolism; D020285:Cryoelectron Microscopy / Q000379:methods; D056629:Dicistroviridae / Q000737:chemistry / Q000235:genetics*; D039642:Eukaryotic Initiation Factors / Q000737:chemistry / Q000378:metabolism; D008958:Models, Molecular; D008968:Molecular Conformation; D008969:Molecular Sequence Data; D009690:Nucleic Acid Conformation; D010454:Peptide Termination Factors / Q000737:chemistry / Q000378:metabolism*; D014176:Protein Biosynthesis; D012333:RNA, Messenger / Q000737:chemistry* / Q000378:metabolism; D012343:RNA, Transfer / Q000737:chemistry / Q000378:metabolism; D012367:RNA, Viral / Q000737:chemistry* / Q000378:metabolism; D012270:Ribosomes / Q000737:chemistry / Q000378:metabolism*
chemical_list: D018528:ATP-Binding Cassette Transporters; D039642:Eukaryotic Initiation Factors; D010454:Peptide Termination Factors; D012333:RNA, Messenger; D012367:RNA, Viral; D012343:RNA, Transfer