title: Structural analysis of an eIF3 subcomplex reveals conserved interactions required for a stable and proper translation pre-initiation complex assembly.
abstract: Translation initiation factor eIF3 acts as the key orchestrator of the canonical initiation pathway in eukaryotes, yet its structure is greatly unexplored. We report the 2.2 Å resolution crystal structure of the complex between the yeast seven-bladed β-propeller eIF3i/TIF34 and a C-terminal α-helix of eIF3b/PRT1, which reveals universally conserved interactions. Mutating these interactions displays severe growth defects and eliminates association of eIF3i/TIF34 and strikingly also eIF3g/TIF35 with eIF3 and 40S subunits in vivo. Unexpectedly, 40S-association of the remaining eIF3 subcomplex and eIF5 is likewise destabilized resulting in formation of aberrant pre-initiation complexes (PICs) containing eIF2 and eIF1, which critically compromises scanning arrest on mRNA at its AUG start codon suggesting that the contacts between mRNA and ribosomal decoding site are impaired. Remarkably, overexpression of eIF3g/TIF35 suppresses the leaky scanning and growth defects most probably by preventing these aberrant PICs to form. Leaky scanning is also partially suppressed by eIF1, one of the key regulators of AUG recognition, and its mutant sui1(G107R) but the mechanism differs. We conclude that the C-terminus of eIF3b/PRT1 orchestrates co-operative recruitment of eIF3i/TIF34 and eIF3g/TIF35 to the 40S subunit for a stable and proper assembly of 48S pre-initiation complexes necessary for stringent AUG recognition on mRNAs.
authors.affiliation: Laboratory of Regulation of Gene Expression, Institute of Microbiology ASCR, v.v.i., Videnska 1083, Prague, 142 20, Czech Republic.
authors.forename: Anna
authors.initials: A
authors.lastname: Herrmannová
authors.forename: Dalia
authors.initials: D
authors.lastname: Daujotyte
authors.forename: Ji-Chun
authors.initials: JC
authors.lastname: Yang
authors.forename: Lucie
authors.initials: L
authors.lastname: Cuchalová
authors.forename: Fabrice
authors.initials: F
authors.lastname: Gorrec
authors.forename: Susan
authors.initials: S
authors.lastname: Wagner
authors.forename: István
authors.initials: I
authors.lastname: Dányi
authors.forename: Peter J
authors.initials: PJ
authors.lastname: Lukavsky
authors.forename: Leos Shivaya
authors.initials: LS
authors.lastname: Valásek
mesh_terms: D018387:Codon, Initiator; D018360:Crystallography, X-Ray; D015977:Eukaryotic Initiation Factor-1 / Q000235:genetics; D039621:Eukaryotic Initiation Factor-3 / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D018628:Gene Dosage; D008958:Models, Molecular; D009154:Mutation; D010442:Peptide Chain Initiation, Translational; D010641:Phenotype; D017434:Protein Structure, Tertiary; D054682:Ribosome Subunits, Small, Eukaryotic / Q000378:metabolism; D029701:Saccharomyces cerevisiae Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D015003:Yeasts / Q000235:genetics / Q000254:growth & development
chemical_list: D018387:Codon, Initiator; D015977:Eukaryotic Initiation Factor-1; D039621:Eukaryotic Initiation Factor-3; C050998:Prt1 protein, S cerevisiae; D029701:Saccharomyces cerevisiae Proteins; C102947:TIF34 protein, S cerevisiae; C026022:eukaryotic peptide initiation factor-1A
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