title: Specific domains in yeast translation initiation factor eIF4G strongly bias RNA unwinding activity of the eIF4F complex toward duplexes with 5'-overhangs.
abstract: During eukaryotic translation initiation, the 43 S ribosomal pre-initiation complex is recruited to the 5'-end of an mRNA through its interaction with the 7-methylguanosine cap, and it subsequently scans along the mRNA to locate the start codon. Both mRNA recruitment and scanning require the removal of secondary structure within the mRNA. Eukaryotic translation initiation factor 4A is an essential component of the translational machinery thought to participate in the clearing of secondary structural elements in the 5'-untranslated regions of mRNAs. eIF4A is part of the 5'-7-methylguanosine cap-binding complex, eIF4F, along with eIF4E, the cap-binding protein, and the scaffolding protein eIF4G. Here, we show that Saccharomyces cerevisiae eIF4F has a strong preference for unwinding an RNA duplex with a single-stranded 5'-overhang versus the same duplex with a 3'-overhang or without an overhang. In contrast, eIF4A on its own has little RNA substrate specificity. Using a series of deletion constructs of eIF4G, we demonstrate that its three previously elucidated RNA binding domains work together to provide eIF4F with its 5'-end specificity, both by promoting unwinding of substrates with 5'-overhangs and inhibiting unwinding of substrates with 3'-overhangs. Our data suggest that the RNA binding domains of eIF4G provide the S. cerevisiae eIF4F complex with a second mechanism, in addition to the eIF4E-cap interaction, for directing the binding of pre-initiation complexes to the 5'-ends of mRNAs and for biasing scanning in the 5' to 3' direction.
authors.affiliation: Department of Biophysics and Biophysical Chemistry, Johns Hopkins University School of Medicine, Baltimore, Maryland 21205-2185, USA.
authors.forename: Vaishnavi
authors.initials: V
authors.lastname: Rajagopal
authors.forename: Eun-Hee
authors.initials: EH
authors.lastname: Park
authors.forename: Alan G
authors.initials: AG
authors.lastname: Hinnebusch
authors.forename: Jon R
authors.initials: JR
authors.lastname: Lorsch
mesh_terms: D020121:5' Untranslated Regions / Q000502:physiology*; D000595:Amino Acid Sequence; D039562:Eukaryotic Initiation Factor-4F / Q000235:genetics / Q000378:metabolism*; D039603:Eukaryotic Initiation Factor-4G / Q000235:genetics / Q000378:metabolism*; D017434:Protein Structure, Tertiary; D012315:RNA Caps / Q000235:genetics / Q000378:metabolism*; D059370:RNA Folding / Q000502:physiology*; D012331:RNA, Fungal / Q000235:genetics / Q000378:metabolism*; D012441:Saccharomyces cerevisiae / Q000235:genetics / Q000378:metabolism*; D029701:Saccharomyces cerevisiae Proteins / Q000235:genetics / Q000378:metabolism*; D017384:Sequence Deletion
chemical_list: D020121:5' Untranslated Regions; D039562:Eukaryotic Initiation Factor-4F; D039603:Eukaryotic Initiation Factor-4G; D012315:RNA Caps; D012331:RNA, Fungal; D029701:Saccharomyces cerevisiae Proteins