title: Dissociation of double-stranded polynucleotide helical structures by eukaryotic initiation factors, as revealed by a novel assay.
abstract: A new technique has been applied to the study of the RNA secondary structure unwinding activity of the eukaryotic initiation factors (eIFs) 4F, 4A, and 4B. Secondary structures were generated at the 5' ends of reovirus and globin mRNA molecules by hybridization with 32P-labeled cDNA molecules 15 nucleotide residues long. The dissociation of the labeled cDNAs from the mRNAs was assayed by a gel filtration chromatography procedure which separates the free cDNAs from mRNAs and mRNA/cDNA hybrids. When the three factors were tested alone, only eIF-4F stimulated dissociation of hybrids. The combination of eIF-4A plus eIF-4B also exhibited a strong hybrid dissociating activity, which was markedly temperature dependent. Under optimum conditions, up to 90% of the hybrid structures are disrupted in 60 min. These results demonstrate for the first time that stable double-stranded regions can be melted and dissociated by eIFs. They also characterize more precisely the first step in the structure unwinding reaction.
authors.affiliation: Department of Biology, Washington University, St. Louis, Missouri 63130.
authors.forename: T G
authors.initials: TG
authors.lastname: Lawson
authors.forename: K A
authors.initials: KA
authors.lastname: Lee
authors.forename: M M
authors.initials: MM
authors.lastname: Maimone
authors.forename: R D
authors.initials: RD
authors.lastname: Abramson
authors.forename: T E
authors.initials: TE
authors.lastname: Dever
authors.forename: W C
authors.initials: WC
authors.lastname: Merrick
authors.forename: R E
authors.initials: RE
authors.lastname: Thach
mesh_terms: D002850:Chromatography, Gel; D004247:DNA / Q000378:metabolism*; D004265:DNA Helicases / Q000032:analysis; D004277:DNA, Single-Stranded / Q000032:analysis; D039601:Eukaryotic Initiation Factor-4A; D039562:Eukaryotic Initiation Factor-4F; D039642:Eukaryotic Initiation Factors; D005914:Globins / Q000032:analysis / Q000235:genetics; D009693:Nucleic Acid Hybridization; D010448:Peptide Initiation Factors / Q000235:genetics / Q000378:metabolism*; D012330:RNA, Double-Stranded / Q000032:analysis; D012333:RNA, Messenger / Q000032:analysis / Q000378:metabolism*; D012087:Reoviridae / Q000032:analysis / Q000235:genetics
chemical_list: D004277:DNA, Single-Stranded; D039562:Eukaryotic Initiation Factor-4F; D039642:Eukaryotic Initiation Factors; D010448:Peptide Initiation Factors; D012330:RNA, Double-Stranded; D012333:RNA, Messenger; C031799:eIF-4B; D005914:Globins; D004247:DNA; D039601:Eukaryotic Initiation Factor-4A; D004265:DNA Helicases