Dynamic Recognition of the mRNA Cap by Saccharomyces cerevisiae eIF4E

• Published in: Structure (London) Vol. 21; no. 12; pp. 2197 - 2207
• Main Authors: O’Leary, Seán E., Petrov, Alexey, Chen, Jin, Puglisi, Joseph D.
• Format: Journal Article
• Language: English
• Published: United States Elsevier Inc 03.12.2013
• Subjects: Carbocyanines; Eukaryotic Initiation Factor-4E - chemistry; Eukaryotic Initiation Factor-4G - chemistry; Eukaryotic Initiation Factor-4G - metabolism; Exoribonucleases - chemistry; Fluorescence Resonance Energy Transfer; Fluorescent Dyes; Nucleic Acid Conformation; Peptide Chain Initiation, Translational; Poly A - chemistry; Poly(A)-Binding Proteins - chemistry; RNA Caps - chemistry; RNA, Fungal - chemistry; Saccharomyces cerevisiae; Saccharomyces cerevisiae - genetics; Saccharomyces cerevisiae - metabolism; Saccharomyces cerevisiae Proteins - chemistry
• ISSN: 0969-2126; 1878-4186
• DOI: 10.1016/j.str.2013.09.016

Recognition of the mRNA 5′ m7G(5′)ppp(5′)N cap is key to translation initiation for most eukaryotic mRNAs. The cap is bound by the eIF4F complex, consisting of a cap-binding protein (eIF4E), a “scaffold” protein (eIF4G), and an RNA helicase (eIF4A). As a central early step in initiation, regulation of eIF4F is crucial for cellular viability. Although the structure and function of eIF4E have been defined, a dynamic mechanistic picture of its activity at the molecular level in the eIF4F⋅mRNA complex is still unavailable. Here, using single-molecule fluorescence, we measured the effects of Saccharomyces cerevisiae eIF4F factors, mRNA secondary structure, and the poly(A)-binding protein Pab1p on eIF4E-mRNA binding dynamics. Our data provide an integrated picture of how eIF4G and mRNA structure modulate eIF4E-mRNA interaction, and uncover an eIF4G- and poly(A)-independent activity of poly(A)-binding protein that prolongs the eIF4E⋅mRNA complex lifetime. [Display omitted] •Used a single-molecule assay to observe S. cerevisiae eIF4E-mRNA interaction directly•Measured the kinetics of assembly of the eIF4E•eIF4G1•eIF4A•mRNA complex•Measured the modulation of eIF4E-RNA binding kinetics by RNA secondary structures•Observed Pab1p-mediated, poly(A)-independent stimulation of eIF4E-RNA binding The eIF4F mRNA cap-binding complex that includes a cap-binding protein (eIF4E) is central to eukaryotic translation initiation. O'Leary et al. probe dynamics of the eIF4E-mRNA interaction and show how eIF4F factors, mRNA, and the poly(A)-binding protein yield distinct dynamic outcomes affecting downstream events.