title: Genome-wide analysis of translational efficiency reveals distinct but overlapping functions of yeast DEAD-box RNA helicases Ded1 and eIF4A.
abstract: DEAD-box RNA helicases eIF4A and Ded1 are believed to promote translation initiation by resolving mRNA secondary structures that impede ribosome attachment at the mRNA 5' end or subsequent scanning of the 5' UTR, but whether they perform unique or overlapping functions in vivo is poorly understood. We compared the effects of mutations in Ded1 or eIF4A on global translational efficiencies (TEs) in budding yeast Saccharomyces cerevisiae by ribosome footprint profiling. Despite similar reductions in bulk translation, inactivation of a cold-sensitive Ded1 mutant substantially reduced the TEs of >600 mRNAs, whereas inactivation of a temperature-sensitive eIF4A variant encoded by tif1-A79V (in a strain lacking the ortholog TIF2) yielded <40 similarly impaired mRNAs. The broader requirement for Ded1 did not reflect more pervasive secondary structures at low temperature, as inactivation of temperature-sensitive and cold-sensitive ded1 mutants gave highly correlated results. Interestingly, Ded1-dependent mRNAs exhibit greater than average 5' UTR length and propensity for secondary structure, implicating Ded1 in scanning through structured 5' UTRs. Reporter assays confirmed that cap-distal stem-loop insertions increase dependence on Ded1 but not eIF4A for efficient translation. While only a small fraction of mRNAs shows a heightened requirement for eIF4A, dependence on eIF4A is correlated with requirements for Ded1 and 5' UTR features characteristic of Ded1-dependent mRNAs. Our findings suggest that Ded1 is critically required to promote scanning through secondary structures within 5' UTRs, and while eIF4A cooperates with Ded1 in this function, it also promotes a step of initiation common to virtually all yeast mRNAs.
authors.affiliation: Laboratory of Gene Regulation and Development, Eunice K. Shriver National Institute of Child Health and Human Development, National Institutes of Health, Bethesda, Maryland 20892, USA;
authors.forename: Neelam Dabas
authors.initials: ND
authors.lastname: Sen
authors.affiliation: Laboratory of Gene Regulation and Development, Eunice K. Shriver National Institute of Child Health and Human Development, National Institutes of Health, Bethesda, Maryland 20892, USA;
authors.forename: Fujun
authors.initials: F
authors.lastname: Zhou
authors.affiliation: Department of Molecular and Cell Biology, University of California, Berkeley, California 94720, USA.
authors.forename: Nicholas T
authors.initials: NT
authors.lastname: Ingolia
authors.affiliation: Laboratory of Gene Regulation and Development, Eunice K. Shriver National Institute of Child Health and Human Development, National Institutes of Health, Bethesda, Maryland 20892, USA;
authors.forename: Alan G
authors.initials: AG
authors.lastname: Hinnebusch
mesh_terms: D020121:5' Untranslated Regions; D053487:DEAD-box RNA Helicases / Q000235:genetics / Q000378:metabolism*; D039562:Eukaryotic Initiation Factor-4F / Q000235:genetics / Q000378:metabolism*; D016681:Genome, Fungal; D009154:Mutation; D009690:Nucleic Acid Conformation; D014176:Protein Biosynthesis; D020365:RNA Helicases; D012331:RNA, Fungal / Q000737:chemistry; D012333:RNA, Messenger / Q000737:chemistry; D012441:Saccharomyces cerevisiae / Q000235:genetics / Q000378:metabolism*; D029701:Saccharomyces cerevisiae Proteins / Q000235:genetics / Q000378:metabolism*
chemical_list: D020121:5' Untranslated Regions; D039562:Eukaryotic Initiation Factor-4F; D012331:RNA, Fungal; D012333:RNA, Messenger; D029701:Saccharomyces cerevisiae Proteins; C504497:TIF2 protein, S cerevisiae; C492152:DED1 protein, S cerevisiae; D053487:DEAD-box RNA Helicases; D020365:RNA Helicases; C504496:TIF1 protein, S cerevisiae