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RNA. Mar 2001; 7(3): 382–394.
PMCID: PMC1370095

The requirement for eukaryotic initiation factor 4A (elF4A) in translation is in direct proportion to the degree of mRNA 5' secondary structure.

Abstract

Eukaryotic initiation factor (elF) 4A functions as a subunit of the initiation factor complex elF4F, which mediates the binding of mRNA to the ribosome. elF4A possesses ATPase and RNA helicase activities and is the prototype for a large family of putative RNA helicases (the DEAD box family). It is thought that the function of elF4A during translation initiation is to unwind the mRNA secondary structure in the 5' UTR to facilitate ribosome binding. However, the evidence to support this hypothesis is rather indirect, and it was reported that elF4A is also required for the translation of mRNAs possessing minimal 5' UTR secondary structure. Were this hypothesis correct, the requirement for elF4A should correlate with the degree of mRNA secondary structure. To test this hypothesis, the effect of a dominant-negative mutant of mammalian elF4A on translation of mRNAs with various degrees of secondary structure was studied in vitro. Here, we show that mRNAs containing stable secondary structure in the 5' untranslated region are more susceptible to inhibition by the elF4A mutant. The mutant protein also strongly inhibits translation from several picornavirus internal ribosome entry sites (IRES), although to different extents. UV crosslinking of elF4F subunits and elF4B to the mRNA cap structure is dramatically reduced by the elF4A mutant and RNA secondary structure. Finally, the elF4A mutant forms a more stable complex with elF4G, as compared to the wild-type elF4A, thus explaining the mechanism by which substoichiometric amounts of mutant elF4A inhibit translation.

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Selected References

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  • Altmann M, Blum S, Pelletier J, Sonenberg N, Wilson TM, Trachsel H. Translation initiation factor-dependent extracts from Saccharomyces cerevisiae. Biochim Biophys Acta. 1990 Aug 27;1050(1-3):155–159. [PubMed]
  • Altmann M, Blum S, Wilson TM, Trachsel H. The 5'-leader sequence of tobacco mosaic virus RNA mediates initiation-factor-4E-independent, but still initiation-factor-4A-dependent translation in yeast extracts. Gene. 1990 Jul 2;91(1):127–129. [PubMed]
  • Baim SB, Sherman F. mRNA structures influencing translation in the yeast Saccharomyces cerevisiae. Mol Cell Biol. 1988 Apr;8(4):1591–1601. [PMC free article] [PubMed]
  • Belsham GJ, Sonenberg N. RNA-protein interactions in regulation of picornavirus RNA translation. Microbiol Rev. 1996 Sep;60(3):499–511. [PMC free article] [PubMed]
  • Benne R, Wong C, Luedi M, Hershey JW. Purification and characterization of initiation factor IF-E2 from rabbit reticulocytes. J Biol Chem. 1976 Dec 10;251(23):7675–7681. [PubMed]
  • Benz J, Trachsel H, Baumann U. Crystal structure of the ATPase domain of translation initiation factor 4A from Saccharomyces cerevisiae--the prototype of the DEAD box protein family. Structure. 1999 Jun 15;7(6):671–679. [PubMed]
  • Berben-Bloemheuvel G, Kasperaitis MA, van Heugten H, Thomas AA, van Steeg H, Voorma HO. Interaction of initiation factors with the cap structure of chimaeric mRNA containing the 5'-untranslated regions of Semliki Forest virus RNA is related to translational efficiency. Eur J Biochem. 1992 Sep 15;208(3):581–587. [PubMed]
  • Blair GE, Dahl HH, Truelsen E, Lelong JC. Functional identity of a mouse ascites and a rabbit reticulocyte initiation factor required for natural mRNA translation. Nature. 1977 Feb 17;265(5595):651–653. [PubMed]
  • Blum S, Schmid SR, Pause A, Buser P, Linder P, Sonenberg N, Trachsel H. ATP hydrolysis by initiation factor 4A is required for translation initiation in Saccharomyces cerevisiae. Proc Natl Acad Sci U S A. 1992 Aug 15;89(16):7664–7668. [PMC free article] [PubMed]
  • Browning KS, Lax SR, Humphreys J, Ravel JM, Jobling SA, Gehrke L. Evidence that the 5'-untranslated leader of mRNA affects the requirement for wheat germ initiation factors 4A, 4F, and 4G. J Biol Chem. 1988 Jul 15;263(20):9630–9634. [PubMed]
  • de la Cruz J, Kressler D, Linder P. Unwinding RNA in Saccharomyces cerevisiae: DEAD-box proteins and related families. Trends Biochem Sci. 1999 May;24(5):192–198. [PubMed]
  • Edery I, Hümbelin M, Darveau A, Lee KA, Milburn S, Hershey JW, Trachsel H, Sonenberg N. Involvement of eukaryotic initiation factor 4A in the cap recognition process. J Biol Chem. 1983 Sep 25;258(18):11398–11403. [PubMed]
  • Edery I, Petryshyn R, Sonenberg N. Activation of double-stranded RNA-dependent kinase (dsl) by the TAR region of HIV-1 mRNA: a novel translational control mechanism. Cell. 1989 Jan 27;56(2):303–312. [PubMed]
  • Fletcher L, Corbin SD, Browning KS, Ravel JM. The absence of a m7G cap on beta-globin mRNA and alfalfa mosaic virus RNA 4 increases the amounts of initiation factor 4F required for translation. J Biol Chem. 1990 Nov 15;265(32):19582–19587. [PubMed]
  • Gehrke L, Auron PE, Quigley GJ, Rich A, Sonenberg N. 5'-Conformation of capped alfalfa mosaic virus ribonucleic acid 4 may reflect its independence of the cap structure or of cap-binding protein for efficient translation. Biochemistry. 1983 Oct 25;22(22):5157–5164. [PubMed]
  • Gingras AC, Raught B, Sonenberg N. eIF4 initiation factors: effectors of mRNA recruitment to ribosomes and regulators of translation. Annu Rev Biochem. 1999;68:913–963. [PubMed]
  • Gorbalenya AE, Koonin EV, Donchenko AP, Blinov VM. Two related superfamilies of putative helicases involved in replication, recombination, repair and expression of DNA and RNA genomes. Nucleic Acids Res. 1989 Jun 26;17(12):4713–4730. [PMC free article] [PubMed]
  • Gradi A, Imataka H, Svitkin YV, Rom E, Raught B, Morino S, Sonenberg N. A novel functional human eukaryotic translation initiation factor 4G. Mol Cell Biol. 1998 Jan;18(1):334–342. [PMC free article] [PubMed]
  • Grifo JA, Tahara SM, Morgan MA, Shatkin AJ, Merrick WC. New initiation factor activity required for globin mRNA translation. J Biol Chem. 1983 May 10;258(9):5804–5810. [PubMed]
  • Haghighat A, Sonenberg N. eIF4G dramatically enhances the binding of eIF4E to the mRNA 5'-cap structure. J Biol Chem. 1997 Aug 29;272(35):21677–21680. [PubMed]
  • Hellen CU, Pestova TV, Wimmer E. Effect of mutations downstream of the internal ribosome entry site on initiation of poliovirus protein synthesis. J Virol. 1994 Oct;68(10):6312–6322. [PMC free article] [PubMed]
  • Hunt SL, Hsuan JJ, Totty N, Jackson RJ. unr, a cellular cytoplasmic RNA-binding protein with five cold-shock domains, is required for internal initiation of translation of human rhinovirus RNA. Genes Dev. 1999 Feb 15;13(4):437–448. [PMC free article] [PubMed]
  • Imataka H, Gradi A, Sonenberg N. A newly identified N-terminal amino acid sequence of human eIF4G binds poly(A)-binding protein and functions in poly(A)-dependent translation. EMBO J. 1998 Dec 15;17(24):7480–7489. [PMC free article] [PubMed]
  • Imataka H, Sonenberg N. Human eukaryotic translation initiation factor 4G (eIF4G) possesses two separate and independent binding sites for eIF4A. Mol Cell Biol. 1997 Dec;17(12):6940–6947. [PMC free article] [PubMed]
  • Jackson RJ. The ATP requirement for initiation of eukaryotic translation varies according to the mRNA species. Eur J Biochem. 1991 Sep 1;200(2):285–294. [PubMed]
  • Jackson RJ, Kaminski A. Internal initiation of translation in eukaryotes: the picornavirus paradigm and beyond. RNA. 1995 Dec;1(10):985–1000. [PMC free article] [PubMed]
  • Jaramillo M, Dever TE, Merrick WC, Sonenberg N. RNA unwinding in translation: assembly of helicase complex intermediates comprising eukaryotic initiation factors eIF-4F and eIF-4B. Mol Cell Biol. 1991 Dec;11(12):5992–5997. [PMC free article] [PubMed]
  • Johnson ER, McKay DB. Crystallographic structure of the amino terminal domain of yeast initiation factor 4A, a representative DEAD-box RNA helicase. RNA. 1999 Dec;5(12):1526–1534. [PMC free article] [PubMed]
  • Kolupaeva VG, Pestova TV, Hellen CU, Shatsky IN. Translation eukaryotic initiation factor 4G recognizes a specific structural element within the internal ribosome entry site of encephalomyocarditis virus RNA. J Biol Chem. 1998 Jul 17;273(29):18599–18604. [PubMed]
  • Koromilas AE, Lazaris-Karatzas A, Sonenberg N. mRNAs containing extensive secondary structure in their 5' non-coding region translate efficiently in cells overexpressing initiation factor eIF-4E. EMBO J. 1992 Nov;11(11):4153–4158. [PMC free article] [PubMed]
  • Lee KA, Guertin D, Sonenberg N. mRNA secondary structure as a determinant in cap recognition and initiation complex formation. ATP-Mg2+ independent cross-linking of cap binding proteins to m7I-capped inosine-substituted reovirus mRNA. J Biol Chem. 1983 Jan 25;258(2):707–710. [PubMed]
  • Lee KA, Sonenberg N. Inactivation of cap-binding proteins accompanies the shut-off of host protein synthesis by poliovirus. Proc Natl Acad Sci U S A. 1982 Jun;79(11):3447–3451. [PMC free article] [PubMed]
  • Linder P, Lasko PF, Ashburner M, Leroy P, Nielsen PJ, Nishi K, Schnier J, Slonimski PP. Birth of the D-E-A-D box. Nature. 1989 Jan 12;337(6203):121–122. [PubMed]
  • Lomakin IB, Hellen CU, Pestova TV. Physical association of eukaryotic initiation factor 4G (eIF4G) with eIF4A strongly enhances binding of eIF4G to the internal ribosomal entry site of encephalomyocarditis virus and is required for internal initiation of translation. Mol Cell Biol. 2000 Aug;20(16):6019–6029. [PMC free article] [PubMed]
  • Lorsch JR, Herschlag D. The DEAD box protein eIF4A. 1. A minimal kinetic and thermodynamic framework reveals coupled binding of RNA and nucleotide. Biochemistry. 1998 Feb 24;37(8):2180–2193. [PubMed]
  • Lorsch JR, Herschlag D. The DEAD box protein eIF4A. 2. A cycle of nucleotide and RNA-dependent conformational changes. Biochemistry. 1998 Feb 24;37(8):2194–2206. [PubMed]
  • Lüking A, Stahl U, Schmidt U. The protein family of RNA helicases. Crit Rev Biochem Mol Biol. 1998;33(4):259–296. [PubMed]
  • Mathews DH, Sabina J, Zuker M, Turner DH. Expanded sequence dependence of thermodynamic parameters improves prediction of RNA secondary structure. J Mol Biol. 1999 May 21;288(5):911–940. [PubMed]
  • Meerovitch K, Svitkin YV, Lee HS, Lejbkowicz F, Kenan DJ, Chan EK, Agol VI, Keene JD, Sonenberg N. La autoantigen enhances and corrects aberrant translation of poliovirus RNA in reticulocyte lysate. J Virol. 1993 Jul;67(7):3798–3807. [PMC free article] [PubMed]
  • Merrick WC, Sonenberg N. Assays for eukaryotic translation factors that bind mRNA. Methods. 1997 Apr;11(4):333–342. [PubMed]
  • Milburn SC, Pelletier J, Sonenberg N, Hershey JW. Identification of the 80-kDa protein that crosslinks to the cap structure of eukaryotic mRNAs as initiation factor eIF-4B. Arch Biochem Biophys. 1988 Jul;264(1):348–350. [PubMed]
  • Nanbru C, Lafon I, Audigier S, Gensac MC, Vagner S, Huez G, Prats AC. Alternative translation of the proto-oncogene c-myc by an internal ribosome entry site. J Biol Chem. 1997 Dec 19;272(51):32061–32066. [PubMed]
  • Parkin NT, Cohen EA, Darveau A, Rosen C, Haseltine W, Sonenberg N. Mutational analysis of the 5' non-coding region of human immunodeficiency virus type 1: effects of secondary structure on translation. EMBO J. 1988 Sep;7(9):2831–2837. [PMC free article] [PubMed]
  • Pause A, Belsham GJ, Gingras AC, Donzé O, Lin TA, Lawrence JC, Jr, Sonenberg N. Insulin-dependent stimulation of protein synthesis by phosphorylation of a regulator of 5'-cap function. Nature. 1994 Oct 27;371(6500):762–767. [PubMed]
  • Pause A, Méthot N, Sonenberg N. The HRIGRXXR region of the DEAD box RNA helicase eukaryotic translation initiation factor 4A is required for RNA binding and ATP hydrolysis. Mol Cell Biol. 1993 Nov;13(11):6789–6798. [PMC free article] [PubMed]
  • Pause A, Méthot N, Svitkin Y, Merrick WC, Sonenberg N. Dominant negative mutants of mammalian translation initiation factor eIF-4A define a critical role for eIF-4F in cap-dependent and cap-independent initiation of translation. EMBO J. 1994 Mar 1;13(5):1205–1215. [PMC free article] [PubMed]
  • Pause A, Sonenberg N. Mutational analysis of a DEAD box RNA helicase: the mammalian translation initiation factor eIF-4A. EMBO J. 1992 Jul;11(7):2643–2654. [PMC free article] [PubMed]
  • Peck ML, Herschlag D. Effects of oligonucleotide length and atomic composition on stimulation of the ATPase activity of translation initiation factor elF4A. RNA. 1999 Sep;5(9):1210–1221. [PMC free article] [PubMed]
  • Pelletier J, Sonenberg N. Insertion mutagenesis to increase secondary structure within the 5' noncoding region of a eukaryotic mRNA reduces translational efficiency. Cell. 1985 Mar;40(3):515–526. [PubMed]
  • Pelletier J, Sonenberg N. Photochemical cross-linking of cap binding proteins to eucaryotic mRNAs: effect of mRNA 5' secondary structure. Mol Cell Biol. 1985 Nov;5(11):3222–3230. [PMC free article] [PubMed]
  • Pestova TV, Shatsky IN, Fletcher SP, Jackson RJ, Hellen CU. A prokaryotic-like mode of cytoplasmic eukaryotic ribosome binding to the initiation codon during internal translation initiation of hepatitis C and classical swine fever virus RNAs. Genes Dev. 1998 Jan 1;12(1):67–83. [PMC free article] [PubMed]
  • Pestova TV, Shatsky IN, Hellen CU. Functional dissection of eukaryotic initiation factor 4F: the 4A subunit and the central domain of the 4G subunit are sufficient to mediate internal entry of 43S preinitiation complexes. Mol Cell Biol. 1996 Dec;16(12):6870–6878. [PMC free article] [PubMed]
  • Pyronnet S, Imataka H, Gingras AC, Fukunaga R, Hunter T, Sonenberg N. Human eukaryotic translation initiation factor 4G (eIF4G) recruits mnk1 to phosphorylate eIF4E. EMBO J. 1999 Jan 4;18(1):270–279. [PMC free article] [PubMed]
  • Pyronnet S, Vagner S, Bouisson M, Prats AC, Vaysse N, Pradayrol L. Relief of ornithine decarboxylase messenger RNA translational repression induced by alternative splicing of its 5' untranslated region. Cancer Res. 1996 Apr 15;56(8):1742–1745. [PubMed]
  • Ray BK, Lawson TG, Kramer JC, Cladaras MH, Grifo JA, Abramson RD, Merrick WC, Thach RE. ATP-dependent unwinding of messenger RNA structure by eukaryotic initiation factors. J Biol Chem. 1985 Jun 25;260(12):7651–7658. [PubMed]
  • Richter-Cook NJ, Dever TE, Hensold JO, Merrick WC. Purification and characterization of a new eukaryotic protein translation factor. Eukaryotic initiation factor 4H. J Biol Chem. 1998 Mar 27;273(13):7579–7587. [PubMed]
  • Rogers GW, Jr, Richter NJ, Merrick WC. Biochemical and kinetic characterization of the RNA helicase activity of eukaryotic initiation factor 4A. J Biol Chem. 1999 Apr 30;274(18):12236–12244. [PubMed]
  • Rozen F, Edery I, Meerovitch K, Dever TE, Merrick WC, Sonenberg N. Bidirectional RNA helicase activity of eucaryotic translation initiation factors 4A and 4F. Mol Cell Biol. 1990 Mar;10(3):1134–1144. [PMC free article] [PubMed]
  • Sachs AB, Sarnow P, Hentze MW. Starting at the beginning, middle, and end: translation initiation in eukaryotes. Cell. 1997 Jun 13;89(6):831–838. [PubMed]
  • Schmid SR, Linder P. D-E-A-D protein family of putative RNA helicases. Mol Microbiol. 1992 Feb;6(3):283–291. [PubMed]
  • Schwemmle M, Schickinger J, Bader M, Sarre TF, Hilse K. A 60-kDa protein from rabbit reticulocytes specifically recognizes the capped 5' end of beta-globin mRNA. Eur J Biochem. 1991 Oct 1;201(1):139–145. [PubMed]
  • Sonenberg N. ATP/Mg++-dependent cross-linking of cap binding proteins to the 5' end of eukaryotic mRNA. Nucleic Acids Res. 1981 Apr 10;9(7):1643–1656. [PMC free article] [PubMed]
  • Sonenberg N. Remarks on the mechanism of ribosome binding to eukaryotic mRNAs. Gene Expr. 1993;3(3):317–323. [PubMed]
  • Sonenberg N, Morgan MA, Merrick WC, Shatkin AJ. A polypeptide in eukaryotic initiation factors that crosslinks specifically to the 5'-terminal cap in mRNA. Proc Natl Acad Sci U S A. 1978 Oct;75(10):4843–4847. [PMC free article] [PubMed]
  • Studier FW, Rosenberg AH, Dunn JJ, Dubendorff JW. Use of T7 RNA polymerase to direct expression of cloned genes. Methods Enzymol. 1990;185:60–89. [PubMed]
  • Svitkin YV, Agol VI. Complete translation of encephalomyocarditis virus RNA and faithful cleavage of virus-specific proteins in a cell-free system from Krebs-2 cells. FEBS Lett. 1978 Mar 1;87(1):7–11. [PubMed]
  • Svitkin YV, Lyapustin VN, Lashkevich VA, Agol VI. Differences between translation products of tick-borne encephalitis virus RNA in cell-free systems from Krebs-2 cells and rabbit reticulocytes: involvement of membranes in the processing of nascent precursors of flavivirus structural proteins. Virology. 1984 Jun;135(2):536–541. [PubMed]
  • Svitkin YV, Ovchinnikov LP, Dreyfuss G, Sonenberg N. General RNA binding proteins render translation cap dependent. EMBO J. 1996 Dec 16;15(24):7147–7155. [PMC free article] [PubMed]
  • Svitkin YV, Pause A, Sonenberg N. La autoantigen alleviates translational repression by the 5' leader sequence of the human immunodeficiency virus type 1 mRNA. J Virol. 1994 Nov;68(11):7001–7007. [PMC free article] [PubMed]
  • Tarun SZ, Jr, Sachs AB. Association of the yeast poly(A) tail binding protein with translation initiation factor eIF-4G. EMBO J. 1996 Dec 16;15(24):7168–7177. [PMC free article] [PubMed]
  • Trachsel H, Erni B, Schreier MH, Staehelin T. Initiation of mammalian protein synthesis. II. The assembly of the initiation complex with purified initiation factors. J Mol Biol. 1977 Nov;116(4):755–767. [PubMed]
  • Tsukiyama-Kohara K, Iizuka N, Kohara M, Nomoto A. Internal ribosome entry site within hepatitis C virus RNA. J Virol. 1992 Mar;66(3):1476–1483. [PMC free article] [PubMed]
  • Wen L, Huang JK, Blackshear PJ. Rat ornithine decarboxylase gene. Nucleotide sequence, potential regulatory elements, and comparison to the mouse gene. J Biol Chem. 1989 May 25;264(15):9016–9021. [PubMed]
  • Wigle DT, Smith AE. Specificity in initiation of protein synthesis in a fractionated mammalian cell-free system. Nat New Biol. 1973 Apr 4;242(118):136–140. [PubMed]
  • Yoder-Hill J, Pause A, Sonenberg N, Merrick WC. The p46 subunit of eukaryotic initiation factor (eIF)-4F exchanges with eIF-4A. J Biol Chem. 1993 Mar 15;268(8):5566–5573. [PubMed]

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