Functional reconstitution of human eukaryotic translation initiation factor 3 (eIF3)

Protein fate in higher eukaryotes is controlled by three complexes that share conserved architectural elements: the proteasome, COP9 signalosome, and eukaryotic translation initiation factor 3 (eIF3). Here we reconstitute the 13-subunit human eIF3 in Escherichia coli, revealing its structural core to be the eight subunits with conserved orthologues in the proteasome lid complex and COP9 signalosome. This structural core in eIF3 binds to the small (40S) ribosomal subunit, to translation initiation factors involved in mRNA cap-dependent initiation, and to the hepatitis C viral (HCV) internal ribosome entry site (IRES) RNA. Addition of the remaining eIF3 subunits enables reconstituted eIF3 to assemble intact initiation complexes with the HCV IRES. Negative-stain EM reconstructions of reconstituted eIF3 further reveal how the approximately 400 kDa molecular mass structural core organizes the highly flexible 800 kDa molecular mass eIF3 complex, and mediates translation initiation.

[1]  C. Robinson,et al.  Symmetrical modularity of the COP9 signalosome complex suggests its multifunctionality. , 2009, Structure.

[2]  A. Nomoto,et al.  Genetic analysis of internal ribosomal entry site on hepatitis C virus RNA: implication for involvement of the highly ordered structure and cell type-specific transacting factors. , 1997, Virology.

[3]  G. Wagner,et al.  Position of eukaryotic initiation factor eIF1 on the 40S ribosomal subunit determined by directed hydroxyl radical probing. , 2003, Genes & development.

[4]  Carol V Robinson,et al.  Structural Characterization of the Human Eukaryotic Initiation Factor 3 Protein Complex by Mass Spectrometry*S , 2007, Molecular & Cellular Proteomics.

[5]  Christopher Irving,et al.  Appion: an integrated, database-driven pipeline to facilitate EM image processing. , 2009, Journal of structural biology.

[6]  J. Doudna,et al.  The j-Subunit of Human Translation Initiation Factor eIF3 Is Required for the Stable Binding of eIF3 and Its Subcomplexes to 40 S Ribosomal Subunits in Vitro* , 2004, Journal of Biological Chemistry.

[7]  J. Doudna,et al.  Structural and mechanistic insights into hepatitis C viral translation initiation , 2007, Nature Reviews Microbiology.

[8]  Michal Sharon,et al.  Structural Organization of the 19S Proteasome Lid: Insights from MS of Intact Complexes , 2006, PLoS biology.

[9]  G. Wagner,et al.  Position of eukaryotic translation initiation factor eIF1A on the 40S ribosomal subunit mapped by directed hydroxyl radical probing , 2009, Nucleic acids research.

[10]  R. Bernards,et al.  The Translation Initiation Factor 3f (eIF3f) Exhibits a Deubiquitinase Activity Regulating Notch Activation , 2010, PLoS biology.

[11]  C. Hellen,et al.  Release of initiation factors from 48S complexes during ribosomal subunit joining and the link between establishment of codon-anticodon base-pairing and hydrolysis of eIF2-bound GTP. , 2004, Genes & development.

[12]  Joseph D. Puglisi,et al.  The Pathway of HCV IRES-Mediated Translation Initiation , 2004, Cell.

[13]  A. Hinnebusch,et al.  The yeast eIF3 subunits TIF32/a, NIP1/c, and eIF5 make critical connections with the 40S ribosome in vivo. , 2003, Genes & development.

[14]  R. Jackson,et al.  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. , 1998, Genes & development.

[15]  Alan G Hinnebusch,et al.  eIF3: a versatile scaffold for translation initiation complexes. , 2006, Trends in biochemical sciences.

[16]  U. Maitra,et al.  Mammalian Translation Initiation Factor eIF1 Functions with eIF1A and eIF3 in the Formation of a Stable 40 S Preinitiation Complex* , 2003, The Journal of Biological Chemistry.

[17]  F. Collart,et al.  A new vector for high-throughput, ligation-independent cloning encoding a tobacco etch virus protease cleavage site. , 2002, Protein expression and purification.

[18]  N. Ban,et al.  Crystal Structure of the Eukaryotic 60S Ribosomal Subunit in Complex with Initiation Factor 6 , 2011, Science.

[19]  J W Hershey,et al.  Identification of partners of TIF34, a component of the yeast eIF3 complex, required for cell proliferation and translation initiation , 1997, The EMBO journal.

[20]  C. Hill,et al.  Proteasome activators. , 2011, Molecular cell.

[21]  S. Formenti,et al.  Translational control in cancer , 2010, Nature Reviews Cancer.

[22]  P. D. de Jong,et al.  Ligation-independent cloning of PCR products (LIC-PCR). , 1990, Nucleic acids research.

[23]  M. Kozak.  Evaluation of the fidelity of initiation of translation in reticulocyte lysates from commercial sources. , 1990, Nucleic acids research.

[24]  W Chiu,et al.  EMAN: semiautomated software for high-resolution single-particle reconstructions. , 1999, Journal of structural biology.

[25]  A. Hinnebusch,et al.  A subcomplex of three eIF3 subunits binds eIF1 and eIF5 and stimulates ribosome binding of mRNA and tRNAiMet , 2001, The EMBO journal.

[26]  Lev L. Kisselev,et al.  In Vitro Reconstitution of Eukaryotic Translation Reveals Cooperativity between Release Factors eRF1 and eRF3 , 2006, Cell.

[27]  A. Hinnebusch,et al.  Complex Formation by All Five Homologues of Mammalian Translation Initiation Factor 3 Subunits from Yeast Saccharomyces cerevisiae * , 1998, The Journal of Biological Chemistry.

[28]  J. Kato,et al.  Mammalian COP9 signalosome , 2009, Genes to cells : devoted to molecular & cellular mechanisms.

[29]  R E Rhoads,et al.  Mutually Cooperative Binding of Eukaryotic Translation Initiation Factor (eIF) 3 and eIF4A to Human eIF4G-1* , 2000, The Journal of Biological Chemistry.

[30]  C. Hellen,et al.  Recycling of Eukaryotic Posttermination Ribosomal Complexes , 2007, Cell.

[31]  C. Hellen,et al.  Canonical eukaryotic initiation factors determine initiation of translation by internal ribosomal entry , 1996, Molecular and cellular biology.

[32]  Aaron K. LeFebvre,et al.  Translation Initiation Factor eIF4G-1 Binds to eIF3 through the eIF3e Subunit* , 2006, Journal of Biological Chemistry.

[33]  K. Zhou,et al.  Mechanism of ribosome recruitment by hepatitis C IRES RNA. , 2001, RNA.

[34]  J. Cate,et al.  Distinct regions of human eIF3 are sufficient for binding to the HCV IRES and the 40S ribosomal subunit. , 2010, Journal of molecular biology.

[35]  M. Glickman,et al.  PCI complexes: Beyond the proteasome, CSN, and eIF3 Troika. , 2009, Molecular cell.

[36]  Fabienne Beuron,et al.  Structural insights into the COP9 signalosome and its common architecture with the 26S proteasome lid and eIF3. , 2010, Structure.

[37]  C. Fraser,et al.  Characterization of eIF3k , 2003 .

[38]  A. Hinnebusch,et al.  Domains of eIF1A that mediate binding to eIF2, eIF3 and eIF5B and promote ternary complex recruitment in vivo , 2003, The EMBO journal.

[39]  K. Ikeda,et al.  Estrogen and phytoestrogen regulate the mRNA expression of adrenomedullin and adrenomedullin receptor components in the rat uterus , 2004, Molecular and Cellular Endocrinology.

[40]  C. Hellen,et al.  Specific Interaction of Eukaryotic Translation Initiation Factor 3 with the 5′ Nontranslated Regions of Hepatitis C Virus and Classical Swine Fever Virus RNAs , 1998, Journal of Virology.

[41]  C. Hellen,et al.  Binding of eukaryotic initiation factor 3 to ribosomal 40S subunits and its role in ribosomal dissociation and anti-association. , 2005, RNA.

[42]  M. Groll,et al.  The 26S proteasome: assembly and function of a destructive machine. , 2010, Trends in biochemical sciences.

[43]  D. Andreev,et al.  Eukaryotic translation initiation machinery can operate in a bacterial-like mode without eIF2 , 2008, Nature Structural &Molecular Biology.

[44]  C. Fraser,et al.  Characterization of eIF3k: a newly discovered subunit of mammalian translation initiation factor elF3. , 2003, European journal of biochemistry.

[45]  G. Wagner,et al.  Structure and interactions of the translation initiation factor eIF1 , 1999, The EMBO journal.

[46]  M. Honda,et al.  Structural requirements for initiation of translation by internal ribosome entry within genome-length hepatitis C virus RNA. , 1996, Virology.

[47]  E. Nogales,et al.  Structural Roles for Human Translation Factor eIF3 in Initiation of Protein Synthesis , 2005, Science.

[48]  N. Sonenberg,et al.  Reconstitution reveals the functional core of mammalian eIF3 , 2007, The EMBO journal.

[49]  R. Jackson,et al.  The mechanism of eukaryotic translation initiation and principles of its regulation , 2010, Nature Reviews Molecular Cell Biology.

[50]  N. Ban,et al.  Crystal Structure of the Eukaryotic 40S Ribosomal Subunit in Complex with Initiation Factor 1 , 2011, Science.

[51]  N. Grigorieff,et al.  Accurate determination of local defocus and specimen tilt in electron microscopy. , 2003, Journal of structural biology.

[52]  J. Doudna,et al.  eIF3j is located in the decoding center of the human 40S ribosomal subunit. , 2007, Molecular cell.

[53]  J Y Lau,et al.  The hepatitis C virus internal ribosome entry site adopts an ion-dependent tertiary fold. , 1999, Journal of molecular biology.

[54]  A. Hinnebusch,et al.  The C-Terminal Region of Eukaryotic Translation Initiation Factor 3a (eIF3a) Promotes mRNA Recruitment, Scanning, and, Together with eIF3j and the eIF3b RNA Recognition Motif, Selection of AUG Start Codons , 2010, Molecular and Cellular Biology.

[55]  Wen Jiang,et al.  EMAN2: an extensible image processing suite for electron microscopy. , 2007, Journal of structural biology.

[56]  Daniel Barsky,et al.  Mass spectrometry reveals modularity and a complete subunit interaction map of the eukaryotic translation factor eIF3 , 2008, Proceedings of the National Academy of Sciences.

[57]  J. Doudna,et al.  The HCV IRES pseudoknot positions the initiation codon on the 40S ribosomal subunit. , 2010, RNA.

[58]  Cara T. Pager,et al.  Hepatitis C Virus Core-Derived Peptides Inhibit Genotype 1b Viral Genome Replication via Interaction with DDX3X , 2010, PloS one.