Translation Elongation and Recoding in Eukaryotes.

Translation Elongation and Recoding in Eukaryotes Thomas E. Dever,1 Jonathan D. Dinman,2 and Rachel Green3 1 Eunice Kennedy Shriver National Institute of Child Health and Human Development, National Institutes of Health, Bethesda, Maryland 20892 2 Department of Cell Biology and Molecular Genetics, University of Maryland, College Park, Maryland 20742 3 Department of Molecular Biology and Genetics, Johns Hopkins University School of Medicine, Baltimore, Maryland 21205 Correspondence: ; ; In this review, we highlight the current understanding of translation elongation and recoding in eukaryotes. In addition to providing an overview of the process, recent advances in our understanding of the role of the factor eIF5A in both translation elongation and termination are discussed. We also highlight mechanisms of translation recoding with a focus on ribosomal frameshifting during elongation. We see that the balance between the basic steps in elongation and the less common recoding events is determined by the kinetics of the different processes as well as by specific sequence determinants. OVERVIEW OF TRANSLATION ELONGATION T he mechanism of translation elongation is conserved in all kingdoms of life. Whereas most of the mechanistic details of the process have been elucidated in studies of bacterial translation (see Rodnina 2018), the key steps are shared between eukaryotes and bacteria. In eukaryotes, translation initiation culminates with formation of an 80S initiation complex in is bound in the P ( pepwhich Met-tRNAMet i tidyl) site of the ribosome. The anticodon of is base-paired with the start the Met-tRNAMet i codon of the messenger RNA (mRNA), and the second codon of the open reading frame (ORF) is in the A (aminoacyl) site of the ribosome. Elongation commences with delivery of the cognate elongating aminoacyl-tRNA (transfer RNA) to the A site of the ribosome (Fig. 1). The eukaryotic translation elongation factor eEF1A, like its bacterial ortholog EF-Tu, is activated upon binding guanosine triphosphate (GTP) and forms a ternary complex upon binding an aminoacyl-tRNA. The eEF1A•GTP•aminoacyl-tRNA complex binds in the A site. Basepairing interactions between the anticodon of the aminoacyl-tRNA and the A-site codon trigger GTP hydrolysis by eEF1A. The eEF1A•GDP complex is released and the aminoacyl-tRNA is accommodated into the A site. High-resolution cryo-electron microscopy (EM) structures of decoding complexes have provided insights into how the bacterial and eukaryotic ribosomes sense proper decoding (Jobe et al. 2018). The 18S ribosomal RNA (rRNA) Editors: Michael B. Mathews, Nahum Sonenberg, and John W.B. Hershey Additional Perspectives on Translation Mechanisms and Control available at www.cshperspectives.org Copyright © 2018 Cold Spring Harbor Laboratory Press; all rights reserved; doi: 10.1101/cshperspect.a032649 Cite this article as Cold Spring Harb Perspect Biol 2018;10:a032649 1 T.E. Dever et al. Aminoacyl-tRNA EF1A EPA EF1A Aminoacyl-tRNA binding eEF1B EF2 EF1A Deacyl-tRNA EF1A EPA Translocation EF2 A-site tRNA accommodation EPA Hypusine eIF5A EPA EF2 Peptide bond formation Diphthamide EPA Figure 1. Model of the eukaryotic translation elongation pathway. At the top, an eEF1A•GTP•aminoacyl-tRNA (transfer RNA) ternary complex binds to the A (aminoacyl) site of an 80S ribosome with the anticodon loop of the tRNA in contact with the messenger RNA (mRNA). Following GTP hydrolysis and release of an eEF1A•GDP binary complex, the aminoacyl-tRNA is accommodated into the A site, and the eEF1A•GDP is recycled to eEF1A•GTP by the exchange factor eEF1B. During catalysis of peptide bond formation, the A- and P ( peptidyl)-site tRNAs shift into hybrid states with the acceptor ends of the tRNAs moving to the P and E sites, respectively. Substrate positioning for peptide bond formation is aided by binding of the factor eIF5A and its hypusine modification (green) in the E site. Following peptide bond formation, the factor eEF2•GTP with its diphthamide modification (magenta) binds in the A site and promotes translocation of the tRNAs into the canonical P and E sites. Following release of the deacylated tRNA from the E site, the next cycle of elongation commences with binding of the appropriate eEF1A•GTP•aminoacyl-tRNA to the A site. Throughout, GTP is depicted as a green ball and GDP as a red ball; also, the large ribosomal subunit (light blue) is displayed transparently to enable visualization of the tRNAs, factors, and mRNA bound to the decoding center at the interface between the large and small subunits and of tRNAs, interacting with the peptidyl transferase center in the large subunit. Note, however, that the positions of the mRNA, tRNAs, and factors are drawn for clarity and are not meant to specify their exact places on the ribosome. helix h44 residues A1824 and A1825 in mammalian (rabbit) ribosomes (A1755 and A1756 in Saccharomyces cerevisiae and A1492 and A1493 in Escherichia coli, respectively) as well as the residue G626 in rabbit ribosomes (G577 in S. cerevisiae and G530 in E. coli) interact with the minor groove of the codon–anticodon helix and stabilize A-site tRNA binding by hydrogen 2 bonding (Ogle et al. 2001; Shao et al. 2016; Loveland et al. 2017). Interestingly, when flipped out of helix 44 (h44), the residues A1824 and A1825 (or A1492 and A1493 in bacteria) interact with the first two codon pairs in the codon–anticodon duplex, enabling the +3 position to participate in wobble interactions related to the degeneracy of the genetic code (Loveland et al. 2017). Cite this article as Cold Spring Harb Perspect Biol 2018;10:a032649 Translation Elongation and Recoding As the h44 residues also flip out to interact with mispaired codon–anticodon helices formed with near-cognate tRNAs in the A site (Demeshkina et al. 2012), it has been proposed that the interaction of G626 (G530 in bacteria) may perform a more crucial function as the latching nucleotide that fixes the codon–anticodon helix in the decoding center of the ribosome (Loveland et al. 2017). In addition to providing insights into decoding, the recent structures of eukaryotic ribosomal complexes have provided insights into GTPase activation of eEF1A as well as of eRF3 in termination complexes and of Hbs1 in ribosome rescue complexes (Shao et al. 2016). In these structures, interactions between the sarcin-ricin loop of the large ribosomal subunit and the Switch 2 loop of the GTPase domains helps position the catalytic His residue to promote GTP hydrolysis (Shao et al. 2016). Moreover, the amino terminus of the eukaryotespecific ribosomal protein eS30 becomes ordered upon cognate codon–anticodon interaction in the A site, and a conserved His residue inserts into the decoding center to form potentially stabilizing contacts (Shao et al. 2016). These novel interactions may contribute to the reported enhanced accuracy of eukaryotic versus bacterial elongation (Kramer et al. 2010). Fin (...truncated)