Protein Synthesis Initiation in Eukaryotic Cells

Abstract

This review summarizes our current understanding of the major pathway for the initiation phase of protein synthesis in eukaryotic cells, with a focus on recent advances. We describe the major scanning or messenger RNA (mRNA) m⁷G cap-dependent mechanism, which is a highly coordinated and stepwise regulated process that requires the combined action of at least 12 distinct translation factors with initiator transfer RNA (tRNA), ribosomes, and mRNAs. We limit our review to studies involving either mammalian or budding yeast cells and factors, as these represent the two best-studied experimental systems, and only include a reference to other organisms where particular insight has been gained. We close with a brief description of what we feel are some of the major unknowns in eukaryotic initiation.

Figure 1.

Overview of the general eukaryotic translation initiation pathway. The pathway for recruiting initiator transfer RNA (tRNA) to the messenger RNA (mRNA) AUG codon in the context of an 80S ribosome (bottom right) is depicted as a series of major steps, labeled with blue text, linked with black arrows. Individual eukaryotic initiation factor (eIF) cartoons and complexes are labeled with black text and nucleotide hydrolysis/inorganic phosphate release reactions are shown by blue arrows. The broad green arrow indicates the direction of scanning toward the AUG codon. The regulatory reactions leading to eIF2 and eIF4E inhibition are shown with plum and red arrows. All steps are described in the main text, starting with eIF2 activation. The timing of release of some factors from initiating ribosomes/mRNA (eIF4F, eIF4B, or eIF3) is not yet clear, so this is not shown.

Figure 2.

Structural models of eukaryotic initiation factor (eIF)3. A composite model of the structure of eIF3 showing the mammalian octamer (right) from PDB 5A5T (des Georges et al. 2015) and the associated eIF3d subunit (PDB 5K4D) (Lee et al. 2016) that is linked via eIF3a to the yeast-like core eIF3bgi complex (PDB 5A5U) (des Georges et al. 2015). The eIF3g is composed of an RNA-recognition motif (RRM) (PDB 2CQ0) and a β-propeller domain similar to eIF3b. eIF3j can interact with the eIF3b RRM. NTD, Amino-terminal domain.

Figure 3.

Model for preinitiation complex (PIC) arrangement at AUG recognition. Three 90° rotated cartoon views of an idealized 48S PIC, modeled on recent structural and functional studies. This model was based on the partial yeast 48S closed complex structure (3JAP) (Llacer et al. 2015) and modified to include findings of others (Luna et al. 2012, 2013; Aylett et al. 2015; des Georges et al. 2015; Simonetti et al. 2016; Obayashi et al. 2017). The entry view (left) shows the eukaryotic initiation factor (eIF)3 yeast-like core subunits, whereas the exit view (right) shows only the mammalian eIF3 octamer complex. eIF2 is shown as semitransparent in the central intersubunit view (middle) to indicate the position of factors otherwise hidden below. For further details of eIF2 interactions, see Figure 4. The position of Rack1 on the ribosome head is shown for orientation purposes only and is not discussed in the text. Factor colors correspond to those used in other figures.

Figure 4.

Altered conformation of initiation factors on AUG recognition. Cartoons derived from cryoelectron microscopy (cryo-EM) analyses of partial preinitiation complexes in a scanning mode (open, left panel) and AUG recognition mode (closed, right panel) (Llacer et al. 2015). Note, in particular, the movement of eukaryotic initiation factor (eIF)1, eIF1A (tail), and eIF2β. eIF3 and part of the 40S head structures were removed for clarity. Images were created with University of California San Francisco (UCSF) Chimera software from PDB files 3JAP and 3JAQ. See text for details.