Box 1 | Cap-dependent translation initiation

From the following article:

Structural and mechanistic insights into hepatitis C viral translation initiation

Christopher S. Fraser and Jennifer A. Doudna

Nature Reviews Microbiology 5, 29-38 (January 2007)

doi:10.1038/nrmicro1558

In most cellular mRNAs, recognition of the start site for protein synthesis occurs by ribosome scanning, a proposed model of which is shown in the figure (reviewed in REFS 67,69,87,88,89). This mechanism requires all of the canonical initiation factors that have been characterized so far and, in contrast to the HCV IRES, also requires the 5'-cap structure found on all cellular mRNAs. Translation initiation begins on the 40S (small) ribosomal subunit, which is composed of an mRNA binding cleft, an aminoacyl (A) site, peptidyl (P) site, and exit (E) site. The mRNA enters the 40S ribosomal subunit through the mRNA entry channel, passes through the A, P and E sites and leaves through the mRNA exit channel. To initiate translation, a free pool of 40S ribosomal subunits, stabilized by association with the large multisubunit initiation factor eIF3, binds to Met-tRNAi and mRNA (reviewed in Refs 50,90,91). The Met-tRNAi is brought to the 40S ribosomal subunit as part of an eIF2–GTP complex, and together with two small initiation factors, eIF1 and eIF1A, forms the 43S preinitiation complex. Assembly of the 43S preinitiation complex on an mRNA by this pathway requires the 5'end of the mRNA, as ribosomes are unable to bind an mRNA that has been covalently circularized92. 43S preinitiation complex formation is enhanced by the presence of the cap-binding protein eIF4E, which in turn associates with the scaffold protein eIF4G. The interaction between eIF4G and eIF3 stabilizes the 43S preinitiation complex, leading to its migration (scanning) to the AUG start codon by the unwinding of any RNA structure present in the 5'-untranslated region (UTR). On pairing between the Met-tRNAi anticodon and the AUG start codon, eIF2 hydrolyses its GTP with the help of the GTPase activating protein eIF5. The eIF2–GDP complex dissociates from the 40S ribosomal complex and is recycled by a guanine nucleotide exchange factor, eIF2B, so that it can associate with a new Met-tRNAi and take part in another round of initiation. The exchange of GDP for GTP by eIF2B is highly regulated in eukaryotic cells, by a mechanism involving the phosphorylation of eIF2 by one of at least four distinct kinases, preventing the efficient recycling of this initiation factor90, 93, 94. Finally, a second GTPase, eIF5B, promotes the joining of the 60S ribosomal subunit to form an 80S initiation complex and the removal of the remaining initiation factors (reviewed in Refs 90,93).

Structural and mechanistic insights into hepatitis C viral translation initiation