Initiation of translation in the majority of eukaryotic cellular mRNAs depends on the 5'-cap (m7GpppN) and involves ribosomal scanning of the 5' untranslated region (5'-UTR) for an initiating AUG start codon. Therefore, this mechanism is often called cap-dependent translation initiation. Proximity to the cap, as well as the nucleotides surrounding an AUG codon, influence the efficiency of the start site recognition during the scanning process. However, if the recognition site is poor enough, scanning ribosomal subunits will ignore and skip potential starting AUGs, a phenomenon called leaky scanning. Leaky scanning allows a single mRNA to encode several proteins that differ in their amino-termini. Merrick (2010) provides an overview of this process and hghlights several features of it that remain incompletely understood.
Several eukaryotic cell and viral mRNAs initiate translation by an alternative mechanism that involves internal initiation rather than ribosomal scanning. These mRNAs contain complex nucleotide sequences, called internal ribosomal entry sites, where ribosomes bind in a cap-independent manner and start translation at the closest downstream AUG codon.
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Once the Met-tRNAi has recognized the AUG, eIF2-bound GTP is hydrolyzed. The reaction is catalyzed by eIF5 (or eIF5B) and is thought to cause dissociation of all other initiation factors and allow joining of the large 60S ribosomal subunit. Release of the initiation factors from 40S leaves the Met-tRNAi in the ribosomal P-site base-paired to the start codon on the mRNA.
The ternary complex (Met-tRNAi:eIF2:GTP) binds to the complex formed by the 40S subunit, eIF3 and eIF1A, to form the 43S complex. eIF1A promotes binding of the ternary complex to the 40S subunit within 43S. The initiator methionyl-tRNA from the ternary complex is positioned at the ribosomal P site.
eIF2B is a guanine nucleotide releasing factor that is required to cause GDP release so that a new GTP molecule can bind and activate eIF2, so that it can be reused.
The precise order of events leading to the circularization of poly (A) mRNA during translation initiation is unknown. Here the association of PABP with the poly (A) mRNA and the association of PABP with eIF4F are represented as occuring simultaneously after formation of the initiation complex. However, it is also possible that these interactions occur during the formation of the translation initiation complex. The binding of eIF4F to the cap and binding of PABP to the poly (A) tail, for example, may occur at the same time. In fact, the eIF4G-PABP interaction helps eIF4F to bind tighter to the cap (Borman et al. 2000.) In addition, eIF4B and eIF4H bind more transiently to the mRNA and may not be part of an initial complex in which PABP has not yet touched eIF4G.
The mRNA-bound ribosomal complex moves along the 5'-untranslated region (5'-UTR) of the mRNA from its initial site to the initiation codon to form a 48S complex, in which the initiation codon (AUG) is base paired to the anticodon of the Met-tRNAi. It is not known whether eIF4A (or another ATPase, such as DED1) facilitates scanning by melting mRNA secondary structures or by actively propelling the ribosome.
The DEAD-box RNA helicase eIF4A, together with the RNA-binding proteins eIF4B or eIF4H, is thought to unwind RNA secondary structures near the 5'-end of the mRNA and in the presence of ATP.
Although the mechanism through which L13a prevents translation initiation has not been determined, Mazumder et al. (2003) have described four alternatives. L13a could (1) inhibit the function of eIF4F, (2) block the recruitment of the 43S preinitiation complex, (3) prevent scanning of the 43S complex to the initiation codon, or 4) interfere with joining of the 60S ribosomal subunit.
The L13a subunit of the 60s ribosome is phosphorylated about 16 hours after INF gamma induction by an unknown kinase. At this time, L13a is also released from the 60s subunit (Mazumder et al.,2003). It is unclear, however, whether phosphorylation occurs before or after the release of L13a. Here, phosphorylation is shown as occurring after release.
The translation initiation complex forms when the 43S complex binds the mRNA that is associated with eIF4F, eIF4B and eIF4H. eIF4G in the eIF4F complex can directly contact eIF3 in the 43S complex. eIF1A is necessary for the formation of this complex.
Once the 60S subunit joins the translation initiation complex, eIF5B hydrolyzes its GTP and is released from the now 80S monosome. The fully assembled 80s ribosome is now ready to start elongation of the polypeptide chain.
The AUG initiation codon in the mRNA is recognized by base pairing with the anticodon of the Met-tRNAi. This reaction requires eIF1, eIF1A, eIF2 and eIF5.
The factor eIF4E within the eIF4F (cap-binding) complex directly binds the 5'-cap on eukaryotic mRNAs. Note that the mRNA is in complex with cytoplasmic proteins constituting an mRNP complex.
The L13a subunit of the 60s ribosome is phosphorylated about 16 hours after INF gamma induction by an unknown kinase. At this time, L13a is also released from the 60s subunit (Mazumder et al.,2003). It is unclear, however, whether phosphorylation occurs before or after the release of L13a. Here, phosphorylation is shown as occurring after release.
Several eukaryotic cell and viral mRNAs initiate translation by an alternative mechanism that involves internal initiation rather than ribosomal scanning. These mRNAs contain complex nucleotide sequences, called internal ribosomal entry sites, where ribosomes bind in a cap-independent manner and start translation at the closest downstream AUG codon.
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DataNodes
Met-tRNAi
mRNAeIF3
eIF1AeIF4F eIF4B eIF4H
PABPmRNA eIF4F eIF4B
eIF4HMet-tRNAi mRNA eIF5B
GTPMet-tRNAi
mRNAeIF4F eIF4B
eIF4HGDP
eIF2BeIF4F eIF4B
eIF4HAnnotated Interactions
Met-tRNAi
mRNAMet-tRNAi
mRNAeIF3
eIF1AeIF4F eIF4B eIF4H
PABPeIF4F eIF4B eIF4H
PABPmRNA eIF4F eIF4B
eIF4HmRNA eIF4F eIF4B
eIF4HMet-tRNAi
mRNAeIF4F eIF4B
eIF4HGDP
eIF2BeIF4F eIF4B
eIF4HeIF4F eIF4B
eIF4H