A knotty problem: Dissecting the molecular mechanics of mRNA recruitment to the eukaryotic ribosome

Abstract

Using a fully reconstituted S. cerevisiae translation initiation system, we are dissecting the molecular mechanics of mRNA recruitment to the 43S pre‐initiation complex (PIC). Efficient recruitment of capped mRNAs requires eIF3 and the eIF4 factors, which stabilize binding of the message and dramatically increase the rate of recruitment. Uncapped mRNAs can be recruited through an alternative pathway that does not require the eIF4 factors. This pathway yields a complex that is unable to effectively locate the start codon. The cap strongly inhibits this alternative mRNA recruitment pathway, imposing a requirement for the eIF4 factors for rapid and stable binding of mRNA to the PIC. Our data suggest that the 5′‐cap serves as both a positive and negative element in mRNA recruitment, promoting initiation in the presence of the full set of mRNA handling factors while preventing binding to the ribosome via an alternative, aberrant pathway requiring only eIF3. Thus the cap ensures that only fully functional pre‐initiation complexes containing all the required factors are assembled on an mRNA.