Pushing the limits of the scanning mechanism for initiation of translation

Abstract

Selection of the translational initiation site in most eukaryotic mRNAs appears to occur via a scanning mechanism which predicts that proximity to the 5' end plays a dominant role in identifying the start codon. This "position effect" is seen in cases where a mutation creates an AUG codon upstream from the normal start site and translation shifts to the upstream site. The position effect is evident also in cases where a silent internal AUG codon is activated upon being relocated closer to the 5' end. Two mechanisms for escaping the first-AUG rule--reinitiation and context-dependent leaky scanning--enable downstream AUG codons to be accessed in some mRNAs. Although these mechanisms are not new, many new examples of their use have emerged. Via these escape pathways, the scanning mechanism operates even in extreme cases, such as a plant virus mRNA in which translation initiates from three start sites over a distance of 900 nt. This depends on careful structural arrangements, however, which are rarely present in cellular mRNAs. Understanding the rules for initiation of translation enables understanding of human diseases in which the expression of a critical gene is reduced by mutations that add upstream AUG codons or change the context around the AUG(START) codon. The opposite problem occurs in the case of hereditary thrombocythemia: translational efficiency is increased by mutations that remove or restructure a small upstream open reading frame in thrombopoietin mRNA, and the resulting overproduction of the cytokine causes the disease. This and other examples support the idea that 5' leader sequences are sometimes structured deliberately in a way that constrains scanning in order to prevent harmful overproduction of potent regulatory proteins. The accumulated evidence reveals how the scanning mechanism dictates the pattern of transcription--forcing production of monocistronic mRNAs--and the pattern of translation of eukaryotic cellular and viral genes.

Deleterious mutation in FDX1L gene is associated with a novel mitochondrial muscle myopathy

“…When a mutation alters the start codon, the scanning mechanism should activate initiation from the next downstream ATG. 12 The next putative in frame start codon is present only 14 codons downstream coding for Met5, however, this methionine residue completely lacks the surrounding Kozak sequence, making it very unlikely alternative initiation site. In agreement with the predicted deleterious effect of the mutation on the translation of the protein, immunoblot assay of the patients' muscle and fibroblasts demonstrated lack of Fdx2 band as compared with controls ( Figure 3a).…”

The amino terminus extension in the long dipeptidyl peptidase 9 isoform contains a nuclear localization signal targeting the active peptidase to the nucleus

“…examples for leaky scanning were reported for several proteins including human-insulin-degrading enzyme (IDe) [44] and human neuropeptide Y [45]. Importantly, leaky scanning occurs if the upstream atG is arranged within a poor match for a Kozak consensus, allowing the translation to initiate also from a second down-stream start codon (for review see [46]). however, the first aUG codon encoding for DPP9-l is arranged within a strong context, meaning a purine (G/a) at position −3 and a G at position +4, arguing against this possibility.…”

Starting the protein synthesis machine: eukaryotic translation initiation

“…We refer the reader to these articles, for an alternate take on the topic and a more comprehensive citation of the primary research literature. (1,(12)(13)(14)(15)(16) Formation of the 43S preinitiation complex Physiological conditions favour the association of 40S and 60S ribosomal subunits to form complete 80S ribosomes. Thus, the first requirement for initiation is to promote a dissociation of vacant ribosomes into their subunits.…”

“…A linear movement along the mRNA 5 0 UTR in search of an AUG triplet, termed 'scanning', was originally proposed by Kozak (16,52) (Fig. 1).…”

“…mammals, recognition of an AUG as a translation start codon also critically depends on its surrounding sequence. The consensus sequence GCC(A/G)CCAUGG (positions where changes have the most pronounced effect are marked in bold) provides an optimal context for initiation (16,(52)(53)(54) Experiments using highly pure or recombinant mammalian initiation factors and ribosomal subunits to reconstitute initiation intermediates have now shed more light on the elusive scanning process. Primer extension by reverse transcriptase is arrested at the leading edge of mRNA-associated ribosomal complexes ('toe-printing'), providing information about their presence and position on the mRNA.…”