It is known that eukaryotic ribosomes are able to translate small ORFs and reinitiate translation at downstream start codons. However, this mechanism is widely considered to be inefficient and it is not commonly taken into account. We compiled a sample of human mRNAs containing small upstream ORFs overlapping with annotated protein coding sequences. Statistical analysis supported the hypothesis on reinitiation of translation at downstream AUG codons and functional significance of potential alternative ORFs. It may be assumed that some 5 0 UTR-located upstream ORFs can deliver ribosomes to alternative translation starts, and they should be taken into consideration in the prediction of human mRNA coding potential.
It is known that the recognition of AUG triplet by eukaryotic ribosomes as a translation start site strongly depends on its nucleotide context. However, the relative significance of different context positions is not fully clear. In particular, it concerns the role of 3'-end part of the context located at the beginning of the protein-coding sequence. The significant bias observed in nucleotide frequencies in positions +4, +5, +6 (corresponding to the second codon of CDS) could result from different reasons and their contribution to start codon recognition and initiation of translation is under discussion. In this study, we conducted a comparative computational analysis of the human mRNA samples containing different nucleotides (adenine, guanine or pyrimidine) in the essential context position -3. It was found that the presence of G in position +4 could be important for the context variant GnnAUG but not for AnnAUG. Interestingly, the second position of proteins encoded by mRNAs with AnnAUG context variant was specifically and significantly enriched with serine whereas the presence of GnnAUG context also correlated with a higher occurrence of alanine and glycine. It is likely that the efficiency of translation initiation process can depend on the interplay between 5'-context part, 3'-context part and the type of amino acid in the second position of the encoded protein.
It is widely considered that the vast majority of eukaryotic mRNAs contain only one open reading frame (ORF) and encode single protein. However, eukaryotic ribosomes can initiate translation at alternative start codons due to leaky scanning or reinitiation mechanisms that provides an opportunity to synthesize several protein products. Recent investigations also demonstrated that alternative translation from nonAUG start codons and AUG codons in a weak nucleotide context could make an important contribution to eukaryotic proteomes. However, accurate prediction of alternative start codons demands detailed investigation of mRNA features influencing their recognition by eukaryotic ribosomes. In this work, we present the results of computational analysis of characteristics of yeast and mammalian mRNAs potentially involved in the recognition of nonAUG start codons. It was found that sequence features of nonAUG started Saccharomyces cerevisiae upstream ORFs (uORFs) were adjusted for efficient translation and these uORFs could frequently encode functional polypeptides. In particular, our initial studies revealed that predicted tertiary structures downstream of nonAUG start sites in mammalian mRNAs were energetically more stable than those predicted for AUG start sites with strong Kozak context. We hypothesize that presence of such stable tertiary structure downstream of nonAUG start sites could be an important factor for the ribosome to recognize noncanonical start codons.
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