Figure 1.

ORFs do not adequately represent translational complexity. (A) Two formal definitions of ORFs. Three reading frames are shown as horizontal bars, with vertical bars corresponding to AUG (green) or stop codons (black). Several examples of start-stop (green arcs) and stop-stop (black arcs) ORFs are shown. (B–D) The relationship between ORFs (top) and expressed proteoforms (bottom) for mRNAs with different locations of starts and stops (middle). Only two relevant reading frames are shown for simplicity. (B) An RNA encoding two proteoforms with alternative N termini owing to utilization of two start codons. Because of multiple potential AUG codons, there are many start-stop ORFs whose conceptual translation does not correspond to encoded proteoforms. A stop-stop ORF does not reflect the existence of alternative proteoforms. (C) In the case of stop codon read-through or selenocysteine insertion, ribosomes may read-through specific stop codons by incorporating an amino acid, yielding a product that cannot be described as a product of a single ORF. (D) Similarly, ribosomal frameshifting generates a trans-frame protein (blue) that does not represent a product of a single ORF.