The mechanism of CUG-codon reassignment. The scheme contrasts the presence of tRNA_CAGs and evolution of CUG codons according to the tRNA loss driven codon reassignment hypothesis with assumptions based on the codon capture and ambiguous intermediate theories. Only the most probable sequence of events is shown for each hypothesis. Alternative but less likely scenarios are presented in Supplemental Figure S12. Mixed models, for example, ambiguous decoding of the CUG codons in the ancestor of the SP and CP clades followed by loss of the ${\text{tRNA}}_{\mathrm{CAG}}^{\mathrm{Leu}}$ in the ancient Pachysolen and capture by the ${\text{tRNA}}_{\mathrm{CAG}}^{\mathrm{Ala}}$, seem extremely unlikely and are not shown. The codon capture theory distinguishes from the tRNA loss driven codon reassignment hypothesis mainly by the order of events (tRNA loss after or before the reduction of CUG codons, respectively), the degree of CUG-codon loss (loss of all codons vs. a reduction of CUG-codon usage, respectively), and the cause of reduction of CUG codons (AT pressure vs. tRNA loss and decreased decoding fidelity, respectively). In contrast to the ambiguous intermediate hypothesis, the tRNA loss driven codon reassignment theory does not assume any ambiguous decoding of the CUG codons. In addition, capture of the CUG codon by different tRNAs is an elemental characteristic of the tRNA loss driven codon reassignment theory and does not need to be split into independent events.