Inference of Sec transitions. (a) Sec-to-Cys transition. To infer such a transition in a SelP sequence, we required the presence of two closest outgroup sequences and a sister SelP sequence containing Sec. (b) Cys-to-Sec transition. To infer gain of Sec from Cys, we required two outgroup sequences and one sister sequence to contain Cys. (c) Loss of Sec. If Sec was present in two outgroup and one sister sequence, whereas the corresponding position in the tested SelP sequence had an amino acid other than Cys or Sec, the inference was Sec loss. (d) Sec/Cys transitions viewed at the level of amino acids and codons. Numbers indicate the number of transitions between indicated amino acids or codons. The size of arrows highlights directionality of these transitions.