Figure 2. Phylogenetic analysis reveals that motifs for ankyrin-dependent axonal clustering evolved sequentially, first in chordates (Na_V channels), then in jawed vertebrates (Kv7.2 and Kv7.3).

(A) Phylogram (minimal evolution) of Na_V channels, showing that all vertebrate channels are derived from chordate NaV1. (B) Clustal alignment of Na_V channel DII–DIII loop sequences, showing presence of anchor motifs in chordate NaV1 and all vertebrate channels. The anchor motifs are boxed (red). Shading indicates each residue’s conservation within a set of 28 chordate and non-chordate (not shown) aligned Na_V sequences: bins represent ≤10, 11–20, 21–30, 31–45, 46–60, and 61–100 % conservation. (C) Phylogeny of KCNQ channels, based on analysis of derived amino acid sequences encoded on exons 5–7. Novel genes identified or cloned (see [12]) are shown in red. The branch marking the inferred first appearance of the KCNQ2/3 anchor motif is shown in red. (D) Alignment of KCNQ2 and KCNQ3 C-terminal intracellular sequences near the anchor motifs. Break (vertical black line) indicates location of 5–8 omitted, poorly conserved residues. The derived Kv7.2/Kv7.3 anchor motif sequences (red boxed region) are similar but non-identical to those of chordate Na_V genes. Otherwise no similarity to the Na_V DII–DIII loop sequence shown in B is present. Shading indicates conservation within the 7 KCNQ sequences aligned: shades represent ≤15, 15–30, 31–45, 46–60, 61–75, 76–90, and 91–100% conservation. In A and C, nodes are labeled with bootstrap values (>50%), nodes associated with gene duplications have red dots, and scale bars indicate substitutions per residue. After [12].