Figure 2. N-linked glycosylation of innexins is highly conserved in chordate animals.

(A) Maximum likelihood phylogeny of innexin proteins from 276 non-chordate species (n=867 innexins). Red branches represent innexins with N-glycosylation consensus sites (NGS) in the extracellular loops (ELs), respectively. Note that each innexin in every chordate species has at least one NGS. The red bars in the inner circle indicate whether extracellular loop 1 or 2 (EL1 and EL2) contains the NGS. The colors of the outer circle represent the phylum classification of the innexins. (B) Fraction of innexins with NGSs in their ELs in the taxonomic groups shown in (A). Multiple sequence alignments of the EL1 of innexins in (C) lancelets and (D) tunicates. Note that lancelets and tunicates have more than two cysteine residues in EL1, like cnidarians and ctenophores. (E) Multiple sequence alignments of the EL2 of innexins in lampreys. Representative multiple sequence alignments of the highly conserved EL2 of (F) Pannexin 1 and the EL1 of (G) Pannexin 2 and (H) Pannexin 3 in vertebrates. Residues that are conserved in all innexins in each alignment are highlighted (*, absolutely conserved; +, physicochemical properties are conserved). Note that the pannexin alignments only contain the sequences of some representative species of each taxonomic group. All identified chordate innexins as well as the complete alignment are provided as supplementary files (Figure 2—source data 2 and Figure 2—source data 3). The cysteine residues (yellow) and the NGSs (red) in each EL are colored. The identified NGSs in EL1 and EL2 of the innexins of all other chordate species are shown in Figure 2—source data 1.