Fig. 1. Structure of the efn-4 locus and sequence comparisons. (A) Genetic and physical maps of the efn-4 region. mab-26/efn-4 is located at the left end of linkage group IV. (B) Alignment of the conserved domain of EFN-4 (residues 25-170) with those of other ephrins, showing point mutations; locations of predicted secondary structure elements are based on structures of ephrin B2 (Toth et al., 2001; Himanen et al., 2001). The ephrin with highest similarity to EFN-4 within the conserved domain is EFN-1 (31% identity, 46% similarity). The EFN-4 conserved domain is approximately equally similar in sequence to the ephrin A and ephrin B subclasses of vertebrate ephrins (25-20% identity; sequences shown are human ephrin A1 and ephrin B2). Alignments were made using ClustalW; identities are in black and similarities in gray. Secondary structure elements are aligned according to Himanen et al. (Himanen et al., 2001). The strong allele ju134 affects a non-conserved threonine residue; the weaker allele e660 affects a conserved proline; and the weakest allele, e1746, affects a non-conserved alanine residue. When aligned according to Toth et al. (Toth et al., 2001) ju134 affects an exposed residue of ß-strand D, which may play a role in ephrin dimerization; e660 affects a residue at the end of ß-strand C in the tetramerization interface; and e1746 affects a buried residue between ß-strand D and -helix E. (C) Phylogenetic tree of C. elegans, Drosophila and vertebrate ephrin sequences. Phylogenetic analysis suggests that the C. elegans ephrins diverged after the last common ancestor of vertebrates and nematodes; among the C. elegans ephrins, EFN-4 is the most rapidly evolving. Vertebrate ephrin sequences used are from humans except for ephrin A6 (chick). This tree was generated using a neighbor-joining algorithm (MEGA); bootstrap values are indicated at branches.