Members of the
netrin gene family have been identified in vertebrates, Drosophila and Caenorhabditis elegans and found to encode secreted molecules involved in axon guidance. Here I use the conserved function of
netrins in triploblasts, coupled with the phylogenetic position of amphioxus (the closest living relative of the vertebrates), to investigate the evolution of an axon guidance cue in chordates. A single amphioxus
netrin gene was isolated by PCR and cDNA library screening and named
AmphiNetrin. The predicted
AmphiNetrin protein showed high identity to other
netrin family members but differed in that the third of three
EGF repeats found in other
netrins was absent. Molecular phylogene-
tic analysis showed that despite the absent
EGF repeat
AmphiNetrin is most closely related to the vertebrate
netrins.
AmphiNetrin expression was identified in embryonic notochord and floor plate, a pattern similar to that of vertebrate
netrin-1 expression.
AmphiNetrin expression was also identified more widely in the posterior larval brain, and in the anterior extension of the notochord that underlies the anterior of the amphioxus brain. All of these areas of expression are correlated with developing axon trajectories: The floor plate with ventrally projecting somatic motor neurons and Rohde cell projections, the posterior brain with the ventral commissure and primary motor centre and the anterior extension of the notochord with ventrally projecting neurons associated with the median eye. Amphioxus is naturally cyclopaedic and also lacks the ventral brain cells that the induction of which results in the splitting of the vertebrate eye field and, when missing, result in cyclopaedia. These cells normally express
netrins required for developing axon tracts in the brain, and the expression of
AmphiNetrin in the anterior extension of the notochord underlying the brain may explain how amphioxus is able to maintain ventral guidance cues while lacking these cells.