Directed neuronal migration is an essential feature of the developing nervous system, during which undifferentiated cells are guided through a changing environment of membrane-bound, matrix-associated, and diffusible cues. Despite extensive investigations into the nature of pathway cues that can affect migratory behavior, the molecular mechanisms that regulate neuronal migration have remained enigmatic. This issue can now be addressed using the enteric nervous system (ENS) of the moth, Manduca sexta. The formation of the ENS requires the migration of an identified population of 300 neurons (the EP cells) along the visceral musculature; unlike other preparations, both the neurons and their muscle band pathways remain directly accessible to manipulation throughout development. Surgical manipulations in vivo have shown that band-specific cues are both necessary and sufficient for EP cell migration. Recently, one candidate pathway molecule has been identified as fasciclin II (MFas II), a member of the immunoglobulin superfamily of homophilic cell adhesion receptors. MFas II is transiently expressed by both the EP cells and their muscle band pathways at the time of migration, while perturbations of MFas II in embryonic culture cause a marked inhibition of migration. Probes against the two major isoforms of MFas II (prepared from full-length clones that have been isolated from a cDNA library) will be used to characterize the developmental expression of the MFas II gene during pathway formation and EP cell migration. A combination of sense and antisense probes, monoclonal and polyclonal antibodies, and synthetic peptides directed against MFas II will then be used to determine its precise functional role in regulating EP cell migration within the developing embryo. A primary cell culture preparation will then be used to define the effects of exogenous MFas II on neuronal migratory behavior in vitro. The onset of EP cell migration also coincides with enhanced tyrosine kinase activity, and MFas II has now been shown to be closely associated with protein tyrosine kinases. Accordingly, the in vivo and in vitro preparations of the EP cells will be used to investigate whether MFas II -dependent aspects of migration require the activation of specific tyrosine kinases. These experiments will clarify the manner in which MFas II affects EP cell migration during embryogenesis, and should lend insight into how similar molecular mechanisms regulate this type of migratory behavior in more complex systems, as well.
