The Role of ephrin-A1/EphA4 signaling in Enteric Neural Crest Cell Migration

Abstract

Enteric neural crest cells (ENCCs) migrate and colonize the gut during enteric nervous system (ENS) development, in which Ret and p75 support ENCC survival, proliferation, migration and differentiation. In nervous system development, cell-cell interaction is regulated by Eph/ephrin signaling that induces attraction or repulsion between cells, crucial to cell segregation and thus cell migration. However, the role of Eph/ephrin signaling in regulating ENCC migration remains unknown. Mutation of Ephs and ephrins have been reported in Hirschsprung disease (HSCR) patients, suggesting the involvement of Eph/ephrin signaling. I examined the role of ephrin-A1/EphA4 signaling using 293T cells to investigate their interaction and regulation of cell segregation. Immunoprecipitation shows that ephrin-A1 can interact with Ret and p75. My results also show that ephrin-A1 mutant protein is able to interact with Ret and p75, and display ligand-receptor binding interaction with EphA4. Cell segregation assay using stably-transfected cells shows that ephrin-A1 mutant-expressing cells demonstrate enhanced cell segregation when co-cultured with EphA4-expressing cells. These findings suggest that ephrin-A1 mutant could disrupt cell segregation regulated by ephrin-A1/EphA4 signaling, while interaction of ephrin-A1 with Ret and p75 suggests that it potentially plays an important role in this cell-cell interaction to regulate ENCC migration.