DISSECTION OF COMPLEX MOLECULAR-INTERACTIONS OF NEUROFASCIN WITH AXONIN-1, F11, AND TENASCIN-R, WHICH PROMOTE ATTACHMENT AND NEURITE FORMATION OF TECTAL CELLS

Neurofascin is a member of the L1 subgroup of the Ig superfamily that promotes axon outgrowth by interactions with neuronal NgCAM-related ce ll adhesion molecule (NrCAM), We used a combination of cellular bindin g assays and neurite outgrowth experiments to investigate mechanisms t hat might modulate the interactions of neurofascin. In addition to NrC AM, we here demonstrate that neurofascin also binds to the extracellul ar matrix glycoprotein tenascin-R (TN-R) and to the Ig superfamily mem bers axonin-1 and F11. Isoforms of neurofascin that are generated by a lternative splicing show different preferences in ligand binding. Whil e interactions of neurofascin with F11 are only slightly modulated, bi nding to axonin-1 and TN-R is strongly regulated by alternatively spli ced stretches located in the NH2-terminal half, and by the proline-ala nine-threonine-rich segment. In vitro neurite outgrowth and cell attac hment assays on a neurofascin-Fc substrate reveal a shift of cellular receptor usage from NrCAM to axonin-1, F11, and at least one additiona l protein in the presence of TN-R, presumably due to competition of th e neurofascin-NrCAM interaction. Thereby, F11 binds to TN-R of the neu rofascin/TN-R complex, but not to neurofascin, whereas axonin-1 is not able to bind directly to the neurofascin/TN-R complex as shown by com petition binding assays, In conclusion, these investigations indicate that the molecular interactions of neurofascin are regulated at differ ent levels, including alternative splicing and by the presence of inte racting proteins.