The long range objective of this project is to elucidate the functional role of neural molecules in the development of common nerve pathways and selective axon fasciculation. Towards this end by immunoaffinity purification with the mAb Lan3-2, the investigator and associates have identified a novel member of the L1 family of CAMs, Tractin, which is a multiple domain cleaved protein with several unique features. It contains 6 Ig-domains, 4 FNIII-like domains, an acidic domain, 12 repeats of a novel collagen-like proline- and glycine-rich sequence motif, a transmembrane domain, and an intracellular tail with an ankyrin and a PDZ-domain binding motif. Tractin is expressed by all neurons but is differentially glycosylated with the Lan3-2 and Laz2-369 glycoepitopes only in sets and subsets of peripheral sensory neurons that form specific fascicles in the CNS. In vivo and in vitro antibody perturbation of these glycoepitopes have demonstrated that they can selectively regulate axonal outgrowth and synapse formation. In addition, at least three other mAbs (Lan2-3. Laz6-212, and Laz7-79) which recognize different glycoepitopes specific to distinct subsets of these neurons have been identified. We will test the hypothesis that these glycoepitopes represent additional posttranslational modifications to Tractin and that such differential glycosylation of a widely expressed neural CAM can functionally assist in regulation neuronal outgrowth and synapse formation of distinct neuronal subpopulations. As Tractin is also expressed by all central neurons these findings suggest that Tractin may function as a major regulator of axon fasciculation, neurite extension, and axonal guidance during early nervous system development. The proposed experiments will test this hypothesis and determine the relative contributions of the different domains of Tractin to these processes in vivo and will identify other proteins with which Tractin interacts to mediate these functions. In addition, expression studies in the S2 cell line will provide novel information about the biosynthesis and mechanisms of posttranslational processing of the L1 family CAMs. Mutations in human and murine L1 lead to severe brain abnormalities; however, the causative developmental mechanisms of these brain defects are not well understood and are likely to involve interaction of L1 with extracellular ligands as well as with intracellular signaling pathways linked to cytoskeletal elements. It is therefore of importance to explore the molecular basis for such interactions in various model systems where such interactions are tractable in order to define the range of structural diversity, functions, and signaling capabilities of L1 family CAMs. Thus, these studies will provide valuable new insights into the underlying causes of aberrant neural connections and abnormal brain development.
