The long term goals of this proposal are to identify and characterize molecules associated with the Schwann cell membrane which mediate the early stages of glial differentiation. Schwann cell differentiation is characterized by an ordered series of carefully synchronized stages which are induced and promoted by direct neuron-glial contact. Initially, induced Schwann cells proliferate and begin to define territories on the bare axolemma. Subsequent morphological changes result in the extension of glial processes, the ensheathment or myelination of axon(s) and the concomitant organization of a robust basal lamina. Because glial differentiation is crucial for proper neural development and regeneration, experiments are designed to identify and characterize antigens which mediate the early, prerequisite stages of Schwann cell differentiation. The proposed experiments examine the hypothesis that each stage of differentiation is mediated by a novel molecule(s) associated with the Schwann cell membrane. Experiment #1 will evaluate the participation of a Schwann cell membrane-associated antigen, 'C4', in the process of ensheathment. Immunoaffinity methods are used to isolate the antigen for biochemical characterization and use as an immunogen. Polyclonal antibodies are used with newly designed in vitro perturbation studies to functionally disrupt the early stages of Schwann cell differentiation. Experiment #2 utilizes newly developed immunization protocols to identify antigens associated with Schwann cell membranes. Moreover, screening techniques are used to distinguish these antigens which perturb normal glial morphology and/or the ensheathment of axonal processes. Experiment #3 examines the hypothesis that a membrane-associated heparin sulfate proteoglycan mediates the organization of Schwann cell morphology during the ensheathment process. Techniques are used to characterize the cellular location of epitopes and their participation in maintenance of cell morphology and interactions with the environment. Experiment #4 will utilize indirect immunohistochemical methods to compare the time course and cellular distribution of Schwann cell antigens expressed in vitro and in vivo during embryogenesis and regeneration. This proposal will identify molecules that mediate the early states of Schwann cell differentiation and thus begin to characterize the mechanisms by which axons are ensheathed.
