Laminins bearing the alpha-2 subunit play central roles in basement membrane assembly and in the differentiation and functional maintenance of skeletal muscle, neuromuscular junction, and peripheral nerve. These roles are thought to depend upon laminin polymerization and anchorage to plasmamembranes. Our recent studies identify plasma membrane sulfatides as critical mediators of anchorage in Schwann cells, enabling basement membrane assembly and laminin-initiated signaling. Inactivation of binding of these glycosphingolipids to the LG modules of laminins-1 and -2 prevents basement membrane assembly in Schwann cells, while intercalation of sulfatides into fibroblasts, which normally lack these extracellular matrices (ECMs), enable formation of functionally-active basement membranes. Sulfatide expression is detected in Schwann cell surfaces in developing peripheral nerve prior to basement membrane assembly, and de-sulfation of sulfatide in DRG neuron-Schwann cell co-cultures prevents basement membrane assembly and nerve myelination. Collectively, these data support the hypothesis that sulfatide-laminin LG modular interactions are important for development of peripheral nerve. Armed with a growing repertoire of recombinant alpha-2 laminin heterotrimers and interactive fragments, and the ability to manipulate cell competency for basement membrane assembly, we propose to dissect molecular mechanisms initiated by neuromuscular laminins using tractable models of Schwann cell ECM assembly and peripheral nerve myelination. In particular, we propose to examine laminin assembly and cellular responses requiring anchorage to sulfatides (aim 1), to dissect laminin alpha-2 LG modular roles ant evaluate laminin-8 and laminin-10 contributions to Schwann cell (aim 2), and study the biological role of neuromuscular laminins and sulfatides in Schwann cell myelination (aim 3). ? ?
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