The muscle fiber is ensheathed by a layer of extracellular matrix material, the basal lamina (BL). The part of the BL which runs through the synaptic cleft between the nerve and muscle, and which differs immunochemically from extrasynaptic BL, plays important roles in the regeneration, maintenance, and function of the neuromuscular junction (NMJ) in the adult. One of these synaptic BL proteins is S-laminin. It is a homolog of the Bl chain of the laminin trimer, and it contains a tripeptide sequence which is specifically adhesive for motoneurons. At the synapse, S-laminin associates with the laminin B2 and A chains, while the B1 chain of laminin is localized to extrasynaptic regions of the muscle. In cultured muscle cells, S-laminin assembles into a trimer with the B2 and A chains, replacing Bl. Surprisingly, there is evidence for two immunochemically distinct populations in cultured muscle cells. The long-term objectives are to learn: (a) how the S-laminin trimer is assembled and localized to the synapse, and (b) how the heterogeneity is generated and if it is related to the function of S-laminin. To do this, the amino acid sequences in S-laminin responsible for localization to the synapse, specific assembly with the laminin B2 and A chains, and the heterogeneity will be determined by construction and expression of interchain cDNA chimeras between S-laminin and laminins Bl or B2. Specific sequences in S-laminin involved in these processes will be determined by site-directed mutagenesis. The cDNA for S-laminin will be expressed either in cultured muscle cells which contain endogenous laminin, or in cells which do not express laminin. In the latter case, cDNAs for the other laminin chains will also be expressed. Functional studies will be done using cultured motor neuron-like cells. The mechanisms involved in forming and maintaining the NMJ are poorly understood. our knowledge of these mechanisms would be increased by determining how S-laminin is assembled at and localized to the postsynaptic apparatus. This in turn would have relevance to the study of neuromuscular diseases.
