The agrin receptor. Localization in the postsynaptic membrane, interaction with agrin, and relationship to the...

Agrin is a component of the synaptic basal lamina that induces the aggregation of acetylcholine receptors (AChRs) and other elements of the postsynaptic membrane. We have determined the localization, binding characteristics, and biochemical profile of the agrin receptor in Torpedo electric organ membranes and defined domains of agrin that bind this receptor. Postsynaptic membranes from Torpedo electric organ bind agrin as judged by depletion of AChR clustering activity from solution. A ligand-based radioimmunoassay shows that agrin binding to postsynaptic membranes is saturable and calcium-dependent. Half-maximal binding is observed at agrin concentrations < or = 10(-10) M. Identification of the bound agrin polypeptides shows that at least one membrane binding domain of agrin is located in a 70-kDa proteolytic fragment. Immunofluorescent visualization and radioimmunoassay of agrin binding demonstrates that the agrin receptor is selectively concentrated in postsynaptic membranes, with little binding detected on nonsynaptic or liver membranes. Agrin binding is greatly reduced if the membranes are pretreated with trypsin, but is unaffected by phosphatidylinositol-specific phospholipase C. Membranes stripped of peripheral proteins by alkaline treatment retain full ligand binding capacity. alpha-Bungarotoxin affinity columns bind AChRs but not agrin receptors. The ratio of agrin receptors to AChRs in postsynaptic membranes is approximately 1:200. We conclude that the agrin receptor is an integral membrane glycoprotein that is selectively concentrated in postsynaptic membranes, but that is not tightly complexed with the AChR. The results also indicate that the biological activity of agrin is mediated through intracellular signal transduction events triggered by ligand binding to the agrin receptor.