THE ALPHA-CHAIN OF LAMININ-1 IS INDEPENDENTLY SECRETED AND DRIVES SECRETION OF ITS BETA-CHAIN AND GAMMA-CHAIN PARTNERS

A mammalian recombinant strategy,vas established to dissect rules of b asement membrane laminin assembly and secretion, The alpha-, beta-, an d gamma-chain subunits of laminin-1 were expressed in all combinations , transiently and/or stably, in a near-null background. In the absence of its normal partners, the alpha chain was secreted as intact protei n and protein that had been cleaved in the coiled-coil domain. In cont rast, the beta and gamma chains, expressed separately or together, rem ained intracellular with formation of beta beta or beta gamma, but not gamma gamma, disulfide-linked dimers, Secretion of the beta and gamma chains required simultaneous expression of all three chains and their assembly into alpha beta gamma heterotrimers. Epitope-tagged recombin ant alpha subunit and recombinant laminin were affinity-purified from the conditioned medium of alpha gamma and alpha beta gamma clones, Rot ary-shadow electron microscopy revealed that the free alpha subunit is a linear structure containing N-terminal and included globules with a foreshortened long arm, while the trimeric species has the typical fo ur-arm morphology of native laminin, We conclude that the alpha chain can be delivered to the extracellular environment as a single subunit, whereas the beta and gamma chains cannot, and that the alpha chain dr ives the secretion of the trimeric molecule, Such an alpha-chain-depen dent mechanism could allow for the regulation of laminin export into a nascent basement membrane, and might serve an important role in contr olling basement membrane formation.