STRUCTURAL-ANALYSIS OF A HUMAN GLIAL VARIANT LAMININ

Astrocytes secrete laminin-like molecules in culture and may represent a major source of laminin in the developing central nervous system, g et these laminins have not been extensively characterized. We previous ly reported the presence of an astrocyte-derived variant laminin in me dia conditioned by human U251 MG astrocytoma cells. This laminin was p artially purified in a highly anionic Mono Q fraction with strong adhe sion activity for fibroblasts and glial cells (Aukhil et al. (1990) Ma trix 10: 98-111). We now show that glial laminin could be dissociated from an anionic species, perhaps an similar to 400-kDa keratan sulfate proteoglycan present in the preparation, by a second round of Mono Q anion exchange chromatography in the presence of 6 M urea. Cell adhesi on activity remained tightly associated with laminin-containing fracti ons, suggesting that glial laminin was responsible for the adhesion ac tivity in the original preparation. Immunochemical and SDS-PAGE gel an alyses of laminin heterotrimers demonstrated that glial laminin contai ned the beta 2 and gamma 1 chains in disulfide-bonded heterotrimeric c omplexes with a 360-kDa chain, a 320-kDa chain, or a postulated simila r to 200-kDa chain. While these chains were not recognized by antibodi es directed against the alpha 1-, alpha 2-, or alpha 3-related laminin chains, rotary shadowed glial laminin molecules appeared to contain a lpha chains, as judged by the presence of an apparent G-domain termina ting the long arm of each laminin molecule. These findings suggest tha t glial laminin contains one or more variant alpha chains, perhaps rel ated to one of the more recently described alpha chains, alpha 3B, alp ha 4, or alpha 5. Together our results implicate human U251 MG glial l aminin as a previously uncharacterized laminin isoform with strong adh esive activity for fibroblasts and glial cells. (C) 1996 Academic Pres s, Inc.