Distinct heparin-binding and neurite-promoting properties of laminin isoforms isolated from chick heart.
Laminin isolated from chick heart is composed of several heterotrimeric variants of 800 and 700 kDa. Here, we used monoclonal antibodies against chick laminin to purify different laminin isoforms from this mixture. Antibody 8D3 specifically removed laminin containing alpha 2 chain from chick heart laminin preparations, leaving behind 700 kDa variants. Using antibody C4 against the laminin beta 2 chain, alpha 2 chain containing variants were further separated into alpha 2 beta 1 gamma 1 and alpha 2 beta 2 gamma 1 laminin, respectively. Laminins containing alpha 2 chain and recognized by antibody 8D3 are cross-shaped molecules. Their expression during embryogenesis is tightly regulated. In 5-day embryos staining with monoclonal antibody 8D3 is restricted to the dermamyotome. Older embryos (8 days) express alpha 2 chain containing variants at myotendinous junction primordia of skeletal muscle, and only late in development these variants are generally expressed in skeletal and heart muscle basement membranes. The 700 kDa laminin variants contain beta 1, beta 2, and gamma 1 subunits affiliated with an immunologically distinct, shorter alpha x chain and appear to be T-shaped in the electron microscope. Whereas laminins with an alpha 2 subunit bind to heparin, variants with the novel alpha x chain do not. Experiments using cultured sympathetic neurons showed that laminins with alpha x chain are less potent than alpha 2 chain containing variants in promoting neurite outgrowth. In contrast, sympathetic neurons cannot discriminate between alpha 2 beta 1 gamma 1 and alpha 2 beta 2 gamma 1 laminin substrates, respectively, and show identical high rates of neurite formation.
Department of Biophysical Chemistry, University of Basel, Switzerland.
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R
Brandenberger
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M
Chiquet
D000818:Animals; D000911:Antibodies, Monoclonal; D002478:Cells, Cultured; D002642:Chick Embryo; D002645:Chickens; D002846:Chromatography, Affinity; D006493:Heparin / Q000378:metabolism*; D007797:Laminin / Q000302:isolation & purification / Q000378:metabolism*; D008970:Molecular Weight; D009206:Myocardium / Q000737:chemistry*; D009414:Nerve Growth Factors / Q000302:isolation & purification / Q000378:metabolism*; D016501:Neurites / Q000502:physiology*; D010446:Peptide Fragments / Q000302:isolation & purification; D011485:Protein Binding; D013329:Structure-Activity Relationship
D000911:Antibodies, Monoclonal; D007797:Laminin; D009414:Nerve Growth Factors; D010446:Peptide Fragments; D006493:Heparin