Evidence for transmembrane anchoring of extracellular matrix at acetylcholine receptor clusters.
Clusters of nicotinic acetylcholine receptor (AChR) in cultured rat myotubes are organized into rectilinear arrays of receptor-rich and receptor-poor domains. Extracellular matrix (ECM) molecules, including fibronectin, heparan sulfate proteoglycan, laminin, and type IV collagen, codistribute with AChR in these clusters. We have examined the stability of this association. We disrupted the AChR clusters in intact myotubes with sodium azide, an energy metabolism inhibitor, and with culture medium free of Ca2+. We also altered or extracted proteins from detergent-isolated AChR clusters by treating with buffers of low ionic strength or alkaline pH or with insoluble chymotrypsin. Each of these treatments dispersed AChR clusters and, simultaneously, caused fibronectin, heparan sulfate proteoglycan, laminin, and type IV collagen to disperse from AChR-rich strips of membrane. Control experiments indicated that insoluble chymotrypsin had no direct effect on the ECM at AChR clusters. It did, however, remove spectrin and the receptor-associated 58-kDa protein from the cytoplasmic surface of receptor clusters. Thus, the ECM at AChR clusters is disrupted by an agent acting at the cytoplasmic surface of the membrane. We discuss the possibility that both AChR and ECM are bound to a common membrane skeleton and the implications this may have for synaptogenesis.
Department of Neurology, University of Maryland School of Medicine, Baltimore 21201.
G M
GM
Dmytrenko
R J
RJ
Bloch
D000818:Animals; D000831:Animals, Newborn; D002462:Cell Membrane / Q000502:physiology; D002478:Cells, Cultured; D046508:Culture Techniques / Q000379:methods; D003598:Cytoskeletal Proteins / Q000032:analysis / Q000378:metabolism; D005109:Extracellular Matrix / Q000502:physiology*; D016326:Extracellular Matrix Proteins / Q000032:analysis / Q000378:metabolism; D005353:Fibronectins / Q000302:isolation & purification / Q000378:metabolism; D006863:Hydrogen-Ion Concentration; D009132:Muscles / Q000166:cytology / Q000502:physiology*; D009994:Osmolar Concentration; D051381:Rats; D017207:Rats, Sprague-Dawley; D011950:Receptors, Cholinergic / Q000378:metabolism*
D003598:Cytoskeletal Proteins; D016326:Extracellular Matrix Proteins; D005353:Fibronectins; D011950:Receptors, Cholinergic