Actin at receptor-rich domains of isolated acetylcholine receptor clusters.
Acetylcholine receptor (AChR) clusters of cultured rat myotubes, isolated by extraction with saponin (Bloch, R. J., 1984, J. Cell Biol. 99:984-993), contain a polypeptide that co-electrophoreses with purified muscle actins. A monoclonal antibody against actin reacts in immunoblots with this polypeptide and with purified actins. In indirect immunofluorescence, the antibody stains isolated AChR clusters only at AChR domains, strips of membrane within clusters that are rich in receptor. It also stains the postsynaptic region of the neuromuscular junction of adult rat skeletal muscle. Semiquantitative immunofluorescence analyses show that labeling by antiactin of isolated analyses show that labeling by antiactin of isolated AChR clusters is specific and saturable and that it varies linearly with the amount of AChR in the cluster. Filaments of purified gizzard myosin also bind preferentially at AChR-rich regions, and this binding is inhibited by MgATP. These experiments suggest that actin is associated with AChR-rich regions of receptor clusters. Depletion of actin by extraction of isolated clusters at low ionic strength selectively releases the actin-like polypeptide from the preparation. Simultaneously, AChRs redistribute within the plane of the membrane of the isolated clusters. Similarly, brief digestion with chymotrypsin reduces immunofluorescence staining and causes AChR redistribution. Treatments that deplete AChR from clusters in intact cells also reduce immunofluorescent staining for actin in isolated muscle membrane fragments. Upon reversal of these treatments, cluster reformation occurs in regions of the membrane that also stain for actin. I conclude that actin is associated with AChR domains and that changes in this association are accompanied by changes in the organization of isolated AChR clusters.
D000199:Actins / Q000302:isolation & purification* / Q000378:metabolism; D000818:Animals; D000831:Animals, Newborn; D000906:Antibodies; D000936:Antigen-Antibody Complex; D004591:Electrophoresis, Polyacrylamide Gel; D008970:Molecular Weight; D009132:Muscles / Q000166:cytology / Q000378:metabolism*; D009218:Myosins / Q000378:metabolism; D051381:Rats; D011919:Rats, Inbred Strains; D011950:Receptors, Cholinergic / Q000302:isolation & purification*
D000199:Actins; D000906:Antibodies; D000936:Antigen-Antibody Complex; D011950:Receptors, Cholinergic; D009218:Myosins