Recruitment of a Nicotinic Acetylcholine Receptor Mutant Lacking Cytoplasmic Tyrosine Residues in Its β Subunit into Agrin-Induced Aggregates

Abstract

During synaptogenesis at the vertebrate skeletal neuromuscular junction, acetylcholine receptors (AChRs) form high-density aggregates opposite the presynaptic terminal in response to nerve-derived agrin. Agrin has been shown to stimulate tyrosine phosphorylation of a muscle-specific receptor tyrosine kinase MuSK and of the AChR β subunit, and tyrosine kinase inhibitors and a tyrosine kinase-deficient mutant of MuSK prevent AChR aggregation. To evaluate the role of tyrosine phosphorylation of the AChR β subunit in receptor aggregation, we replaced all three putative cytoplasmic tyrosine residues of the AChR β subunit with phenylalanine residues and expressed the mutant receptors in cultured myotubes. Upon agrin treatment, transfected myotubes formed AChR aggregates that contained receptors with mutant β subunits. Thus, AChRs can be recruited into agrin-induced specializations by protein–protein interactions that do not depend on tyrosine phosphorylation of the AChR β subunit.