title: Regulation of muscle acetylcholine receptor turnover by β subunit tyrosine phosphorylation.
abstract: At the neuromuscular junction (NMJ), the postsynaptic localization of muscle acetylcholine receptor (AChR) is regulated by neural signals and occurs via several processes including metabolic stabilization of the receptor. However, the molecular mechanisms that influence receptor stability remain poorly defined. Here, we show that neural agrin and the tyrosine phosphatase inhibitor, pervanadate slow the degradation of surface receptor in cultured muscle cells. Their action is mediated by tyrosine phosphorylation of the AChR β subunit, as agrin and pervandate had no effect on receptor half-life in AChR-β(3F/3F) muscle cells, which have targeted mutations of the β subunit cytoplasmic tyrosines. Moreover, in wild type AChR-β(3Y) muscle cells, we found a linear relationship between average receptor half-life and the percentage of AChR with phosphorylated β subunit, with half-lives of 12.7 and 23 h for nonphosphorylated and phosphorylated receptor, respectively. Surprisingly, pervanadate increased receptor half-life in AChR-β(3Y) myotubes in the absence of clustering, and agrin failed to increase receptor half-life in AChR-β(3F/3F) myotubes even in the presence of clustering. The metabolic stabilization of the AChR was mediated specifically by phosphorylation of βY390 as mutation of this residue abolished β subunit phosphorylation but did not affect δ subunit phosphorylation. Receptor stabilization also led to higher receptor levels, as agrin increased surface AChR by 30% in AChR-β(3Y) but not AChR-β(3F/3F) myotubes. Together, these findings identify an unexpected role for agrin-induced phosphorylation of β(Y390) in downregulating AChR turnover. This likely stabilizes AChR at developing synapses, and contributes to the extended half-life of AChR at adult NMJs.
authors: Department of Physiology and Membrane Biology, University of California Davis, Davis, CA, USA.
authors: John B
authors: JB
authors: Rudell
authors: Michael J
authors: MJ
authors: Ferns
mesh_terms: D018171:Agrin / Q000502:physiology*; D000818:Animals; D002462:Cell Membrane / Q000378:metabolism; D002478:Cells, Cultured / Q000187:drug effects / Q000378:metabolism; D006207:Half-Life; D051379:Mice; D008822:Mice, Transgenic; D018485:Muscle Fibers, Skeletal / Q000378:metabolism; D009124:Muscle Proteins / Q000378:metabolism; D018482:Muscle, Skeletal / Q000166:cytology / Q000378:metabolism*; D016297:Mutagenesis, Site-Directed; D009469:Neuromuscular Junction / Q000378:metabolism; D010766:Phosphorylation; D019000:Phosphotyrosine / Q000378:metabolism; D025941:Protein Interaction Mapping; D011499:Protein Processing, Post-Translational; D055550:Protein Stability; D021122:Protein Subunits; D011978:Receptors, Nicotinic / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D011993:Recombinant Fusion Proteins / Q000235:genetics / Q000378:metabolism; D013569:Synapses / Q000378:metabolism; D014638:Vanadates / Q000494:pharmacology
chemical_list: D018171:Agrin; C495964:Chrnb1 protein, mouse; D009124:Muscle Proteins; D021122:Protein Subunits; D011978:Receptors, Nicotinic; D011993:Recombinant Fusion Proteins; C051609:peripheral membrane protein 43K; C076715:pervanadate; D019000:Phosphotyrosine; D014638:Vanadates