title: Crosslinking-induced endocytosis of acetylcholine receptors by quantum dots.
abstract: In a majority of patients with myasthenia gravis (MG), anti-acetylcholine receptor (AChR) antibodies target postsynaptic AChR clusters and thus compromise the membrane integrity of neuromuscular junctions (NMJs) and lead to muscle weakness. Antibody-induced endocytosis of AChRs in the postsynaptic membrane represents the initial step in the pathogenesis of MG; however, the molecular mechanisms underlying AChR endocytosis remain largely unknown. Here, we developed an approach to mimic the pathogenic antibodies for inducing the crosslinking and internalization of AChRs from the postsynaptic membrane. Using biotin-α-bungarotoxin and quantum dot (QD)-streptavidin, cell-surface and internalized AChRs could be readily distinguished by comparing the size, fluorescence intensity, trajectory, and subcellular localization of the QD signals. QD-induced AChR endocytosis was mediated by clathrin-dependent and caveolin-independent mechanisms, and the trafficking of internalized AChRs in the early endosomes required the integrity of microtubule structures. Furthermore, activation of the agrin/MuSK (muscle-specific kinase) signaling pathway strongly suppressed QD-induced internalization of AChRs. Lastly, QD-induced AChR crosslinking potentiated the dispersal of aneural AChR clusters upon synaptic induction. Taken together, our results identify a novel approach to study the mechanisms of AChR trafficking upon receptor crosslinking and endocytosis, and demonstrate that agrin-MuSK signaling pathways protect against crosslinking-induced endocytosis of AChRs.
authors.affiliation: Division of Life Science, State Key Laboratory of Molecular Neuroscience, The Hong Kong University of Science & Technology, Clear Water Bay, Hong Kong, China ; Department of Physiology, Yong Loo Lin School of Medicine, National University of Singapore, Singapore.
authors.forename: Chi Wai
authors.initials: CW
authors.lastname: Lee
authors.affiliation: Division of Life Science, State Key Laboratory of Molecular Neuroscience, The Hong Kong University of Science & Technology, Clear Water Bay, Hong Kong, China.
authors.forename: Hailong
authors.initials: H
authors.lastname: Zhang
authors.affiliation: Division of Life Science, State Key Laboratory of Molecular Neuroscience, The Hong Kong University of Science & Technology, Clear Water Bay, Hong Kong, China ; Department of Physiology, Faculty of Medicine, The University of Hong Kong, Pok Fu Lam, Hong Kong, China.
authors.forename: Lin
authors.initials: L
authors.lastname: Geng
authors.affiliation: Division of Life Science, State Key Laboratory of Molecular Neuroscience, The Hong Kong University of Science & Technology, Clear Water Bay, Hong Kong, China.
authors.forename: H Benjamin
authors.initials: HB
authors.lastname: Peng
mesh_terms: D018171:Agrin / Q000378:metabolism; D000818:Animals; D002966:Clathrin / Q000378:metabolism; D003432:Cross-Linking Reagents / Q000737:chemistry / Q000494:pharmacology*; D004705:Endocytosis / Q000187:drug effects*; D051379:Mice; D008870:Microtubules / Q000187:drug effects / Q000378:metabolism; D010316:Particle Size; D045663:Quantum Dots / Q000737:chemistry; D020794:Receptor Protein-Tyrosine Kinases / Q000378:metabolism; D011950:Receptors, Cholinergic / Q000737:chemistry* / Q000378:metabolism*; D015398:Signal Transduction / Q000187:drug effects; D013569:Synapses / Q000187:drug effects / Q000378:metabolism
chemical_list: D018171:Agrin; D002966:Clathrin; D003432:Cross-Linking Reagents; D011950:Receptors, Cholinergic; C511695:MuSK protein, mouse; D020794:Receptor Protein-Tyrosine Kinases