Nicotinic acetylcholine receptor is internalized via a Rac-dependent, dynamin-independent endocytic pathway.
Endocytosis of the nicotinic acetylcholine receptor (AChR) is a proposed major mechanism of neuromodulation at neuromuscular junctions and in the pathology of synapses in the central nervous system. We show that binding of the competitive antagonist alpha-bungarotoxin (alphaBTX) or antibody-mediated cross-linking induces the internalization of cell surface AChR to late endosomes when expressed heterologously in Chinese hamster ovary cells or endogenously in C2C12 myocytes. Internalization occurs via sequestration of AChR-alphaBTX complexes in narrow, tubular, surface-connected compartments, which are indicated by differential surface accessibility of fluorescently tagged alphaBTX-AChR complexes to small and large molecules and real-time total internal reflection fluorescence imaging. Internalization occurs in the absence of clathrin, caveolin, or dynamin but requires actin polymerization. alphaBTX binding triggers c-Src phosphorylation and subsequently activates the Rho guanosine triphosphatase Rac1. Consequently, inhibition of c-Src kinase activity, Rac1 activity, or actin polymerization inhibits internalization via this unusual endocytic mechanism. This pathway may regulate AChR levels at ligand-gated synapses and in pathological conditions such as the autoimmune disease myasthenia gravis.
Kumari
Sudha
S
National Centre for Biological Sciences, Tata Institute of Fundamental Research, Bangalore, India.
Borroni
Virginia
V
Chaudhry
Ashutosh
A
Chanda
Baron
B
Massol
Ramiro
R
Mayor
Satyajit
S
Barrantes
Francisco J
FJ
D000199:Actins / Q000378:metabolism; D000818:Animals; D000906:Antibodies / Q000494:pharmacology; D001692:Biological Transport / Q000187:drug effects; D002038:Bungarotoxins / Q000494:pharmacology; D002451:Cell Compartmentation / Q000187:drug effects; D002460:Cell Line; D003432:Cross-Linking Reagents / Q000494:pharmacology; D015536:Down-Regulation / Q000187:drug effects; D034281:Dynamins / Q000037:antagonists & inhibitors / Q000378:metabolism; D004705:Endocytosis / Q000187:drug effects; D011992:Endosomes / Q000187:drug effects / Q000378:metabolism; D020868:Gene Silencing / Q000187:drug effects; D007700:Kinetics; D008247:Lysosomes / Q000187:drug effects / Q000378:metabolism; D051379:Mice; D010766:Phosphorylation / Q000187:drug effects; D016392:Proto-Oncogene Proteins pp60(c-src) / Q000378:metabolism; D011978:Receptors, Nicotinic / Q000378:metabolism*; D020744:rac GTP-Binding Proteins / Q000378:metabolism*
D000199:Actins; D000906:Antibodies; D002038:Bungarotoxins; D003432:Cross-Linking Reagents; D011978:Receptors, Nicotinic; D016392:Proto-Oncogene Proteins pp60(c-src); D020744:rac GTP-Binding Proteins; D034281:Dynamins