Antibody to epitope tag induces internalization of human muscarinic subtype 1 receptor.
We have studied the effect of an antibody against the epitope EYMPME on the internalization of the human muscarinic cholinergic receptor hm1 tagged with the epitope at the amino terminus. The antibody to the tag induces internalization of the hm1 receptor within minutes after exposure of human embryonic kidney 293 cells transfected with the tagged receptor. This antibody-induced internalization is reversible following removal of the antibody. In contrast to hm1 internalization induced by the agonist carbachol, internalization induced by antibody is not blocked by the muscarinic antagonist atropine. The mechanism of antibody-mediated internalization does not appear to involve receptor dimerization by the antibody, as Fab fragments derived from the antibody also induce internalization. The pathway of antibody-induced internalization, similar to the agonist-induced process, is mediated by clathrin-coated vesicles. Furthermore, antibody treatment does not result in any second messenger production, as measured by phosphoinositide accumulation. Our data show that internalization of a G protein-coupled receptor can be triggered by interaction of the amino terminus of the receptor with an exogenous ligand and can occur independently of second messenger production. This result suggests that the receptor can exist in multiple conformations, each mediating distinct downstream events.
Tolbert
L M
LM
Department of Biopharmaceutical Sciences and Pharmaceutical Chemistry, University of California, San Francisco 94143-0446, USA.
Lameh
J
J
D000906:Antibodies / Q000276:immunology*; D002460:Cell Line; D004705:Endocytosis / Q000502:physiology*; D000939:Epitopes / Q000235:genetics / Q000276:immunology*; D006801:Humans; D018613:Microscopy, Confocal; D009154:Mutation / Q000502:physiology; D010446:Peptide Fragments / Q000276:immunology; D011976:Receptors, Muscarinic / Q000235:genetics / Q000276:immunology / Q000502:physiology*; D015290:Second Messenger Systems / Q000502:physiology; D016324:Sequence Tagged Sites
D000906:Antibodies; D000939:Epitopes; D010446:Peptide Fragments; D011976:Receptors, Muscarinic