ANTIBODY TO EPITOPE TAG INDUCES INTERNALIZATION OF HUMAN MUSCARINIC SUBTYPE-1 RECEPTOR

We have studied the effect of an antibody against the epitope EYMPME o n the internalization of the human muscarinic cholinergic receptor hm1 tagged with the epitope at the amino terminus. The antibody to the ta g induces internalization of the hm1 receptor within minutes after exp osure of human embryonic kidney 293 cells transfected with the tagged receptor. This antibody-induced internalization is reversible followin g removal of the antibody. In contrast to hm1 internalization induced by the agonist carbachol, internalization induced by antibody is not b locked by the muscarinic antagonist atropine. The mechanism of antibod y-mediated internalization does not appear to involve receptor dimeriz ation by the antibody, as Fab fragments derived from the antibody also induce internalization. The pathway of antibody-induced internalizati on, similar to the agonist-induced process, is mediated by clathrin-co ated vesicles. Furthermore, antibody treatment does not result in any second messenger production, as measured by phosphoinositide accumulat ion. Our data show that internalization of a G protein-coupled recepto r can be triggered by interaction of the amino terminus of the recepto r with an exogenous ligand and can occur independently of second messe nger production, This result suggests that the receptor can exist in m ultiple conformations, each mediating distinct downstream events.