Intracellular trafficking of angiotensin II and its AT1 and AT2 receptors: evidence for selective sorting of receptor and ligand.
Angiotensin II (Ang II) binds to two different receptor subtypes, AT1 and AT2 receptors. In many cases, receptor stimulation by Ang II is followed by a rapid desensitization of the intracellular signal transduction and a decrease in cell surface receptor number. The present study was designed to examine by immunofluorescence microscopy the cellular trafficking pathways of Ang II and its AT1a and AT2 receptors in human embryonal kidney 293 cells stably expressing these receptor subtypes. Fluorescently labeled Ang II and AT1a receptors were rapidly internalized into endosomes. AT2 receptors were localized in the plasma membrane and did not undergo endocytosis upon agonist stimulation. After removal of agonist, AT1a receptors recycled to the plasma membrane, whereas fluorescently labeled Ang II was targeted to the lysosomal pathway. Even though no further loss of surface receptor was measurable by ligand binding at steady state, fluorescein-Ang II was continuously internalized, and cycling of receptor between endosomal vesicles and the plasma membrane was detected by antibody feeding. These experiments provide evidence for subtype-specific receptor sorting and internalization of Ang II and its AT1a receptor as a receptor-ligand complex, and they suggest that the sequestration of receptors into endosomes is in dynamic equilibrium with receptor cycling to the plasma membrane. Finally, internalization of AT1a receptors and Ang II persists after desensitization mechanisms have attenuated Ca2+ and inositol 1,4,5-trisphosphate signaling.
Hein
L
L
Falk Cardiovascular Research Center and Department of Medicine, Stanford University School of Medicine, California 94305, USA.
Meinel
L
L
Pratt
R E
RE
Dzau
V J
VJ
Kobilka
B K
BK
D000643:Ammonium Chloride / Q000494:pharmacology; D000804:Angiotensin II / Q000378:metabolism*; D000818:Animals; D001692:Biological Transport; D002118:Calcium / Q000378:metabolism; D002462:Cell Membrane / Q000378:metabolism; D002738:Chloroquine / Q000494:pharmacology; D004705:Endocytosis; D000939:Epitopes; D019793:Fluorescein; D005452:Fluoresceins / Q000737:chemistry; D019085:Fluorescent Antibody Technique, Direct; D006801:Humans; D015544:Inositol 1,4,5-Trisphosphate / Q000378:metabolism; D008247:Lysosomes / Q000378:metabolism; D051379:Mice; D044140:Receptor, Angiotensin, Type 1; D044139:Receptor, Angiotensin, Type 2; D011945:Receptors, Angiotensin / Q000187:drug effects / Q000276:immunology / Q000378:metabolism*
D000939:Epitopes; D005452:Fluoresceins; D044140:Receptor, Angiotensin, Type 1; D044139:Receptor, Angiotensin, Type 2; D011945:Receptors, Angiotensin; D000643:Ammonium Chloride; D000804:Angiotensin II; D015544:Inositol 1,4,5-Trisphosphate; D002738:Chloroquine; D002118:Calcium; D019793:Fluorescein