Internalization of beta-adrenergic receptor in A431 cells involves non-coated vesicles.
To characterize the mechanism of internalization of beta-adrenergic catecholamine receptors on human epidermoid A431 carcinoma cells, their distribution was analyzed by immunocytochemistry using the monoclonal anti-receptor antibody BRK2. In preconfluent cultures, the receptors appeared to be randomly distributed on the cell surface. Exposure to the agonist isoproterenol induced an overall decrease in the number of cell surface receptors as determined by binding experiments and visualized by immunofluorescence. When cells were incubated at 4 degrees C with BRK2 and anti-mouse IgG-gold and then transferred at 37 degrees C, non-coated invaginations and vesicles were labeled. The addition of isoproterenol resulted in an increased rate of internalization of the receptor-BRK2-anti-IgG-gold complex. When incubation with the two antibody reagents was prolonged (with or without isoproterenol), non-coated vesicles fused in the endosomal compartment, and receptors were transferred to multivesicular bodies and lysosomes. At no stage in this process was there any indication that clathrin-coated pits or vesicles participated. Furthermore, we found that an intracellular potassium depletion treatment known to inhibit endocytosis, did not affect the normal pattern of desensitization of beta-adrenergic receptors.
Raposo
G
G
Laboratory of Electron Microscopy, Institut Jacques Monod, University of Paris VII, CNRS, Paris, France.
Dunia
I
I
Delavier-Klutchko
C
C
Kaveri
S
S
Strosberg
A D
AD
Benedetti
E L
EL
D000911:Antibodies, Monoclonal / Q000032:analysis; D004705:Endocytosis; D005455:Fluorescent Antibody Technique; D006801:Humans; D007150:Immunohistochemistry; D007545:Isoproterenol / Q000494:pharmacology; D008247:Lysosomes / Q000032:analysis / Q000378:metabolism; D008854:Microscopy, Electron; D011188:Potassium / Q000494:pharmacology; D011943:Receptors, Adrenergic, beta / Q000032:analysis / Q000378:metabolism*; D014407:Tumor Cells, Cultured
D000911:Antibodies, Monoclonal; D011943:Receptors, Adrenergic, beta; D007545:Isoproterenol; D011188:Potassium