CARBOXYL-TERMINAL DOMAINS DETERMINE INTERNALIZATION AND RECYCLING CHARACTERISTICS OF BOMBESIN RECEPTOR CHIMERAS

To investigate the role of the carboxyl terminus in the regulation of the bombesin (BN) receptor, we constructed two chimeric receptors with carboxyl termini transferred from either m3 muscarinic cholinergic (m 3 ACh) (BMC) or cholecystokinin A (CCKA) (BCC) receptors and expressed them in Chinese hamster ovary cells, Previous studies showed that ago nist treatment caused rapid internalization of CCKA but not m3 ACh rec eptors in these cells, In the current study we conducted separate anal yses of ligand and receptor internalization and analyzed receptor recy cling. Ligand internalization was assessed using acid washing. BN and CCKA receptors internalized ligand with 80 +/- 3 and 85 +/- 7% in an a cid-resistant compartment at equilibrium, Ligand internalization of ch imeric receptors generally assumed the properties of the donor recepto rs. Thus, BCC receptors internalized ligand to a similar extent as wil d-type CCKA receptors (75 +/- 3%), whereas, BMC receptors showed reduc ed ligand internalization (38 +/- 1%), Receptor internalization was mo re directly assessed by determining agonist-induced loss of surface bi nding. BN and CCKA receptors were largely internalized (56 +/- 8 and 5 0 +/- 7%, respectively). BCC receptors were also extensively internali zed (82 +/- 3%). In contrast, BMC receptors were minimally internalize d (22 +/- 8%), Receptor recycling was assessed as recovery from agonis t induced loss of binding, BN, CCKA, and BMC receptors showed rapid re cycling, In contrast, BCC receptors did not recycle, These data indica te that carboxyl-terminal structures determine both internalization of ligand-receptor complexes and subsequent receptor recycling.