DETERMINANTS OF [ARG(2)]PTH-(1-34) BINDING AND SIGNALING IN THE TRANSMEMBRANE REGION OF THE PARATHYROID-HORMONE RECEPTOR

Previously, we reported that [Arg(2)]PTH-(1-34) bound to the rat osteo sarcoma cell line, ROS 17/2.8, with 2-fold higher apparent affinity th an it did to the opossum kidney cell line, OK, yet the analog was only a weak partial agonist for cAMP stimulation with ROS 17/2.8 cells, wh ereas it was a full cAMP agonist with OK cells. These results suggeste d that the rat and opossum PTH receptors differ in a region recognized by the hormone's amino-terminus. In this report we show that the clon ed PTH receptors derived from ROS 17/2.8 and OK cells, expressed in CO S-7 cells, also displayed altered responses to [Arg(2)] PTH-(1-34). Th us, [Arg(2)]PTH-(1-34) bound to the cloned rat PTH receptor with 7-fol d higher affinity than it did to the cloned opossum PTH receptor, and in cAMP stimulation assays, it was a much weaker agonist with the rat receptor than it was with the opossum receptor. Studies with rat/oposs um PTH receptor chimeras suggested that the membrane-spanning region o f the receptor contributed to the different binding and signaling resp onses to [Arg(2)]PTH-(1-34). Point mutation analysis identified three sites in or near the extracellular ends of transmembrane domains V and VI, which specifically affected [Arg(2)] PTH-(1-34) binding and signa ling.