Multiple ramp domains are required for generation of amylin receptor phenotype from the calcitonin receptor gene product

Calcitonin (CT), calcitonin gene-related peptide (CGRP), amylin, and adreno medullin constitute a family of structurally related peptides that signal v ia either the calcitonin receptor-like receptor or the CT receptor, with re ceptor phenotype determined by coexpression of one of the three receptor ac tivity-modifying proteins (RAMPs). The nature of the interaction between th e receptor and RAMP was investigated using chimeras between RAMP1 and RAMP2 where the amino-terminal domain of RAMP1 was attached to the transmembrane domain and carboxy terminus of RAMP2 and called RAMP1/2, and vice versa fo r RAMP2/1, Cotransfection of wild-type or chimeric RAMPs with the insert-ne gative isoform of the human CT receptor (hCTR(I1-)) into COS-7 cells result ed in the expression of I-125-rat amylin binding sites. Highest specific bi nding was observed when either RAMP1 or RAMP2/1 were cotransfected, indicat ing the importance of the RAMP transmembrane domain and/or carboxy terminus for the degree to which amylin receptors are expressed. In contrast, the p henotype generated was primarily determined by the amino terminus, with sim ilar RAMP1- and RAMP1/2-induced receptor phenotypes that had higher affinit y for human CGRP alpha and lower affinity for human calcitonin than the RAM P2- and RAMP2/1-induced receptors. (C) 2000 Academic Press.