RAMP-altered class B GPCR hormone binding and signaling

SUMMARY/ABSTRACT: Three receptor activity-modifying proteins (RAMP1-3) in humans form heterodimers with the class B G protein-coupled receptors (GPCRs) calcitonin receptor (CTR) and calcitonin-like receptor (CLR) and modulate their responses to six CGRP family peptide hormones. This complex system of seven receptors and six ligands is a model for understanding class B GPCR signaling mechanisms and modulation of GPCR function by accessory membrane proteins. CGRP family peptide signaling through CTR, CTR- RAMP1/2/3, or CLR-RAMP1/2/3, is involved in diverse processes including calcium homeostasis, blood glucose regulation, pain signaling, neuroimmune communication, and development and maintenance of the cardiovascular and lymphatic systems. Consequently, the receptors are drug targets for osteoporosis, diabetes, obesity, migraine headache, and cardiovascular and lymphatic disorders. We have used biochemistry, pharmacology, and structural biology approaches to study how the peptides bind and activate the receptors, how the RAMPs determine ligand selectivity, and how the peptide-bound receptors couple to G protein transducer to promote cAMP signaling. Our studies advanced our understanding of each of these areas and enabled us to engineer novel peptide antagonists and agonists with useful properties like increased receptor affinity, altered receptor selectivity, or long-acting signaling. These have been powerful tools for studying the receptors and may have promise as therapeutic leads. More recently, we discovered temporal differences in the signaling properties of the CGRP family peptides that likely contribute to their diverse biology. Some of the peptides promote cAMP signaling of short duration, whereas others elicit long duration signaling. Despite these advances, several areas need more study. How RAMPs determine ligand selectivity is only partially understood. Our understanding of the kinetics of peptide binding and cAMP signaling is incomplete, and the molecular basis of long duration signaling is unclear. Our understanding of the interactions of the receptors with the two other families of transducer proteins that mediate GPCR signaling, the GPCR kinases (GRKs) and b-arrestins, is rudimentary. Moreover, there is still a need for better pharmacological tools for studying the receptors. Accordingly, our goals are to further define the mechanisms by which RAMPs determine CTR and CLR ligand selectivity, further characterize the kinetics of the molecular events leading to cAMP signaling, and define the basis for long duration cAMP signaling. In addition, we will continue to engineer novel peptide ligands and we will study the receptor interactions with GRK and b-arrestin transducers.