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Laboratory Overview
Our laboratory works on signal transduction at the receptor for the neuropeptide calcitonin gene-related peptide (CGRP). CGRP is one of the most potent vasodilators known, and has been implicated in hypertension, migraine, and chronic pain. Despite the clinical implications of CGRP's biological actions, therapeutic strategies targeting CGRP have been hindered by the lack of an identified CGRP receptor. CGRP binding results in increased cAMP levels in cells and tissue, and a candidate G protein-coupled receptor has been identified called the calcitonin receptor-like receptor (CRLR). However, CRLR was initially non-functional when transfected into mammalian tissue culture cells.
Our laboratory has recently discovered a novel protein required for signal transduction at CGRP receptors which we have named the CGRP-receptor component protein (RCP). RCP is a low molecular weight intracellular membrane-associated protein that couples the CGRP receptor to the cellular signal transduction pathway. We demonstrated this mechanism by making stable cell lines that expressed RCP antisense constructs, and observed a correlation between loss of RCP protein expression and a loss of CGRP-induced cAMP production. However, there was no loss of CGRP binding in the RCP-antisense cells, indicating that RCP did not function as a chaperone protein but was instead coupling the CGRP receptor to the cellular signal transduction pathway. RCP co-immunoprecipitates with the CGRP receptor (CRLR) demonstrating a direct interaction between the two proteins. It has recently been demonstrated that CRLR requires a chaperone protein named receptor activating protein (RAMP1) for routing to the cell surface. Our model for a functional CGRP receptor therefore must include at least three proteins in a complex: the ligand binding protein (CRLR), a chaperone protein (RAMP1), and a coupling protein for signal transduction (RCP). RCP thus represents a novel signal transduction protein, and represents a new level of regulation for signal transduction at G protein-coupled receptors.
Our laboratory is investigating the mechanism of RCP action by: (1) determining the molecular/ biochemical requirements for RCP function, (2) determining the proteins that interact with RCP in a functional receptor complex using yeast two-hybrid strategies, (3) determining the role of RCP and CGRP in vivo, using targeted homologous recombination to generate transgenic RCP knockout mice.