title: Controlling signaling with a specifically designed Gi-coupled receptor.
abstract: We are developing a system to control G protein signaling in vivo to regulate a broad range of physiologic responses. Our system utilizes G protein-coupled peptide receptors engineered to respond exclusively to synthetic small molecule ligands and not to their natural ligand(s). These engineered receptors are designated RASSLs (receptor activated solely by a synthetic ligand). We have made two prototype RASSLs that are based on the human kappa opioid receptor. Small molecule drugs that activate the kappa receptor are nonaddictive and safe to administer in vivo. Binding and signaling assays reveal 200-2000-fold reductions in the ability of our RASSLs to bind or be activated by dynorphin, an endogenous peptide ligand of the kappa opioid receptor. In a high-throughput signaling assay, these prototype RASSLs expressed in Chinese hamster ovary K1 cells showed little or no response to a panel of 21 opioid peptides but still signaled normally in response to small molecule drugs such as spiradoline. Activation of a RASSL by spiradoline also caused proliferation of rat-1a tissue culture cells. These data provide evidence that G protein-coupled receptors can be made into RASSLs. The potential in vivo applications for RASSLs include the positive enrichment of transfected cells and the development of new animal models of disease.
authors.affiliation: Gladstone Institute for Cardiovascular Disease, Departments of Medicine and Pharmacology, University of California, San Francisco, CA 94141-9100, USA.
authors.forename: P
authors.initials: P
authors.lastname: Coward
authors.forename: H G
authors.initials: HG
authors.lastname: Wada
authors.forename: M S
authors.initials: MS
authors.lastname: Falk
authors.forename: S D
authors.initials: SD
authors.lastname: Chan
authors.forename: F
authors.initials: F
authors.lastname: Meng
authors.forename: H
authors.initials: H
authors.lastname: Akil
authors.forename: B R
authors.initials: BR
authors.lastname: Conklin
mesh_terms: D000262:Adenylyl Cyclases / Q000378:metabolism; D000595:Amino Acid Sequence; D000818:Animals; D001667:Binding, Competitive; D016466:CHO Cells; D019556:COS Cells; D006224:Cricetinae; D015195:Drug Design; D006801:Humans; D008969:Molecular Sequence Data; D009294:Narcotics / Q000378:metabolism; D017433:Protein Structure, Secondary; D017473:Receptors, Opioid, kappa / Q000737:chemistry / Q000378:metabolism*; D011993:Recombinant Fusion Proteins / Q000737:chemistry; D015398:Signal Transduction
chemical_list: D009294:Narcotics; D017473:Receptors, Opioid, kappa; D011993:Recombinant Fusion Proteins; C110260:Ro1 protein, synthetic; C110261:Ro2 protein, recombinant; D000262:Adenylyl Cyclases