GaL1 (Ga14) Couples the Opioid Receptor-Like1 Receptor to Stimulation of Phospholipase C

In most tissues and cells the opioid receptor-like (ORL1) receptor regulates effectors primarily through the pertussis toxin (PTX)-sensitive guanine nucleotide-binding regulatory proteins (G proteins) Gi/Go. Many Gi-coupled receptors possess additional capability to interact with one or more PTX-insensitive G proteins. Using the bg-induced stimulation of type 2 adenylyl cyclase as a readout, we screened the ability of ORL1 receptor to interact with a panel of PTX-insensitive G proteins. In the presence of PTX, activation of the ORL1 receptor resulted in the stimulation of type 2 adenylyl cyclase only in HEK 293 cells coexpressing the a subunit of Gz, G12, G14, or G16, but not in cells coexpressing G11, G13, or Gq. Coupling to both Gz and G16 was expected because close relatives of the ORL1 receptor, the opioid receptors, are known to couple productively to these G proteins. ORL1 receptor coupling to either G12 or G14 has not been demonstrated. As predicted by the type 2 adenylyl cyclase assays, activation of the ORL1 receptor resulted in the formation of inositol phosphates in COS-7 cells transiently cotransfected with Ga14. The ORL1 receptor-mediated stimulation of phospholipase C was found to be Ga14 dependent, agonist dose dependent, ligand selective, and PTX insensitive. We conclude that G14 can link the ORL1 receptor to regulation of phopholipase C. The opioid receptor-like (ORL1) receptor is a guanine nucleotide-binding regulatory protein (G protein)-coupled receptor whose complementary DNA sequence bears substantial homology to those of the opioid receptors (Mollereau et al., 1994). Despite its resemblance to opioid receptors, the ORL1 receptor does not bind any of the known opioid ligands with high affinity and appears to regulate distinct physiological functions. The endogenous ligand for the ORL1 receptor has been identified as a novel heptadecapeptide termed nociceptin (Meunier et al., 1995), or orphanin FQ (Reinscheid et al., 1995; henceforth referred to as nociceptin/OFQ). Nociceptin/OFQ has been shown to regulate nociception (Meunier et al., 1995) and may also be involved in numerous physiological processes as diverse as blood pressure (Champion et al., 1997) and feeding (Pomonis et al., 1996). The diverse function of the ORL1 receptor must invariably be mediated via the heterotrimeric (abg) signal-transducing G proteins. Like opioid receptors, the ORL1 receptor uses the pertussis toxin (PTX)-sensitive Gi/Go proteins to inhibit adenylyl cyclase (Mollereau et al., 1994) and to regulate the activity of ion channels (Connor et al., 1996; Mattes et al., 1996). The ORL1 receptor can thus be considered as a multifunctional “Gicoupled” receptor. It has become increasingly apparent that multifunctional receptors possess the capacity to interact with G proteins belonging to more than one subfamily. Many Gi-coupled receptors can signal through G proteins other than the Gi/Go proteins. The m-opioid receptor, a close relative of the ORL1 receptor, can interact with G proteins from both the Gi and Gq subfamily (Chan et al., 1995; Offermanns and Simon, 1995; Lee et al., 1998). Among the seven known G proteins that can interact with the m-opioid receptor, at least two are PTX insensitive (Gz and G16). Given that the ORL1 receptor is closely related to the opioid receptor family, it may likewise activate PTX-insensitive G proteins to regulate disparate effector pathways. To gain further insights on the signaling capacity of the ORL1 receptor, we screened for potential coupling of the receptor to a panel of PTX-insensitive G