Neurokinin 1 receptor desensitization and resensitization: is it all happening at the membrane? Focus on "Protein phosphatase 2A mediates resensitization of the neurokinin 1 receptor".

G PROTEIN-COUPLED RECEPTORS (GPCRs, also designated as heptahelical receptors or 7TMRs) are integral membrane proteins characterized by seven transmembrane -helices. GPCRs, with 800 genes, represent the largest transmembrane receptor superfamily in the human genome (11). Many intercellular messengers, including hormones, neurotransmitters, chemokines, growth and developmental factors, bioactive lipids, proteases, amino acids, nucleotides, biogenic amines and ions, initiate their characteristic effects on their target cells by binding to 350 nonchemosensory GPCRs and thereby function as neurotransmitters, local regulators, or systemic hormones. Accordingly, GPCRs mediate the majority ( 80%) of signal transduction across cell membranes in the organism (7). GPCRs and their cognate agonists are not only implicated in the regulation of postmitotic functions of differentiated cells but also act as growth factors and are implicated in autocrine/ paracrine stimulation of proliferation of cancer cells (16). At present, GPCRs are the most prominent targets for therapeutic drugs as they play an important role in a variety of diseases from cancer and diabetes, to neurodegenerative, inflammatory, gastrointestinal, cardiovascular, and behavioral disorders. GPCRs are therefore of utmost interest in homeostatic regulation, pathophysiologic states, and drug development. Repeated or sustained application of agonists rapidly decreases signaling by the corresponding GPCR in response to subsequent challenge with the same ligand, a process known as homologous desensitization. A widely accepted model envisages that inactivation of GPCR signaling in the setting of homologous desensitization is initiated by G protein-coupled receptor kinase (GRK) which phosphorylates the COOH-terminal tail and/or third intracellular loop of activated GPCRs (12), thereby promoting the recruitment of -arrestins and formation of a molecular complex between the activated (ligand-bound) and phosphorylated GPCR and arrestin (Fig. 1). The binding of -arrestin to the activated GPCR uncouples the receptor from the -subunit of heterotrimeric G proteins, thereby causing desensitization. -Arrestin also recruits cellular proteins (e.g., clathrin) that mediate GPCR internalization, traffic, and intracellular sorting (6, 8), leading to recycling of a resensitized (dephosphorylated) GPCR or, alternatively, to GPCR degradation (Fig. 1). In recent years, it has become apparent that GPCRs also mediate intracellular signaling through G protein-independent pathways, e.g., via -arrestin (13), and that the mechanisms leading to GPCR desensitization, trafficking, and resensitization are far more complex than initially thought. There are multiple GRK isoforms (GRK 1–7) that bind the active conformation of GPCRs in different cell types (5). In turn, different GRKs phosphorylate different residues in the receptor, leading to different outcomes. GRKs act not only through phosphorylation-dependent (14) but also via phosphorylationindependent (4) mechanisms, including sequestration of heterotrimeric G proteins (19). GPCRs can also be phosphorylated by other protein kinases, including protein kinase A (PKA) and members of the PKC family, in a cell context-dependent manner. There are at least two nonvisual arrestins ( -arrestin 1 and -arrestin 2, also termed arrestin 2 and arrestin 3, respectively) which can recruit different proteins and thereby not only initiate GPCR desensitization and internalization but also promote signaling by the internalized receptor (21). Thus, GPCR function is regulated at multiple levels by a large number of interacting proteins, including kinases, phosphatases, and scaffolds that assemble molecular complexes and fine tune the transduction of the GPCR signal initiated at the cell surface by the binding of an agonistic ligand. Tachykinins are a family of biologically active peptides distributed in the central and peripheral nervous system. Three distinct GPCR subtypes have been identified for tachykinins, designated as neurokinin 1 receptor (NK1R), NK2R, and NK3R. The most potent of the tachykinins, the undecapeptide substance P (SP), binds preferentially to NK1R. Many reports have demonstrated that repeated application of SP induces acute homologous desensitization of NK1R signaling followed by resensitization (1, 9, 15, 20). In line with other GPCR/ligand systems, this process has been envisaged to proceed through three steps: 1) internalization of the ligand-bound, -arrestincomplexed, and phosphorylated NK1R; 2) release of ligand, dissociation of the NK1R/ -arrestin complex, and NK1R dephosphorylation in endosomes; and 3) recycling of the receptor to the cell surface. The study by Murphy, Roosterman, and colleagues in this issue of American Journal of PhysiologyCell Physiology (9) addresses the contribution of internalization and recycling to the resensitization of NK1R function. A comparison of the time courses of recovery of SP binding sites at the cell surface and SP-induced Ca mobilization led the authors to the surprising conclusion that reactivation of NK1R located at the plasma membrane is the primary mechanism of resensitization of SP-induced Ca signaling. To study the mechanisms involved, Murphy, Roosterman, and colleagues (9) utilize several approaches to demonstrate that SP induces interaction of serine/threonine-protein phosphatase 2A (PP2A) and -arrestin1 at the plasma membrane and that -arrestin1, PP2A, and NK1R form a molecular complex, as shown by immunoprecipitation of surface-localAddress for reprint requests and other correspondence: E. Rozengurt, 900 Veteran Ave., Warren Hall Rm. 11-124, Dept. of Medicine, UCLA School of Medicine, Los Angeles, CA 90095-1786 (e-mail: [email protected]). Am J Physiol Cell Physiol 301: C772–C774, 2011; doi:10.1152/ajpcell.00274.2011. Editorial Focus