Search PubMed⌕ Search

SEARCH · Search PubMed

Results for “GPCR”

Search indexed PubMed citations on genomics, clinical trials, systematic reviews and public health. Explore titles, authors and supplied subject terms, then open the PubMed record.

Quote a phrase for an exact phrase match. Source license links do not imply unrestricted reuse.

At least 721 records · Page 40Linked to original sources
PubMed2002

Phospholipase C is required for glucose-induced calcium influx in budding yeast.

Intracellular calcium is a second messenger involved in several processes in yeast, such as mating, nutrient sensing, stress response and cell cycle events. It was reported that glucose addition stimulates a rapid increase in free calcium level in yeast. To investigate the calcium level variations induced by different stimuli we used a reporter system based on the photoprotein aequorin. Glucose addition (50 mM) to nutrient-starved cells induced an increase in free intracellular calcium concentration, mainly due to an influx from external medium. The increase of calcium reached its maximum 100-120 s after the stimulus. A concentration of about 20 mM glucose was required for a 50% increase in intracellular calcium. This response was completely abolished in strain plc1 Delta and in the isogenic wild-type strain treated with 3-nitrocoumarin, a phosphatidylinositol-specific phospholipase C inhibitor, suggesting that Plc1p is essential for glucose-induced calcium increase. This suggests that Plc1p should have a significant role in transducing glucose signal. The calcium influx induced by addition of high glucose on cells previously stimulated with low glucose levels was inhibited in strains with a deletion in the GPR1 or GPA2 genes, which suggests that glucose would be detected through the Gpr1p/Gpa2p receptor/G protein-coupled (GPCR) complex. Moreover, the signal was completely abolished in a strain unable to phosphorylate glucose, which is consistent with the reported requirement of glucose phosphorylation for GPCR complex activation.

Aequorin↗
PubMed2002

Endothelin suppresses cell migration via the JNK signaling pathway in a manner dependent upon Src kinase, Rac1, and Cdc42.

Cell migration is a complex phenomenon that is stimulated by chemoattractive factors such as chemokines, a family of ligands for G protein-coupled receptors (GPCRs). In contrast, factors that suppress cell migration, and the mechanism of their action, remain largely unknown. In this study, we show that endothelin, a GPCR ligand, inhibits cell motility in a manner dependent upon signaling through the c-Jun N-terminal kinase (JNK) pathway. We further demonstrate that this effect is dependent upon Src kinase and small GTPases Rac1 and Cdc42. These findings provide new insight into GPCR-mediated regulation of cell migration.

Cell Movement↗
PubMed2003

Platelet-derived growth factor (PDGF)-induced chemotaxis does not require the G protein-coupled receptor S1P1 in murine embryonic fibroblasts and vascular smooth muscle cells.

Sphingosine 1-phosphate (S1P), a bioactive lipid mediator, signals via G protein-coupled receptors (GPCR). The prototypical S1P receptor, S1P1 (also known as EDG-1), a Gi-linked receptor, is critical for vascular maturation during development. Recent work suggested that platelet-derived growth factor (PDGF)-induced cell migration required the S1P1 receptor, representing a novel mechanism for cross-talk between receptor tyrosine kinases and GPCRs. Since both S1P and PDGF are implicated in vascular smooth muscle cell (VSMC) pathobiology and development, we investigated this issue in rat VSMC and in embryonic fibroblasts derived from S1P1 null mice. Our data suggest that the S1P1 receptor is critical for S1P-induced, Gi-dependent migration but not for PDGF-BB-induced, receptor tyrosine kinase-dependent chemotaxis in VSMC. In addition, lack of S1P1 receptor in mouse embryonic fibroblasts did not significantly affect PDGF-induced cell migration. These data question the generality of the concept that S1P1 GPCR is a critical downstream component of PDGF-induced chemotaxis.

Animals↗
PubMed2003

The 'magic tail' of G protein-coupled receptors: an anchorage for functional protein networks.

All cell types express a great variety of G protein-coupled receptors (GPCRs) that are coupled to only a limited set of G proteins. This disposition favors cross-talk between transduction pathways. However, GPCRs are organized into functional units. They promote specificity and thus avoid unsuitable cross-talk. New methodologies (mostly yeast two-hybrid screens and proteomics) have been used to discover more than 50 GPCR-associated proteins that are involved in building these units. In addition, these protein networks participate in the trafficking, targeting, signaling, fine-tuning and allosteric regulation of GPCRs. To date, proteins that interact with the GPCR C-terminus are the most abundant and are the focus of this review.

Allosteric Regulation↗
PubMed2003

Photolysis of intracellular caged sphingosine-1-phosphate causes Ca2+ mobilization independently of G-protein-coupled receptors.

Sphingosine-1-phosphate (S1P), the product of sphingosine kinase, activates several widely expressed G-protein-coupled receptors (GPCR). S1P might also play a role as second messenger, but this hypothesis has been challenged by recent findings. Here we demonstrate that intracellular S1P can mobilize Ca(2+) in intact cells independently of S1P-GPCR. Within seconds, S1P generated by the photolysis of caged S1P raised the intracellular free Ca(2+) concentration in HEK-293, SKNMC and HepG2 cells, in which the response to extracellularly applied S1P was either blocked or absent. Ca(2+) transients induced by photolysis of caged S1P were caused by Ca(2+) mobilization from thapsigargin-sensitive stores. These results provide direct evidence for a true intracellular action of S1P.

Calcium↗
PubMed2004

Heavier-than-air flying machines are impossible.

Many G protein-coupled receptor (GPCR) models have been built over the years. The release of the structure of bovine rhodopsin in August 2000 enabled us to analyze models built before that period to learn more about the models we build today. We conclude that the GPCR modelling field is riddled with 'common knowledge' similar to Lord Kelvin's remark in 1895 that "heavier-than-air flying machines are impossible", and we summarize what we think are the (im)possibilities of modelling GPCRs using the coordinates of bovine rhodopsin as a template. Associated WWW pages: www.gpcr.org/articles/2003_mod

Amino Acid Sequence↗
PubMed1996

Characterization of a human gene related to genes encoding somatostatin receptors.

We report the identification of a gene, named SLC-1(1), encoding a novel G protein-coupled receptor (GPCR). A customized search procedure of a database of expressed sequence tags (dbEST) retrieved a human cDNA sequence that partially encoded a GPCR. A genomic DNA fragment identical to the cDNA was obtained and used to screen a library to isolate the full-length coding region of the gene. This gene was intronless in its open reading frame, and encoded a receptor of 402 amino acids, and shared -40% amino acid identity in the transmembrane (TM) regions to the five known human somatostatin receptors. Northern blot analysis revealed that SLC-1 is expressed in human brain regions, including the forebrain and hypothalamus. Expression in the rat was highest in brain, followed by heart, kidney, and ovary. Expression of SLC-1 in COS-7 cells failed to show specific binding to radiolabelled Tyr1-somatostatin-14, naloxone, bremazocine, 1,3-di(2-tolyl)-guanidine (DTG), or haloperidol. A repeat polymorphism of the form (CA)n was discovered in the 5'-untranslated region (UTR) of the gene and SLC-1 was mapped to chromosome 22, q13.3.

Amino Acid Sequence↗
PubMed1998

G protein-coupled receptor kinase 2 (GRK2): mechanisms of regulation and physiological functions.

G protein-coupled receptor kinase 2 (GRK2) plays a key role in determining the rate and extent of G protein-coupled receptor (GPCR) desensitization and resensitization. Recent data indicate that GRK2 activity, subcellular distribution and expression are tightly regulated. The important physiological function of GRK2 as a modulator of the efficacy of GPCR signal transduction systems is exemplified by its relevance in cardiovascular physiopathology as well as by its emerging role in the regulation of chemokine receptors.

Animals↗
PubMed1999

Discovery of a receptor related to the galanin receptors.

We report the isolation of a cDNA clone named GPR54, which encodes a novel G protein-coupled receptor (GPCR). A PCR search of rat brain cDNA retrieved a clone partially encoding a GPCR. In a library screening this clone was used to isolate a cDNA with an open reading frame (ORF) encoding a receptor of 396 amino acids long which shared significant identities in the transmembrane regions with rat galanin receptors GalR1 (45%), GalR3 (45%) and GalR2 (44%). Northern blot and in situ hybridization analyses revealed that GPR54 is expressed in brain regions (pons, midbrain, thalamus, hypothalamus, hippocampus, amygdala, cortex, frontal cortex, and striatum) as well as peripheral regions (liver and intestine). In COS cell expression of GPR54 no specific binding was observed for 125I-galanin. A recent BLAST search with the rat GPR54 ORF nucleotide sequence recovered the human orthologue of GPR54 in a 3.5 Mb contig localized to chromosome 19p13.3.

Amino Acid Sequence↗
PubMed1998

Receptor binding thermodynamics as a tool for linking drug efficacy and affinity.

Determination of drug-receptor binding constants (association, KA, or dissociation, KD = l/KA) by radiochemical specific binding assays has proved to be an invaluable tool for screening of potential active drugs. Simple determination of KA (or KD) values makes it possible, however, to calculate the standard free energy delta G degree = -RTln KA = RTln KD (T = 298.15 K) of the binding equilibrium but not that of its two components as defined by the Gibbs equation delta G degree = delta H degree - T delta S degree, where delta H degree and delta S degree are the equilibrium standard enthalpy and entropy, respectively. This incomplete knowledge is highly inconvenient from a pure thermodynamic point of view as delta H degree and delta S degree carry much information on the details of the drug-receptor interaction and the interplay of both reaction partners with the solvent. In recent times it has been shown that the relative delta H degree and delta S degree magnitudes can often give a simple 'in vitro' way for discriminating 'the effect', that is the manner in which the drug interferes with the signal transduction pathways. This particular effect, called 'thermodynamic discrimination', results from the fact that binding of antagonists may be enthalpy-driven and that of agonists entropy-driven, or vice versa. The first case of thermodynamic discrimination was reported for the beta-adrenergic G-protein coupled receptor (GPCR) and only recently has it been confirmed for adenosine A1 and A2a receptors. Only very recently has the binding thermodynamics of ligand-gated ion channel receptors (LGICR) been investigated and data for four receptors have been reported showing that all of them are thermodynamically discriminated. While it seems difficult at present to find a reasonable explanation for the thermodynamic discrimination phenomenon in GPCR, some hypotheses can be suggested for LGICR. Since global delta H degree and delta S degree values of the binding process are expected to be heavily affected by rearrangements occurring in the solvent, thermodynamic discrimination in LGICR is at least logically understandable admitting that the observed delta H degree (and then delta S degree) values are determined by both specific binding and abrupt variation of water-accessible receptor surfaces consequent to the setting up of the channel opening.

GTP-Binding Proteins↗
PubMed2003

The human formyl peptide receptor as model system for constitutively active G-protein-coupled receptors.

According to the two-state model of G-protein-coupled receptor (GPCR) activation, GPCRs isomerize from an inactive (R) state to an active (R*) state. In the R* state, GPCRs activate G-proteins. Agonist-independent R/R* isomerization is referred to as constitutive activity and results in an increase in basal G-protein activity, i.e. GDP/GTP exchange. Agonists stabilize the R* state and further increase, whereas inverse agonists stabilize the R state and decrease, basal G-protein activity. Constitutive activity is observed in numerous wild-type GPCRs and disease-causing GPCR mutants with increased constitutive activity. The human formyl peptide receptor (FPR) exists in several isoforms (FPR-26, FPR-98 and FPR-G6) and activates chemotaxis and cytotoxic cell functions of phagocytes through G(i)-proteins. Studies in HL-60 leukemia cell membranes demonstrated inhibitory effects of Na(+) and pertussis toxin on basal G(i)-protein activity, suggesting that the FPR is constitutively active. However, since HL-60 cells express several constitutively active chemoattractant receptors, analysis of constitutive FPR activity was difficult. Sf9 insect cells do not express chemoattractant receptors and G(i)-proteins and provide a sensitive reconstitution system for FPR/G(i)-protein coupling. Such expression studies showed that FPR-26 is much more constitutively active than FPR-98 and FPR-G6 as assessed by the relative inhibitory effects of Na(+) and of the inverse agonist cyclosporin H on basal G(i)-protein activity. Site-directed mutagenesis studies suggest that the E346A exchange in the C-terminus critically determines dimerization and constitutive activity of FPR. Moreover, N-glycosylation of the N-terminus seems to be important for constitutive FPR activity. Finally, we discuss some future directions of research.

Amino Acid Sequence↗
PubMed2003

CaVbeta subunit-mediated up-regulation of CaV2.2 currents triggered by D2 dopamine receptor activation.

Voltage-dependent Ca(2+) channels (VDCCs) are subject to modulation by a number of pathways, including membrane-delimited inhibition by heterotrimeric G-proteins and modulation through phosphorylation by diverse kinases. Here we report that in the Xenopus oocyte expression system Ca(V)2.2 channels undergo a sustained, linear and irreversible run-up lasting up to 30 min, which is potentiated during G-protein-mediated inhibition by activation of co-expressed G-protein coupled receptors (GPCRs). This up-regulation is not a result of receptor desensitization, but is associated with a hyperpolarization of the voltage for activation and depends on the presence of accessory subunits such that beta subunits promote, and alpha2delta subunits oppose the current increase. We have investigated the involvement of G-proteins and found that over-expression of Galpha(o) subunits or Galpha-transducin reduced the amount of agonist-mediated up-regulation. However, we have found no evidence for the involvement of any second messenger pathways in the increase of current run-up in the presence of a GPCR agonist. Taken together, our data suggest that the effect reported herein involves an enhancement of the GTPase activity of endogenous Galpha subunits, which is triggered by GPCR activation and mediated by accessory Ca(V)beta subunits. It may involve an increased association of Ca(V)beta subunits with alpha1 subunits in the plasma membrane or trafficking of channels to the plasma membrane.

Animals↗
PubMed2002

Age-dependent agonist-specific dysregulation of membrane-mediated signal transduction: emergence of the gate theory of aging.

Although a general mechanism for the limited responsiveness of senescent cells has yet to be established, reduced responsiveness may in part be ascribed to deficits in the apparatus required for cell surface receptor-mediated signal transduction. Age-related changes of receptor-mediated signal transduction occur at many levels, and are known to include quantitative and qualitative changes in growth factor receptors, G-protein coupled receptors, and many other downstream signaling molecules. Here, we emphasize the prime role of the cellular surface in the perception and transmission of external stimuli in response to the aging process. As major means of cellular signal transduction, the receptor tyrosine kinase (RTK) system and the G protein-coupled receptor (GPCR) system of senescent cells were investigated. We observed that the RTK system was severely damaged, while the GPCR system was only partially inactivated by aging. These results suggest that the agonist-dependent dysregulation of and imbalance of signal transduction pathways might be responsible for the functional deterioration of senescent cells, and indicate a possibility of the functional recovery of senescent cells through agonist-specific signal system activation. Moreover, those data evoke the emerging concept that the senescent phenotype may be modulated by the membrance-associated signal system, implying the gate theory of aging.

Aging↗
PubMed1998

Regulation of mitogen-activated protein kinase cascades in the heart.

Using primary cultures of neonatal rat ventricular myocytes and isolated adult rat hearts as models, we have characterized extensively the regulation of MAPKs in the heart. The ERKs are activated primarily by GPCR agonists acting through PKC. These agonists can also activate the JNKs although the mechanism is unclear. Cellular stresses stimulate strong activation of the JNKs, but also cause some stimulation of ERKs. Activation of p38-MAPK has so far only been demonstrated in intact adult hearts subjected to stresses and probably leads to activation of MAPKAPK2. Both cellular stresses and GPCR agonists induce phosphorylation of c-Jun, but only the latter causes upregulation of c-Jun protein.

Animals↗
PubMed2003

G protein-coupled receptors in invertebrates: a state of the art.

G protein-coupled receptors (GPCRs) constitute one of the largest and most ancient superfamilies of membrane-spanning proteins. We focus on neuropeptide GPCRs, in particular on those of invertebrates. In general, such receptors mediate the responses of signaling molecules that constitute the highest hierarchical position in the regulation of physiological processes. Until recently, only a few of these receptors were identified in invertebrates. However, the availability of a plethora of genomic information has boosted the discovery of novel members in several invertebrate species, such as Drosophila, in which 18 neuropeptide GPCRs have been characterized. The finalization of genomic projects in other invertebrates will lead to a similar expansion of GPCR understanding. Many new insights regarding neuropeptide regulation have followed from the discovery of their cognate receptors. Furthermore, information on GPCR signaling is still fragmentary and the elucidation of these pathways in model insects such as Drosophila will lead to further insights in other species, including mammals. In this review we present the current status of what is known about invertebrate GPCRs, discuss some novel perceptions that follow from the identified members, and, finally, present some future prospects.

Amino Acid Sequence↗
PubMed2006

Cellular and molecular biology of orphan G protein-coupled receptors.

The superfamily of G protein-coupled receptors (GPCRs) is the largest and most diverse group of membrane-spanning proteins. It plays a variety of roles in pathophysiological processes by transmitting extracellular signals to cells via heterotrimeric G proteins. Completion of the human genome project revealed the presence of approximately 168 genes encoding established nonsensory GPCRs, as well as 207 genes predicted to encode novel GPCRs for which the natural ligands remained to be identified, the so-called orphan GPCRs. Eighty-six of these orphans have now been paired to novel or previously known molecules, and 121 remain to be deorphaned. A better understanding of the GPCR structures and classification; knowledge of the receptor activation mechanism, either dependent on or independent of an agonist; increased understanding of the control of GPCR-mediated signal transduction; and development of appropriate ligand screening systems may improve the probability of discovering novel ligands for the remaining orphan GPCRs.

Animals↗
PubMed2004

Structure-based design, synthesis, and activity of peptide inhibitors of RGS4 GAP activity.

One of the principal roles of the multifunctional regulator of G-protein signaling (RGS) proteins is to terminate G-protein-coupled receptor (GPCR) signaling by binding to the G-protein Galpha subunit, thus acting as GTPase-activating proteins (GAPs). In principle, then, selective inhibitors of this GAP function would have potential as therapeutic agents, as they could be used to augment the effects of endogenous or exogenous GPCR agonists. Using the published RGS4-G(ialpha1) X-ray structure, we have designed and synthesized a series of cyclic peptides, modeled on the G(ialpha) switch I region, that inhibit RGS4 GAP activity, presumably by blocking the interaction between RGS4 and G(ialpha1). These compounds should prove useful for elucidating RGS-mediated activity and serve as a starting point for the development of a novel class of therapeutic agent.

Amino Acid Sequence↗
PubMed2004

Analysis of differential modulatory activities of GRK2 and GRK4 on Galphaq-coupled receptor signaling.

G-protein-coupled receptor kinases (GRK) contain a regulator of G-protein signaling (RGS)-like domain located at the N terminus (GRK-Nter) of their sequence. This domain is present in all the GRK subtypes, but the RGS-like domain of GRK2 was documented to be functionally active, as it is able to interact selectively with Galphaq (both in vitro and in cells) and to inhibit Galphaq-dependent signaling. In contrast GRK4, GRK5, and GRK6 are unable to interact with Galphaq. This article describes the methodology to investigate the modulatory activity of GRK2 and GRK4 on GPCR-stimulated Galphaq signaling. This analysis is essentially based on three types of experiments: (a) study of the effect of the GRK-Nter on GPCR-dependent signaling; (b) analysis of the binding of GRK-Nter to Galphaq in vitro; and (c) analysis of the interaction of GRK with Galphaq in cells.

Cell Line↗