Gene Ontology annotation through association of InterPro records with GO terms
Annotation inferences using phylogenetic trees
Gene Ontology annotation based on curation of immunofluorescence data
Automatic transfer of experimentally verified manual GO annotation data to orthologs using Ensembl Compara
Combined Automated Annotation using Multiple IEA Methods
Cloning and chromosomal mapping of four putative novel human G-protein-coupled receptor genes.
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GPR23, the gene later deorphanized as LPAR4, was initially isolated as a human putative GPCR and mapped to Xq13-q21.1.
"This resulted in the isolation of genes GPR21, GPR22 and GPR23."
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The initial human GPR23 study established chromosomal position but did not identify a ligand or signaling function.
"Fluorescence in situ hybridization (FISH) was used to map GPR20 to chromosome 8q, region 24.3-24.2, GPR21 to chromosome 9, region q33, GPR22 to chromosome 7, region q22-q31.1, and GPR23 to chromosome X, region q13-q21.1."
Cloning of a human heptahelical receptor closely related to the P2Y5 receptor.
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The human receptor historically called P2Y5-like/P2Y9 was cloned as a 370-aa heptahelical GPCR.
"Starting from this partial sequence, we have isolated a complete clone and identified a 1113 base pair open reading frame encoding a new G-coupled receptor that we have called P2Y5-like."
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Forty tested nucleotides and nucleosides failed to activate the receptor in four signaling assays, arguing against interpreting the historical P2Y name as demonstrated purinergic activity.
"None of the 40 nucleotides and nucleosides tested was able to elicit a response in any of four functional assays: inositol phosphate formation, stimulation or inhibition of cAMP formation, and extracellular acidification measured with a microphysiometer."
Identification of p2y9/GPR23 as a novel G protein-coupled receptor for lysophosphatidic acid, structurally distant from the Edg family.
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Human p2y9/GPR23 was deorphanized as LPA4 by direct, saturable 1-oleoyl-LPA binding in membranes of receptor-expressing cells.
"Membrane fractions of RH7777 cells transiently expressing p2y9/GPR23 displayed a specific binding for 1-oleoyl-LPA with a Kd value of around 45 nm."
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LPAR4 discriminates among LPA molecular species, preferring 1-oleoyl-LPA in the reported rank order.
"Competition binding and reporter gene assays showed that p2y9/GPR23 preferred structural analogs of LPA with a rank order of 1-oleoyl- > 1-stearoyl- > 1-palmitoyl- > 1-myristoyl- > 1-alkyl- > 1-alkenyl-LPA."
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In heterologous CHO cells, LPA-bound LPAR4 elevated intracellular calcium and stimulated adenylyl cyclase.
"In Chinese hamster ovary cells expressing p2y9/GPR23, 1-oleoyl-LPA induced an increase in intracellular Ca2+ concentration and stimulated adenylyl cyclase activity."
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The original human tissue survey found LPAR4 transcript most abundant in ovary.
"Quantitative real-time PCR demonstrated that mRNA of p2y9/GPR23 was significantly abundant in ovary compared with other tissues."
LPA4/p2y9/GPR23 mediates rho-dependent morphological changes in a rat neuronal cell line.
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LPAR4 drove Gq/11-dependent calcium mobilization in rat B103 neuroblastoma transfectants but did not alter adenylyl cyclase in that system.
"In B103 cells stably expressing LPA(4), we observed G(q/11)-dependent calcium mobilization, but LPA did not affect adenylyl cyclase activity."
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LPA induced Rho-dependent neurite retraction, aggregation, and cadherin-dependent adhesion through transfected LPAR4.
"In LPA(4) transfectants, LPA induced dramatic morphological changes, i.e. neurite retraction, cell aggregation, and cadherin-dependent cell adhesion, which involved Rho-mediated signaling pathways."
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The study inferred LPAR4 coupling to Gq/11 and G12/13 but not Gi/o in this rat neuronal-cell context.
"Thus, our results demonstrated that LPA(4) as well as LPA(1) couple to G(q/11) and G(12/13), whereas LPA(4) differs from LPA(1) in that it does not couple to G(i/o)."
LPA(4)/GPR23 is a lysophosphatidic acid (LPA) receptor utilizing G(s)-, G(q)/G(i)-mediated calcium signaling and G(12/13)-mediated Rho activation.
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Epitope-tagged LPAR4 showed concentration-dependent LPA responses and specific membrane binding, independently confirming it as an LPA receptor.
"Using epitope-tagged LPA(4), pharmacological intervention, and G protein mini-genes, we provide independent confirmatory evidence that supports LPA(4) as a fourth LPA receptor, including LPA concentration-dependent responses and specific membrane binding."
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The study attributed LPAR4-dependent cAMP increases to Gs, calcium mobilization to Gq and a pertussis-toxin-sensitive pathway, and Rho-dependent cytoskeletal outputs to G12/13.
"Importantly, we further demonstrate new LPA-dependent activities of LPA(4) that include the following: receptor internalization; G(12/13)- and Rho-mediated neurite retraction and stress fiber formation; G(q) protein and pertussis toxin-sensitive calcium mobilization and activation of a nonselective cation conductance; and cAMP increases mediated by G(s)."
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LPA stimulation induced internalization of epitope-tagged LPAR4 in the study system.
"Importantly, we further demonstrate new LPA-dependent activities of LPA(4) that include the following: receptor internalization; G(12/13)- and Rho-mediated neurite retraction and stress fiber formation; G(q) protein and pertussis toxin-sensitive calcium mobilization and activation of a nonselective cation conductance; and cAMP increases mediated by G(s)."
Role of LPA4/p2y9/GPR23 in negative regulation of cell motility.
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Mouse Lpar4 loss enhanced LPA-driven fibroblast migration and shifted Akt/Rac/Rho signaling, whereas receptor reconstitution reduced motility.
"Consistent with negative modulation of the phosphatidylinositol 3 kinase pathway by LPA(4), LPA(4) deficiency potentiated Akt and Rac but decreased Rho activation induced by LPA. Reconstitution of LPA(4) converted LPA(4)-negative cells into a less motile phenotype."
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Ectopic LPAR4 inhibited migration and invasion in human cancer cells and antagonized LPA1-driven motility in receptor-null B103 cells.
"In support of the biological relevance of these observations, ectopic expression of LPA(4) strongly inhibited migration and invasion of human cancer cells. When coexpressed with LPA(1) in B103 neuroblastoma cells devoid of endogenous LPA receptors, LPA(4) attenuated LPA(1)-driven migration and invasion, indicating functional antagonism between the two subtypes of LPA receptors."
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The first targeted mouse Lpar4 knockout had no gross apparent phenotype, bounding strong developmental claims from this model.
"Although LPA(4)-deficient mice displayed no apparent abnormalities, LPA(4)-deficient mouse embryonic fibroblasts (MEFs) were hypersensitive to LPA-induced cell migration."
Autotaxin promotes cancer invasion via the lysophosphatidic acid receptor 4: participation of the cyclic AMP/EPAC/Rac1 signaling pathway in invadopodia formation.
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In fibrosarcoma cells, autotaxin-derived LPA signaled through LPAR4 to promote invadopodia through a cAMP-EPAC-Rac1 pathway.
"We further provide evidence that LPA(4) signaling in fibrosarcoma cells regulates invadopodia formation downstream of ATX, a process mediated through the activation of EPAC by cyclic AMP and subsequent Rac1 activation."
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LPAR4 knockdown supported a requirement for the receptor in invasion and in vivo metastasis formation in this cancer model.
"Results using LPA(4) shRNA support the requirement of the LPA(4) receptor for cell invasion and in vivo metastasis formation."
Strategy for the identification of GPR23/LPA4 receptor agonists and inverse agonists.
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Human GPR23 expressed in CHO cells reproduced LPA-dependent cAMP and calcium responses.
"In Chinese hamster ovary cells expressing the human GPR23, LPA induced an increase in cellular cyclic adenosine monophosphate (cAMP) and calcium levels."
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Orthogonal reporter and radioligand-binding screens yielded LPAR4 agonists, inverse agonists, and a negative modulator.
"Here we report the identification of novel GPR23 agonists, inverse agonists, and a negative modulator from 2 high-throughput screens, a beta-lactamase reporter screen, and a [3H]LPA-binding screen."
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Candidate mechanisms and selectivity were tested with reporter, cAMP, and other LPA-receptor assays.
"Several screening hits were selected for mechanism of action studies using the beta-lactamase reporter assay and a cAMP assay. An evaluation of their selectivity at the other LPA receptors was also conducted."
Development of a GPR23 cell-based β-lactamase reporter assay.
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An inducible reporter assay detected constitutive GPR23/LPAR4 activity.
"This report describes how a tetracycline-inducible system was utilized in conjunction with a sensitive β-lactamase reporter gene to develop an assay in which constitutive activity of the receptor could be monitored."
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Screening with the reporter identified the first reported small-molecule inverse agonists for GPR23.
"This assay was then utilized to screen a 1.1 million compound library to identify the first small molecule inverse agonists for the receptor."
Lysophosphatidic Acid Receptor 4 Is Transiently Expressed during Cardiac Differentiation and Critical for Repair of the Damaged Heart.
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LPAR4 expression transiently peaks in cardiac progenitors during both mouse and human PSC differentiation.
"During in vitro differentiation of mouse and human PSCs toward cardiac lineage, LPAR4 expression peaked after 3-7 days of differentiation in cardiac progenitors and then declined."
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Sequential LPAR4 stimulation and downstream p38 inhibition enhanced cardiac differentiation in mouse and human PSC cultures.
"Sequential stimulation and inhibition of LPAR4 using these agents enhanced the in vitro efficiency of cardiac differentiation from mouse and human PSCs."
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The in vivo cardiac-repair result was demonstrated in mice and should not be transferred directly to human physiology.
"Importantly, in vivo, this sequential stimulation and inhibition of LPAR4 reduced the infarct size and rescued heart dysfunction in mice."
Multiplexed mapping of the interactome of GPCRs with receptor activity-modifying proteins.
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The RAMP interactome screen tested every GPCR-RAMP pair by ectopic coexpression and membrane-solubilization assays.
"All potential GPCR-RAMP interacting pairs were expressed ectopically, solubilized and analyzed using the multiplexed suspension bead array (SBA) strategy."
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The library screen used one biological replicate per GPCR-only or GPCR-RAMP sample, with separate detection schemes supplying the two replicates.
"For the DuET library–based GPCR-RAMP interactome screen, we used one biological replicate of each of the four unique GPCR-containing samples (each GPCR alone and each GPCR with each of the three RAMPs) in two replicates. Each replicate represented one detection scheme."
Liganded Gq/11-activating GPCRs act as GEFs for Gq/11
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Reactome places Gq/11-linked GPCRs upstream of PLC, IP3, and calcium mobilization.
"PLC hydrolyzes phosphatidylinositol (PIP2) to diacyl glycerol (DAG) and inositol triphosphate (IP3)."
P2Y9 receptor can bind to LPA
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Reactome identifies human GPR23/P2Y9 as LPAR4 and models LPA binding with signaling through multiple G proteins.
"P2Y9 has been reported to bind lysophosphatidic acid (LPA) as a ligand and elicit numerous effects via multiple G proteins (Lee CW et al, 2007)."
Liganded Gq-activating GPCRs bind inactive heterotrimeric Gq
The Ligand:GPCR:Gq complex dissociates