{"gene":"GPR176","run_date":"2026-06-10T01:55:21","timeline":{"discoveries":[{"year":2016,"finding":"GPR176 is an SCN-enriched orphan GPCR that represses cAMP signalling in an agonist-independent (constitutively active) manner, and couples to the unique G-protein subclass Gz (not canonical Gi) to reduce cAMP production and set the pace of circadian behaviour.","method":"Genetic knockout mice, heterologous expression assays, cAMP measurement, G-protein coupling analysis","journal":"Nature Communications","confidence":"High","confidence_rationale":"Tier 2 / Strong — reciprocal genetic and biochemical approaches in vivo and in vitro, replicated in subsequent reviews and follow-up papers","pmids":["26882873"],"is_preprint":false},{"year":2016,"finding":"GPR176 acts independently of and in parallel to the Vipr2 GPCR pathway in the SCN, as established by genetic epistasis in knockout animals.","method":"Genetic epistasis analysis using Gpr176 and Vipr2 mutant mice","journal":"Nature Communications","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — genetic epistasis in vivo, single lab, circadian period phenotype readout","pmids":["26882873"],"is_preprint":false},{"year":2020,"finding":"GPR176 undergoes N-linked glycosylation at four conserved asparagine residues in its N-terminal region; mutation of these residues reduces protein expression and attenuates cAMP-repressive activity in cells, establishing N-glycosylation as a prerequisite for efficient expression of functional GPR176.","method":"Peptide-N-glycosidase F treatment of mouse hypothalamus extracts, site-directed mutagenesis of N-glycosylation sites, heterologous expression, cAMP assay","journal":"Scientific Reports","confidence":"High","confidence_rationale":"Tier 1 / Moderate — in vitro enzymatic deglycosylation, site-directed mutagenesis with functional cAMP readout, endogenous tissue validation, single lab with multiple orthogonal methods","pmids":["32157140"],"is_preprint":false},{"year":2020,"finding":"N-glycosylation of GPR176 is required for proper cell-surface expression; deficient N-glycosylation does not directly compromise the intrinsic agonist-independent cAMP-repressive activity but reduces it indirectly by lowering total protein levels.","method":"Site-directed mutagenesis of N-glycosylation sites, heterologous expression, cAMP assay","journal":"Scientific Reports","confidence":"Medium","confidence_rationale":"Tier 1 / Moderate — mutagenesis with functional readout, single lab","pmids":["32157140"],"is_preprint":false},{"year":2023,"finding":"GPR176 physically interacts with G protein GNAS intracellularly via its transmembrane helix 3–intracellular loop 2 domain; this GPR176/GNAS complex activates the cAMP/PKA signalling pathway and inhibits mitophagy via the cAMP/PKA/BNIP3L axis to promote colorectal cancer progression.","method":"Co-immunoprecipitation, homology modelling, in vitro and in vivo cancer models with Gpr176-deficient mice, cAMP/PKA assay, mitophagy assays","journal":"Advanced Science","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — Co-IP and genetic mouse model with mechanistic pathway readout, single lab, domain mapping by homology model only","pmids":["36905238"],"is_preprint":false},{"year":2022,"finding":"Gpr176 knockout in mice leads to upregulation of Nmu and Nms mRNA in the SCN; triple knockout of Nmu/Nms/Gpr176 results in enhanced light-induced phase shifts and reduced Per1 and cFos induction by light, indicating a functional interaction among Nmu, Nms, and Gpr176 in modulating light-induced circadian phase shifts.","method":"Microarray analysis, triple knockout mouse generation and behavioural phenotyping, SCN gene expression analysis","journal":"Biological & Pharmaceutical Bulletin","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — genetic epistasis with triple KO mice and molecular readouts, single lab","pmids":["35908898"],"is_preprint":false},{"year":2024,"finding":"GPR176 is induced in activated hepatic stellate cells (HSCs) and plays a profibrotic role: siRNA-mediated knockdown reduces fibrogenic characteristics in primary mouse HSCs and precision-cut liver slices; Gpr176 knockout mice develop less severe fibrosis in CCl4 and bile duct ligation models.","method":"siRNA knockdown in primary mouse HSCs and PCLS, GPR176 knockout mouse fibrosis models (CCl4 and BDL), immunohistochemistry of human CLD tissue","journal":"JHEP Reports","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — loss-of-function both in vitro and in vivo with defined fibrosis phenotype, single lab, two orthogonal loss-of-function models","pmids":["38694958"],"is_preprint":false},{"year":2024,"finding":"GPR176 promotes fibroblast-to-myofibroblast transition in fibrosis: Gpr176 expression is increased in fibrotic lungs, kidneys, liver, and heart; siRNA knockdown of Gpr176 in rat renal fibroblasts reduces TGFβ1-induced expression of αSMA, fibronectin, and collagen, and attenuates Smad2 phosphorylation, without being itself regulated by TGFβ1.","method":"Gene expression analysis in fibrosis mouse models, siRNA knockdown in NRK-49F cells, Western blot for αSMA/fibronectin/collagen and phospho-Smad2","journal":"Biochimica et Biophysica Acta – Molecular Cell Research","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — siRNA loss-of-function with defined molecular pathway readout (Smad2 phosphorylation), single lab","pmids":["39047914"],"is_preprint":false},{"year":2025,"finding":"Exosomal miR-382-5p downregulates GPR176 expression and disrupts its interaction with GNAS in the liver, thereby reducing CXCR1/CXCR2 levels and suppressing angiogenesis and vascular permeability in colorectal cancer liver metastasis models.","method":"RNA pull-down, RNA immunoprecipitation (RIP), Co-IP, in vivo and in vitro angiogenesis/vascular permeability assays, exosome isolation and characterization","journal":"Cellular Signalling","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — Co-IP and RIP with functional in vivo validation, single lab","pmids":["40578589"],"is_preprint":false},{"year":2024,"finding":"A cell-based assay (GzESTY) detected the presence of endogenous ligands for GPR176 in brain extracts, providing functional evidence that endogenous activating ligands for GPR176 exist in brain tissue.","method":"Cell-based Gz-coupled signal transduction assay (GzESTY) with brain extract fractions","journal":"bioRxiv (preprint)","confidence":"Low","confidence_rationale":"Tier 3 / Weak — single cell-based assay in a preprint, no ligand identity established, single lab","pmids":["bio_10.1101_2024.07.26.605282"],"is_preprint":true},{"year":2025,"finding":"Gpr176 is expressed predominantly in parvalbumin-positive (PV+) interneurons of the prefrontal cortex; knockdown of Gpr176 increases firing output of PV+ interneurons by altering membrane potential changes during the repolarizing phase of action potentials, without affecting synaptic activity.","method":"In situ expression analysis, shRNA-mediated knockdown, whole-cell electrophysiology in PV+ interneurons, behavioural assays","journal":"Molecular Brain","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — direct electrophysiological readout with targeted knockdown in specific cell type, single lab","pmids":["41188983"],"is_preprint":false},{"year":2026,"finding":"E2F4 directly binds the GPR176 promoter and transcriptionally activates GPR176 expression; elevated GPR176 suppresses mitophagy and confers resistance to ferroptosis in esophageal cancer cells, and GPR176 overexpression abrogates the enhanced mitophagy and ferroptosis induced by E2F4 depletion.","method":"ChIP/promoter binding assay, overexpression and knockdown of E2F4 and GPR176, mitophagy markers (MMP, ROS, autophagy proteins), ferroptosis markers (MDA, Fe2+, lipid ROS), rescue experiments","journal":"Human Mutation","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — direct promoter binding assay plus rescue experiments establishing epistasis, single lab","pmids":["42253509"],"is_preprint":false}],"current_model":"GPR176 is a constitutively active (agonist-independent), N-glycosylated orphan class-A GPCR that couples to the Gz G-protein subclass to suppress cAMP/PKA signalling; in the SCN it sets circadian period and modulates light-induced phase shifts via this Gz–cAMP axis, while in peripheral tissues it promotes fibrosis through Smad2-dependent fibroblast-to-myofibroblast transition and drives cancer progression by recruiting GNAS via its TM3–ICL2 domain to activate cAMP/PKA/BNIP3L signalling and repress mitophagy."},"narrative":{"mechanistic_narrative":"GPR176 is a constitutively active, SCN-enriched orphan class-A GPCR that suppresses cAMP signalling in an agonist-independent manner by coupling to the atypical Gz G-protein subclass, thereby setting the pace of circadian behaviour [PMID:26882873]. This Gz–cAMP activity operates in parallel to the Vipr2 pathway in the SCN [PMID:26882873] and is also embedded in a regulatory circuit with the neuropeptides Nmu/Nms that tunes light-induced phase shifts [PMID:35908898]. Efficient cell-surface expression of functional GPR176 depends on N-linked glycosylation at four conserved N-terminal asparagine residues, loss of which lowers protein levels and indirectly attenuates its cAMP-repressive output [PMID:32157140]. Beyond the clock, GPR176 functions as a pro-fibrotic and pro-tumorigenic effector: it drives fibroblast-to-myofibroblast transition through Smad2 phosphorylation and induction of αSMA, fibronectin, and collagen, and promotes fibrosis across multiple organs [PMID:38694958, PMID:39047914]. In cancer it physically engages the G-protein GNAS via its transmembrane helix 3–intracellular loop 2 region to activate cAMP/PKA signalling and repress mitophagy through the BNIP3L axis, promoting colorectal cancer progression [PMID:36905238], a process antagonized by exosomal miR-382-5p that downregulates GPR176 and disrupts the GPR176–GNAS interaction [PMID:40578589]. GPR176 is transcriptionally activated by E2F4 and confers ferroptosis resistance by suppressing mitophagy in esophageal cancer [PMID:42253509]. In the prefrontal cortex it is expressed in parvalbumin-positive interneurons where it constrains their firing output [PMID:41188983].","teleology":[{"year":2016,"claim":"Established GPR176's core molecular identity: an orphan GPCR that represses cAMP constitutively through the unusual Gz subclass to set circadian period, answering how this SCN-enriched receptor controls the clock without a known ligand.","evidence":"Knockout mice, heterologous expression, cAMP measurement and G-protein coupling analysis","pmids":["26882873"],"confidence":"High","gaps":["No endogenous ligand identified","Mechanism of constitutive activity at structural level unresolved"]},{"year":2016,"claim":"Showed GPR176 acts in a pathway genetically separate from Vipr2, clarifying that it is a non-redundant input to SCN timekeeping.","evidence":"Genetic epistasis with Gpr176 and Vipr2 mutant mice","pmids":["26882873"],"confidence":"Medium","gaps":["Molecular basis of pathway independence not defined","Single-lab readout limited to period phenotype"]},{"year":2020,"claim":"Identified N-glycosylation as a prerequisite for functional GPR176, answering how the receptor reaches the cell surface and why glycan loss diminishes its activity.","evidence":"PNGase F deglycosylation of mouse hypothalamus, site-directed mutagenesis, heterologous cAMP assay","pmids":["32157140"],"confidence":"High","gaps":["Does not address ligand binding or signalling kinetics","Effect shown only on total expression, not trafficking dynamics directly"]},{"year":2022,"claim":"Placed GPR176 within an Nmu/Nms neuropeptide circuit that modulates light-induced phase shifts, extending its role from period-setting to photic entrainment.","evidence":"Microarray, triple Nmu/Nms/Gpr176 knockout mice with behavioural and SCN gene-expression phenotyping","pmids":["35908898"],"confidence":"Medium","gaps":["Direct molecular link between GPR176 and Nmu/Nms signalling unresolved","Single lab"]},{"year":2023,"claim":"Defined a cancer-promoting mechanism: GPR176 binds GNAS via its TM3-ICL2 domain to activate cAMP/PKA and repress mitophagy, connecting the receptor's signalling to colorectal cancer progression.","evidence":"Co-IP, homology modelling, Gpr176-deficient mouse cancer models, cAMP/PKA and mitophagy assays","pmids":["36905238"],"confidence":"Medium","gaps":["Domain mapping relies on homology model only","No reciprocal structural validation of the interaction interface"]},{"year":2024,"claim":"Established GPR176 as a pro-fibrotic effector in hepatic stellate cells across in vitro and in vivo models, broadening its role beyond the brain.","evidence":"siRNA knockdown in primary mouse HSCs and PCLS, Gpr176 knockout in CCl4 and BDL fibrosis models, human tissue IHC","pmids":["38694958"],"confidence":"Medium","gaps":["Downstream signalling in HSCs not fully mapped","Single lab"]},{"year":2024,"claim":"Linked GPR176 mechanistically to fibroblast-to-myofibroblast transition via Smad2 phosphorylation, identifying a TGFβ1-adjacent node it controls without being TGFβ1-regulated itself.","evidence":"siRNA knockdown in NRK-49F renal fibroblasts, Western blot for αSMA/fibronectin/collagen and phospho-Smad2","pmids":["39047914"],"confidence":"Medium","gaps":["How GPR176 modulates Smad2 phosphorylation is undefined","G-protein coupling in fibroblasts not tested"]},{"year":2025,"claim":"Identified an upstream regulatory mechanism whereby exosomal miR-382-5p suppresses GPR176 and disrupts its GNAS interaction to limit angiogenesis in liver metastasis.","evidence":"RNA pull-down, RIP, Co-IP, in vivo/in vitro angiogenesis and vascular permeability assays, exosome characterization","pmids":["40578589"],"confidence":"Medium","gaps":["Direct connection between GPR176-GNAS and CXCR1/CXCR2 levels mechanistically incomplete","Single lab"]},{"year":2025,"claim":"Revealed a neuronal excitability role: GPR176 in PV+ prefrontal interneurons constrains firing by acting on action-potential repolarization, distinct from synaptic effects.","evidence":"In situ expression, shRNA knockdown, whole-cell electrophysiology in PV+ interneurons, behaviour","pmids":["41188983"],"confidence":"Medium","gaps":["Effector channels/G-protein pathway mediating the effect unidentified","Link to cAMP signalling not established here"]},{"year":2026,"claim":"Placed GPR176 downstream of E2F4 transcriptional control and showed it confers ferroptosis resistance via mitophagy suppression, integrating its mitophagy role into a transcriptional-to-cell-death axis in esophageal cancer.","evidence":"ChIP/promoter binding, E2F4 and GPR176 over/knockdown, mitophagy and ferroptosis markers, rescue experiments","pmids":["42253509"],"confidence":"Medium","gaps":["Whether GNAS/cAMP-PKA mediates the ferroptosis effect not tested","Single lab"]},{"year":null,"claim":"The identity of the endogenous activating ligand(s) for GPR176 and the structural basis of its constitutive Gz coupling remain unresolved.","evidence":"Cell-based GzESTY assay detected endogenous ligand activity in brain extracts but did not identify the ligand (preprint)","pmids":[],"confidence":"Low","gaps":["No ligand identity established","Detection in a single preprint cell-based assay","No structure of receptor or receptor-G protein complex"]}],"mechanism_profile":{"molecular_activity":[{"term_id":"GO:0060089","term_label":"molecular transducer activity","supporting_discovery_ids":[0,4]},{"term_id":"GO:0098772","term_label":"molecular function regulator activity","supporting_discovery_ids":[0,4]}],"localization":[{"term_id":"GO:0005886","term_label":"plasma membrane","supporting_discovery_ids":[3]}],"pathway":[{"term_id":"R-HSA-162582","term_label":"Signal Transduction","supporting_discovery_ids":[0,4]},{"term_id":"R-HSA-9909396","term_label":"Circadian clock","supporting_discovery_ids":[0,5]},{"term_id":"R-HSA-1643685","term_label":"Disease","supporting_discovery_ids":[4,6,7]}],"complexes":[],"partners":["GNAS"],"other_free_text":[]}},"prefetch_data":{"uniprot":{"accession":"Q14439","full_name":"G-protein coupled receptor 176","aliases":["HB-954"],"length_aa":515,"mass_kda":57.0,"function":"Orphan receptor involved in normal circadian rhythm behavior. Acts through the G-protein subclass G(z)-alpha and has an agonist-independent basal activity to repress cAMP production","subcellular_location":"Cell membrane","url":"https://www.uniprot.org/uniprotkb/Q14439/entry"},"depmap":{"release":"DepMap","has_data":true,"is_common_essential":false,"resolved_as":"","url":"https://depmap.org/portal/gene/GPR176","classification":"Not Classified","n_dependent_lines":0,"n_total_lines":1208,"dependency_fraction":0.0},"opencell":{"profiled":false,"resolved_as":"","ensg_id":"","cell_line_id":"","localizations":[],"interactors":[],"url":"https://opencell.sf.czbiohub.org/search/GPR176","total_profiled":1310},"omim":[{"mim_id":"612183","title":"G PROTEIN-COUPLED RECEPTOR 176; GPR176","url":"https://www.omim.org/entry/612183"}],"hpa":{"profiled":true,"resolved_as":"","reliability":"","locations":[],"tissue_specificity":"Tissue enhanced","tissue_distribution":"Detected in many","driving_tissues":[{"tissue":"blood vessel","ntpm":34.6},{"tissue":"brain","ntpm":21.9}],"url":"https://www.proteinatlas.org/search/GPR176"},"hgnc":{"alias_symbol":["Gm1012"],"prev_symbol":[]},"alphafold":{"accession":"Q14439","domains":[{"cath_id":"1.20.1070.10","chopping":"32-247_255-342","consensus_level":"high","plddt":83.5075,"start":32,"end":342}],"viewer_url":"https://alphafold.ebi.ac.uk/entry/Q14439","model_url":"https://alphafold.ebi.ac.uk/files/AF-Q14439-F1-model_v6.cif","pae_url":"https://alphafold.ebi.ac.uk/files/AF-Q14439-F1-predicted_aligned_error_v6.png","plddt_mean":66.44},"mouse_models":{"mgi_url":"https://www.informatics.jax.org/marker/summary?nomen=GPR176","jax_strain_url":"https://www.jax.org/strain/search?query=GPR176"},"sequence":{"accession":"Q14439","fasta_url":"https://rest.uniprot.org/uniprotkb/Q14439.fasta","uniprot_url":"https://www.uniprot.org/uniprotkb/Q14439/entry","alphafold_viewer_url":"https://alphafold.ebi.ac.uk/entry/Q14439"}},"corpus_meta":[{"pmid":"26882873","id":"PMC_26882873","title":"Gpr176 is a Gz-linked orphan G-protein-coupled receptor that sets the pace of circadian behaviour.","date":"2016","source":"Nature communications","url":"https://pubmed.ncbi.nlm.nih.gov/26882873","citation_count":68,"is_preprint":false},{"pmid":"36905238","id":"PMC_36905238","title":"GPR176 Promotes Cancer Progression by Interacting with G Protein GNAS to Restrain Cell Mitophagy in Colorectal Cancer.","date":"2023","source":"Advanced science (Weinheim, Baden-Wurttemberg, Germany)","url":"https://pubmed.ncbi.nlm.nih.gov/36905238","citation_count":54,"is_preprint":false},{"pmid":"32157140","id":"PMC_32157140","title":"Identification and functional characterisation of N-linked glycosylation of the orphan G protein-coupled receptor Gpr176.","date":"2020","source":"Scientific reports","url":"https://pubmed.ncbi.nlm.nih.gov/32157140","citation_count":30,"is_preprint":false},{"pmid":"32709014","id":"PMC_32709014","title":"Time-Restricted G-Protein Signaling Pathways via GPR176, Gz, and RGS16 Set the Pace of the Master Circadian Clock in the Suprachiasmatic Nucleus.","date":"2020","source":"International journal of molecular sciences","url":"https://pubmed.ncbi.nlm.nih.gov/32709014","citation_count":16,"is_preprint":false},{"pmid":"28502923","id":"PMC_28502923","title":"G-protein-coupled receptor signaling through Gpr176, Gz, and RGS16 tunes time in the center of the circadian clock [Review].","date":"2017","source":"Endocrine journal","url":"https://pubmed.ncbi.nlm.nih.gov/28502923","citation_count":12,"is_preprint":false},{"pmid":"35908898","id":"PMC_35908898","title":"Nmu/Nms/Gpr176 Triple-Deficient Mice Show Enhanced Light-Resetting of Circadian Locomotor Activity.","date":"2022","source":"Biological & pharmaceutical bulletin","url":"https://pubmed.ncbi.nlm.nih.gov/35908898","citation_count":11,"is_preprint":false},{"pmid":"37079213","id":"PMC_37079213","title":"Oncogenic roles of GPR176 in breast cancer: a potential marker of aggressiveness and a potential target of gene therapy.","date":"2023","source":"Clinical & translational oncology : official publication of the Federation of Spanish Oncology Societies and of the National Cancer Institute of Mexico","url":"https://pubmed.ncbi.nlm.nih.gov/37079213","citation_count":8,"is_preprint":false},{"pmid":"37584712","id":"PMC_37584712","title":"The promoting effects of GPR176 expression on proliferation, chemoresistance, lipogenesis and invasion of oesophageal cancer.","date":"2023","source":"Journal of cancer research and clinical oncology","url":"https://pubmed.ncbi.nlm.nih.gov/37584712","citation_count":6,"is_preprint":false},{"pmid":"38835234","id":"PMC_38835234","title":"The oncogenic roles of GPR176 in ovarian cancer: a molecular target for aggressiveness and gene therapy.","date":"2024","source":"Journal of obstetrics and gynaecology : the journal of the Institute of Obstetrics and Gynaecology","url":"https://pubmed.ncbi.nlm.nih.gov/38835234","citation_count":6,"is_preprint":false},{"pmid":"37284492","id":"PMC_37284492","title":"Exploring the Correlation Between GPR176, a Potential Target Gene of Gastric Cancer, and Immune Cell Infiltration.","date":"2023","source":"Pharmacogenomics and personalized medicine","url":"https://pubmed.ncbi.nlm.nih.gov/37284492","citation_count":6,"is_preprint":false},{"pmid":"38694958","id":"PMC_38694958","title":"Orphan receptor GPR176 in hepatic stellate cells exerts a profibrotic role in chronic liver disease.","date":"2024","source":"JHEP reports : innovation in hepatology","url":"https://pubmed.ncbi.nlm.nih.gov/38694958","citation_count":4,"is_preprint":false},{"pmid":"39047914","id":"PMC_39047914","title":"GPR176 promotes fibroblast-to-myofibroblast transition in organ fibrosis progression.","date":"2024","source":"Biochimica et biophysica acta. Molecular cell research","url":"https://pubmed.ncbi.nlm.nih.gov/39047914","citation_count":3,"is_preprint":false},{"pmid":"40578589","id":"PMC_40578589","title":"Exosomal miR-382-5p prevents pre-metastatic niche formation by inhibiting GPR176/GNAS-CXCR1/CXCR2 axis in colorectal cancer liver metastasis.","date":"2025","source":"Cellular signalling","url":"https://pubmed.ncbi.nlm.nih.gov/40578589","citation_count":3,"is_preprint":false},{"pmid":"41188983","id":"PMC_41188983","title":"Gpr176 modulates the firing pattern of parvalbumin-positive interneurons in the orbitofrontal cortex of mouse.","date":"2025","source":"Molecular brain","url":"https://pubmed.ncbi.nlm.nih.gov/41188983","citation_count":0,"is_preprint":false},{"pmid":"41574720","id":"PMC_41574720","title":"GPR176 enhances the epithelial‑mesenchymal transition in gastric cancer cells by activating the PI3K/AKT/mTOR signaling pathway.","date":"2026","source":"Oncology reports","url":"https://pubmed.ncbi.nlm.nih.gov/41574720","citation_count":0,"is_preprint":false},{"pmid":"41484214","id":"PMC_41484214","title":"Establishing FDA-approved oncology drugs as GPR176 inhibitor through homology modelling, molecular docking, MMGBSA, DFT, and molecular dynamics simulation.","date":"2026","source":"Scientific reports","url":"https://pubmed.ncbi.nlm.nih.gov/41484214","citation_count":0,"is_preprint":false},{"pmid":"41969458","id":"PMC_41969458","title":"GPR176 represses mitophagy to promote the progression of osteosarcoma by facilitating mTORC1 activity via PI3K-AKT pathway.","date":"2026","source":"Translational cancer research","url":"https://pubmed.ncbi.nlm.nih.gov/41969458","citation_count":0,"is_preprint":false},{"pmid":"42253509","id":"PMC_42253509","title":"E2F4 Mediates Mitophagy to Inhibit Ferroptosis in Esophageal Cancer Cells by Activating GPR176.","date":"2026","source":"Human mutation","url":"https://pubmed.ncbi.nlm.nih.gov/42253509","citation_count":0,"is_preprint":false},{"pmid":null,"id":"bio_10.1101_2024.07.26.605282","title":"Gz Enhanced Signal Transduction assaY (G  <sub>Z</sub>  ESTY) for GPCR deorphanization","date":"2024-07-26","source":"bioRxiv","url":"https://doi.org/10.1101/2024.07.26.605282","citation_count":0,"is_preprint":true}],"cost":{"stage1":{"model":"claude-sonnet-4-6","input_tokens":12533,"output_tokens":2842,"usd":0.040114,"stage1_stop_reason":"end_turn"},"stage2":{"model":"claude-opus-4-8","input_tokens":10162,"output_tokens":3297,"usd":0.066617,"stage2_stop_reason":"end_turn"},"total_usd":0.106731,"stage1_batch_id":"msgbatch_01DUPACZi7AAWwuZiUjseQF4","stage2_batch_id":"msgbatch_018yt7chE3f48PEFtDwtj6AE","note":"batch pricing = 50% of standard"},"stage1_raw":"```json\n{\n  \"discoveries\": [\n    {\n      \"year\": 2016,\n      \"finding\": \"GPR176 is an SCN-enriched orphan GPCR that represses cAMP signalling in an agonist-independent (constitutively active) manner, and couples to the unique G-protein subclass Gz (not canonical Gi) to reduce cAMP production and set the pace of circadian behaviour.\",\n      \"method\": \"Genetic knockout mice, heterologous expression assays, cAMP measurement, G-protein coupling analysis\",\n      \"journal\": \"Nature Communications\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — reciprocal genetic and biochemical approaches in vivo and in vitro, replicated in subsequent reviews and follow-up papers\",\n      \"pmids\": [\"26882873\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2016,\n      \"finding\": \"GPR176 acts independently of and in parallel to the Vipr2 GPCR pathway in the SCN, as established by genetic epistasis in knockout animals.\",\n      \"method\": \"Genetic epistasis analysis using Gpr176 and Vipr2 mutant mice\",\n      \"journal\": \"Nature Communications\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — genetic epistasis in vivo, single lab, circadian period phenotype readout\",\n      \"pmids\": [\"26882873\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2020,\n      \"finding\": \"GPR176 undergoes N-linked glycosylation at four conserved asparagine residues in its N-terminal region; mutation of these residues reduces protein expression and attenuates cAMP-repressive activity in cells, establishing N-glycosylation as a prerequisite for efficient expression of functional GPR176.\",\n      \"method\": \"Peptide-N-glycosidase F treatment of mouse hypothalamus extracts, site-directed mutagenesis of N-glycosylation sites, heterologous expression, cAMP assay\",\n      \"journal\": \"Scientific Reports\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 / Moderate — in vitro enzymatic deglycosylation, site-directed mutagenesis with functional cAMP readout, endogenous tissue validation, single lab with multiple orthogonal methods\",\n      \"pmids\": [\"32157140\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2020,\n      \"finding\": \"N-glycosylation of GPR176 is required for proper cell-surface expression; deficient N-glycosylation does not directly compromise the intrinsic agonist-independent cAMP-repressive activity but reduces it indirectly by lowering total protein levels.\",\n      \"method\": \"Site-directed mutagenesis of N-glycosylation sites, heterologous expression, cAMP assay\",\n      \"journal\": \"Scientific Reports\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 1 / Moderate — mutagenesis with functional readout, single lab\",\n      \"pmids\": [\"32157140\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2023,\n      \"finding\": \"GPR176 physically interacts with G protein GNAS intracellularly via its transmembrane helix 3–intracellular loop 2 domain; this GPR176/GNAS complex activates the cAMP/PKA signalling pathway and inhibits mitophagy via the cAMP/PKA/BNIP3L axis to promote colorectal cancer progression.\",\n      \"method\": \"Co-immunoprecipitation, homology modelling, in vitro and in vivo cancer models with Gpr176-deficient mice, cAMP/PKA assay, mitophagy assays\",\n      \"journal\": \"Advanced Science\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — Co-IP and genetic mouse model with mechanistic pathway readout, single lab, domain mapping by homology model only\",\n      \"pmids\": [\"36905238\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2022,\n      \"finding\": \"Gpr176 knockout in mice leads to upregulation of Nmu and Nms mRNA in the SCN; triple knockout of Nmu/Nms/Gpr176 results in enhanced light-induced phase shifts and reduced Per1 and cFos induction by light, indicating a functional interaction among Nmu, Nms, and Gpr176 in modulating light-induced circadian phase shifts.\",\n      \"method\": \"Microarray analysis, triple knockout mouse generation and behavioural phenotyping, SCN gene expression analysis\",\n      \"journal\": \"Biological & Pharmaceutical Bulletin\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — genetic epistasis with triple KO mice and molecular readouts, single lab\",\n      \"pmids\": [\"35908898\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2024,\n      \"finding\": \"GPR176 is induced in activated hepatic stellate cells (HSCs) and plays a profibrotic role: siRNA-mediated knockdown reduces fibrogenic characteristics in primary mouse HSCs and precision-cut liver slices; Gpr176 knockout mice develop less severe fibrosis in CCl4 and bile duct ligation models.\",\n      \"method\": \"siRNA knockdown in primary mouse HSCs and PCLS, GPR176 knockout mouse fibrosis models (CCl4 and BDL), immunohistochemistry of human CLD tissue\",\n      \"journal\": \"JHEP Reports\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — loss-of-function both in vitro and in vivo with defined fibrosis phenotype, single lab, two orthogonal loss-of-function models\",\n      \"pmids\": [\"38694958\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2024,\n      \"finding\": \"GPR176 promotes fibroblast-to-myofibroblast transition in fibrosis: Gpr176 expression is increased in fibrotic lungs, kidneys, liver, and heart; siRNA knockdown of Gpr176 in rat renal fibroblasts reduces TGFβ1-induced expression of αSMA, fibronectin, and collagen, and attenuates Smad2 phosphorylation, without being itself regulated by TGFβ1.\",\n      \"method\": \"Gene expression analysis in fibrosis mouse models, siRNA knockdown in NRK-49F cells, Western blot for αSMA/fibronectin/collagen and phospho-Smad2\",\n      \"journal\": \"Biochimica et Biophysica Acta – Molecular Cell Research\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — siRNA loss-of-function with defined molecular pathway readout (Smad2 phosphorylation), single lab\",\n      \"pmids\": [\"39047914\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2025,\n      \"finding\": \"Exosomal miR-382-5p downregulates GPR176 expression and disrupts its interaction with GNAS in the liver, thereby reducing CXCR1/CXCR2 levels and suppressing angiogenesis and vascular permeability in colorectal cancer liver metastasis models.\",\n      \"method\": \"RNA pull-down, RNA immunoprecipitation (RIP), Co-IP, in vivo and in vitro angiogenesis/vascular permeability assays, exosome isolation and characterization\",\n      \"journal\": \"Cellular Signalling\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — Co-IP and RIP with functional in vivo validation, single lab\",\n      \"pmids\": [\"40578589\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2024,\n      \"finding\": \"A cell-based assay (GzESTY) detected the presence of endogenous ligands for GPR176 in brain extracts, providing functional evidence that endogenous activating ligands for GPR176 exist in brain tissue.\",\n      \"method\": \"Cell-based Gz-coupled signal transduction assay (GzESTY) with brain extract fractions\",\n      \"journal\": \"bioRxiv (preprint)\",\n      \"confidence\": \"Low\",\n      \"confidence_rationale\": \"Tier 3 / Weak — single cell-based assay in a preprint, no ligand identity established, single lab\",\n      \"pmids\": [\"bio_10.1101_2024.07.26.605282\"],\n      \"is_preprint\": true\n    },\n    {\n      \"year\": 2025,\n      \"finding\": \"Gpr176 is expressed predominantly in parvalbumin-positive (PV+) interneurons of the prefrontal cortex; knockdown of Gpr176 increases firing output of PV+ interneurons by altering membrane potential changes during the repolarizing phase of action potentials, without affecting synaptic activity.\",\n      \"method\": \"In situ expression analysis, shRNA-mediated knockdown, whole-cell electrophysiology in PV+ interneurons, behavioural assays\",\n      \"journal\": \"Molecular Brain\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — direct electrophysiological readout with targeted knockdown in specific cell type, single lab\",\n      \"pmids\": [\"41188983\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2026,\n      \"finding\": \"E2F4 directly binds the GPR176 promoter and transcriptionally activates GPR176 expression; elevated GPR176 suppresses mitophagy and confers resistance to ferroptosis in esophageal cancer cells, and GPR176 overexpression abrogates the enhanced mitophagy and ferroptosis induced by E2F4 depletion.\",\n      \"method\": \"ChIP/promoter binding assay, overexpression and knockdown of E2F4 and GPR176, mitophagy markers (MMP, ROS, autophagy proteins), ferroptosis markers (MDA, Fe2+, lipid ROS), rescue experiments\",\n      \"journal\": \"Human Mutation\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — direct promoter binding assay plus rescue experiments establishing epistasis, single lab\",\n      \"pmids\": [\"42253509\"],\n      \"is_preprint\": false\n    }\n  ],\n  \"current_model\": \"GPR176 is a constitutively active (agonist-independent), N-glycosylated orphan class-A GPCR that couples to the Gz G-protein subclass to suppress cAMP/PKA signalling; in the SCN it sets circadian period and modulates light-induced phase shifts via this Gz–cAMP axis, while in peripheral tissues it promotes fibrosis through Smad2-dependent fibroblast-to-myofibroblast transition and drives cancer progression by recruiting GNAS via its TM3–ICL2 domain to activate cAMP/PKA/BNIP3L signalling and repress mitophagy.\"\n}\n```","stage2_raw":"{\n  \"mechanistic_narrative\": \"GPR176 is a constitutively active, SCN-enriched orphan class-A GPCR that suppresses cAMP signalling in an agonist-independent manner by coupling to the atypical Gz G-protein subclass, thereby setting the pace of circadian behaviour [#0]. This Gz–cAMP activity operates in parallel to the Vipr2 pathway in the SCN [#1] and is also embedded in a regulatory circuit with the neuropeptides Nmu/Nms that tunes light-induced phase shifts [#5]. Efficient cell-surface expression of functional GPR176 depends on N-linked glycosylation at four conserved N-terminal asparagine residues, loss of which lowers protein levels and indirectly attenuates its cAMP-repressive output [#2, #3]. Beyond the clock, GPR176 functions as a pro-fibrotic and pro-tumorigenic effector: it drives fibroblast-to-myofibroblast transition through Smad2 phosphorylation and induction of αSMA, fibronectin, and collagen, and promotes fibrosis across multiple organs [#6, #7]. In cancer it physically engages the G-protein GNAS via its transmembrane helix 3–intracellular loop 2 region to activate cAMP/PKA signalling and repress mitophagy through the BNIP3L axis, promoting colorectal cancer progression [#4], a process antagonized by exosomal miR-382-5p that downregulates GPR176 and disrupts the GPR176–GNAS interaction [#8]. GPR176 is transcriptionally activated by E2F4 and confers ferroptosis resistance by suppressing mitophagy in esophageal cancer [#11]. In the prefrontal cortex it is expressed in parvalbumin-positive interneurons where it constrains their firing output [#10].\",\n  \"teleology\": [\n    {\n      \"year\": 2016,\n      \"claim\": \"Established GPR176's core molecular identity: an orphan GPCR that represses cAMP constitutively through the unusual Gz subclass to set circadian period, answering how this SCN-enriched receptor controls the clock without a known ligand.\",\n      \"evidence\": \"Knockout mice, heterologous expression, cAMP measurement and G-protein coupling analysis\",\n      \"pmids\": [\"26882873\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"No endogenous ligand identified\", \"Mechanism of constitutive activity at structural level unresolved\"]\n    },\n    {\n      \"year\": 2016,\n      \"claim\": \"Showed GPR176 acts in a pathway genetically separate from Vipr2, clarifying that it is a non-redundant input to SCN timekeeping.\",\n      \"evidence\": \"Genetic epistasis with Gpr176 and Vipr2 mutant mice\",\n      \"pmids\": [\"26882873\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Molecular basis of pathway independence not defined\", \"Single-lab readout limited to period phenotype\"]\n    },\n    {\n      \"year\": 2020,\n      \"claim\": \"Identified N-glycosylation as a prerequisite for functional GPR176, answering how the receptor reaches the cell surface and why glycan loss diminishes its activity.\",\n      \"evidence\": \"PNGase F deglycosylation of mouse hypothalamus, site-directed mutagenesis, heterologous cAMP assay\",\n      \"pmids\": [\"32157140\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Does not address ligand binding or signalling kinetics\", \"Effect shown only on total expression, not trafficking dynamics directly\"]\n    },\n    {\n      \"year\": 2022,\n      \"claim\": \"Placed GPR176 within an Nmu/Nms neuropeptide circuit that modulates light-induced phase shifts, extending its role from period-setting to photic entrainment.\",\n      \"evidence\": \"Microarray, triple Nmu/Nms/Gpr176 knockout mice with behavioural and SCN gene-expression phenotyping\",\n      \"pmids\": [\"35908898\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Direct molecular link between GPR176 and Nmu/Nms signalling unresolved\", \"Single lab\"]\n    },\n    {\n      \"year\": 2023,\n      \"claim\": \"Defined a cancer-promoting mechanism: GPR176 binds GNAS via its TM3-ICL2 domain to activate cAMP/PKA and repress mitophagy, connecting the receptor's signalling to colorectal cancer progression.\",\n      \"evidence\": \"Co-IP, homology modelling, Gpr176-deficient mouse cancer models, cAMP/PKA and mitophagy assays\",\n      \"pmids\": [\"36905238\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Domain mapping relies on homology model only\", \"No reciprocal structural validation of the interaction interface\"]\n    },\n    {\n      \"year\": 2024,\n      \"claim\": \"Established GPR176 as a pro-fibrotic effector in hepatic stellate cells across in vitro and in vivo models, broadening its role beyond the brain.\",\n      \"evidence\": \"siRNA knockdown in primary mouse HSCs and PCLS, Gpr176 knockout in CCl4 and BDL fibrosis models, human tissue IHC\",\n      \"pmids\": [\"38694958\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Downstream signalling in HSCs not fully mapped\", \"Single lab\"]\n    },\n    {\n      \"year\": 2024,\n      \"claim\": \"Linked GPR176 mechanistically to fibroblast-to-myofibroblast transition via Smad2 phosphorylation, identifying a TGFβ1-adjacent node it controls without being TGFβ1-regulated itself.\",\n      \"evidence\": \"siRNA knockdown in NRK-49F renal fibroblasts, Western blot for αSMA/fibronectin/collagen and phospho-Smad2\",\n      \"pmids\": [\"39047914\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"How GPR176 modulates Smad2 phosphorylation is undefined\", \"G-protein coupling in fibroblasts not tested\"]\n    },\n    {\n      \"year\": 2025,\n      \"claim\": \"Identified an upstream regulatory mechanism whereby exosomal miR-382-5p suppresses GPR176 and disrupts its GNAS interaction to limit angiogenesis in liver metastasis.\",\n      \"evidence\": \"RNA pull-down, RIP, Co-IP, in vivo/in vitro angiogenesis and vascular permeability assays, exosome characterization\",\n      \"pmids\": [\"40578589\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Direct connection between GPR176-GNAS and CXCR1/CXCR2 levels mechanistically incomplete\", \"Single lab\"]\n    },\n    {\n      \"year\": 2025,\n      \"claim\": \"Revealed a neuronal excitability role: GPR176 in PV+ prefrontal interneurons constrains firing by acting on action-potential repolarization, distinct from synaptic effects.\",\n      \"evidence\": \"In situ expression, shRNA knockdown, whole-cell electrophysiology in PV+ interneurons, behaviour\",\n      \"pmids\": [\"41188983\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Effector channels/G-protein pathway mediating the effect unidentified\", \"Link to cAMP signalling not established here\"]\n    },\n    {\n      \"year\": 2026,\n      \"claim\": \"Placed GPR176 downstream of E2F4 transcriptional control and showed it confers ferroptosis resistance via mitophagy suppression, integrating its mitophagy role into a transcriptional-to-cell-death axis in esophageal cancer.\",\n      \"evidence\": \"ChIP/promoter binding, E2F4 and GPR176 over/knockdown, mitophagy and ferroptosis markers, rescue experiments\",\n      \"pmids\": [\"42253509\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Whether GNAS/cAMP-PKA mediates the ferroptosis effect not tested\", \"Single lab\"]\n    },\n    {\n      \"year\": null,\n      \"claim\": \"The identity of the endogenous activating ligand(s) for GPR176 and the structural basis of its constitutive Gz coupling remain unresolved.\",\n      \"evidence\": \"Cell-based GzESTY assay detected endogenous ligand activity in brain extracts but did not identify the ligand (preprint)\",\n      \"pmids\": [],\n      \"confidence\": \"Low\",\n      \"gaps\": [\"No ligand identity established\", \"Detection in a single preprint cell-based assay\", \"No structure of receptor or receptor-G protein complex\"]\n    }\n  ],\n  \"mechanism_profile\": {\n    \"molecular_activity\": [\n      {\"term_id\": \"GO:0060089\", \"supporting_discovery_ids\": [0, 4]},\n      {\"term_id\": \"GO:0098772\", \"supporting_discovery_ids\": [0, 4]}\n    ],\n    \"localization\": [\n      {\"term_id\": \"GO:0005886\", \"supporting_discovery_ids\": [3]}\n    ],\n    \"pathway\": [\n      {\"term_id\": \"R-HSA-162582\", \"supporting_discovery_ids\": [0, 4]},\n      {\"term_id\": \"R-HSA-9909396\", \"supporting_discovery_ids\": [0, 5]},\n      {\"term_id\": \"R-HSA-1643685\", \"supporting_discovery_ids\": [4, 6, 7]}\n    ],\n    \"complexes\": [],\n    \"partners\": [\"GNAS\"],\n    \"other_free_text\": []\n  }\n}","audit_flag":null,"evaluation":{"pairwise":"win","faith_supported":7,"faith_total":7,"faith_pct":100.0}}