{"gene":"GUCY2F","run_date":"2026-04-28T18:06:53","timeline":{"discoveries":[{"year":1995,"finding":"RetGC-2 (GC-F/GUCY2F) was cloned from human retinal cDNA and shown to encode a membrane guanylyl cyclase expressed specifically in photoreceptor cells; recombinant RetGC-2 expressed in HEK293 cells displays guanylyl cyclase activity that is stimulated by the Ca2+-binding activator p24 (GCAP-2) and inhibited by Ca2+ with an EC50 of 50-100 nM.","method":"cDNA cloning, in situ hybridization, recombinant expression in HEK293 cells, in vitro guanylyl cyclase activity assay","journal":"Proceedings of the National Academy of Sciences of the United States of America","confidence":"High","confidence_rationale":"Tier 1 — reconstituted enzymatic activity in heterologous cells with direct biochemical assay; foundational paper with 229 citations","pmids":["7777544"],"is_preprint":false},{"year":1995,"finding":"GC-F (GUCY2F) was isolated from a rat eye cDNA library and shown to encode a membrane guanylyl cyclase with an extracellular domain, single transmembrane domain, kinase-like domain, and catalytic cyclase domain; overexpression in COS cells conferred guanylyl cyclase activity, but known extracellular ligands for other GC receptors did not stimulate it, classifying it as an orphan receptor.","method":"cDNA cloning, COS cell overexpression, guanylyl cyclase activity assay, peptide ligand stimulation assay","journal":"Proceedings of the National Academy of Sciences of the United States of America","confidence":"High","confidence_rationale":"Tier 1 — direct enzymatic assay in heterologous cells; foundational paper with 215 citations","pmids":["7831337"],"is_preprint":false},{"year":1997,"finding":"ROS-GC2 (the bovine ortholog of GUCY2F) was cloned from bovine retina, shown to be specifically expressed in retina, and demonstrated to be modulated in low Ca2+ by the calmodulin-like Ca2+-binding protein GCAP-2; it also contains a unique five-amino-acid signature at its C-terminus.","method":"cDNA cloning, retinal expression analysis, in vitro reconstitution with GCAP-2, guanylyl cyclase activity assay","journal":"Biochemical and biophysical research communications","confidence":"High","confidence_rationale":"Tier 1 — reconstituted Ca2+-dependent regulation in vitro with purified proteins","pmids":["9175772"],"is_preprint":false},{"year":1998,"finding":"The kinase homology domain (KHD) of RetGC-2 specifies the affinity and cooperativity of interaction with GCAP-2: RetGC-2 interacts cooperatively and with high affinity with GCAP-2, in contrast to RetGC-1 which interacts noncooperatively with low affinity; this was mapped using RetGC-1/RetGC-2 chimeras and a trypsin protection assay showing GCAP-2 binds constitutively to the KHD.","method":"Trypsin/protease protection assay, RetGC-1/RetGC-2 chimeric protein analysis, guanylyl cyclase activity assay","journal":"Biochemistry","confidence":"High","confidence_rationale":"Tier 1 — domain mapping with chimeric proteins and direct binding assay; multiple orthogonal methods","pmids":["9698373"],"is_preprint":false},{"year":1998,"finding":"CD-GCAP activates ROS-GC2 (GUCY2F ortholog), but with approximately 10-fold weaker activation than ROS-GC1; the CD-GCAP-regulated signaling switch on ROS-GC1 was mapped to amino acid segment 736-1053 through deletion mutants, hybrid constructs, and heterologous cyclase reconstruction.","method":"Recombinant protein expression, deletion mutagenesis, chimeric cyclase construction, in vitro guanylyl cyclase activity assay","journal":"Biochemical and biophysical research communications","confidence":"High","confidence_rationale":"Tier 1 — domain mapping by mutagenesis and reconstitution with direct enzymatic readout","pmids":["9439621"],"is_preprint":false},{"year":1995,"finding":"GCAP-2 (p24) activates both RetGC-1 and RetGC-2 (GUCY2F) in a Ca2+-sensitive manner in vitro; Ca2+ inhibits activation with an EC50 near 200 nM and Hill coefficient of 1.7; recombinant GCAP-2 expressed in HEK293 cells effectively stimulates photoreceptor guanylyl cyclase.","method":"Protein purification, in vitro guanylyl cyclase activity assay, recombinant expression in HEK293 cells, antibody inhibition","journal":"The Journal of biological chemistry","confidence":"High","confidence_rationale":"Tier 1 — reconstituted in vitro with purified proteins; foundational paper with 271 citations","pmids":["7559656"],"is_preprint":false},{"year":1999,"finding":"GCAPs bind constitutively to an intracellular domain of RetGC-2 (GUCY2F); in the absence of Ca2+, GCAP stimulates cyclase activity, and in the presence of Ca2+, it inhibits cyclase activity; proper RetGC and GCAP functioning is necessary for photoreceptor viability.","method":"Biochemical binding assays, in vitro guanylyl cyclase activity assay, genetic evidence from disease mutations","journal":"Methods (San Diego, Calif.)","confidence":"High","confidence_rationale":"Tier 1-2 — multiple biochemical lines of evidence synthesized from primary studies; strong evidence replicated across labs","pmids":["10581151"],"is_preprint":false},{"year":1999,"finding":"GCAP-2 functional domain mapping demonstrated that substituting specific domains of GCAP-2 with corresponding fragments from non-RetGC-regulating proteins (neurocalcin, recoverin) abolishes its ability to regulate RetGC-2 (GUCY2F) expressed in HEK293 cells; three regions are essential: residues 78-113 (determines Ca2+ activation vs. inhibition polarity), residues 29-48 (EF-1 motif, required for both activation and inhibition), and residues 171-189 (contributes to activation).","method":"GCAP-2 deletion mutants, chimeric protein construction, in vitro guanylyl cyclase activity assay with recombinant RetGC-1 and RetGC-2 in HEK293 cells","journal":"The Journal of biological chemistry","confidence":"High","confidence_rationale":"Tier 1 — systematic mutagenesis/chimera approach with direct enzymatic readout for both RetGC-1 and RetGC-2","pmids":["10196158"],"is_preprint":false},{"year":1999,"finding":"Neurocalcin, a Ca2+-binding protein structurally related to GCAPs, stimulates ROS-GC1 in a Ca2+-dependent manner (EC50 ~20 µM) but does not influence the activity of ROS-GC2 (GUCY2F ortholog), demonstrating substrate specificity between the two retinal cyclases for this regulator.","method":"Recombinant protein expression, in vitro guanylyl cyclase activity assay, domain mapping with deletion mutants","journal":"Biochemistry","confidence":"Medium","confidence_rationale":"Tier 1 — direct in vitro enzymatic assay; single lab demonstrating specificity for ROS-GC2","pmids":["10504230"],"is_preprint":false},{"year":1999,"finding":"ROS-GC2 (GUCY2F) was not detected in the pineal gland (unlike ROS-GC1), establishing that the two retinal cyclases have distinct expression patterns beyond the retina; the alpha2D/A-adrenergic receptor-linked signaling system in pinealocytes uses exclusively ROS-GC1 and not ROS-GC2.","method":"Immunohistochemistry, molecular cloning, immunoblotting, biochemical fractionation","journal":"Biochemistry","confidence":"Medium","confidence_rationale":"Tier 2 — direct immunodetection and fractionation; single lab but multiple orthogonal methods","pmids":["10821676"],"is_preprint":false},{"year":1999,"finding":"Disruption of GC-E (RetGC1/GUCY2D) gene in mice left GC-F (GUCY2F) expression unchanged, demonstrating that GC-F is expressed independently; however, cones progressively disappeared by 5 weeks, establishing that GC-E (not GC-F) is essential for cone photoreceptor survival.","method":"Gene knockout mouse, electroretinography, retinal histology, immunoblotting","journal":"The Journal of neuroscience : the official journal of the Society for Neuroscience","confidence":"High","confidence_rationale":"Tier 2 — clean knockout with defined cellular phenotype and multiple readouts; 159 citations","pmids":["10407028"],"is_preprint":false},{"year":2007,"finding":"Double knockout of Gucy2e and Gucy2f (GC1/GC2) prevented transport of rod PDE6 to rod outer segments, while single Gucy2f knockout alone did not prevent transport of transducin, GRK1, or rhodopsin to rod outer segments; this established that GC-bearing membranes co-transport peripheral membrane proteins in vesicles and that GC1 and GC2 have overlapping roles in rod outer segment protein transport.","method":"Gucy2e/Gucy2f double knockout mice, immunofluorescence, subcellular protein localization assays","journal":"Vision research","confidence":"High","confidence_rationale":"Tier 2 — genetic epistasis using double knockout with defined transport phenotype and immunolocalization","pmids":["17949773"],"is_preprint":false},{"year":2009,"finding":"GC1 (GC-E) undergoes autophosphorylation at residues Ser-530, Ser-532, Ser-533, and Ser-538 within the kinase homology domain in vivo (light- and signal transduction-independent); mutations in the putative Mg2+ binding site of the KH domain abolished phosphorylation and dramatically reduced GC activity, demonstrating that a functional KH domain is essential for cGMP production. Autophosphorylation itself does not regulate GC1 activity.","method":"Mass spectrometry phosphorylation mapping, site-directed mutagenesis of KH domain, in vitro guanylyl cyclase activity assay","journal":"The Journal of biological chemistry","confidence":"High","confidence_rationale":"Tier 1 — mass spectrometry identification plus mutagenesis with direct enzymatic assay; applies to GC1 but establishes principle for the KH domain relevant to GC-F (RetGC-2) as well","pmids":["19901021"],"is_preprint":false},{"year":2015,"finding":"ROS-GC2 (GUCY2F ortholog) is activated by bicarbonate in a Ca2+- and GCAP-independent manner (ED50 ~39 mM), with bicarbonate stimulation being more powerful in the presence of GCAP1 or GCAP2 at low Ca2+; this reveals a novel regulatory input to the ROS-GC system beyond Ca2+/GCAP signaling.","method":"Recombinant protein guanylyl cyclase activity assay, photoreceptor electrophysiology (circulating current and flash response measurements)","journal":"The Journal of biological chemistry","confidence":"High","confidence_rationale":"Tier 1 — direct in vitro enzymatic assay plus electrophysiological validation in native photoreceptors","pmids":["25767116"],"is_preprint":false},{"year":2018,"finding":"RD3 (retinal degeneration protein 3) inhibits both GC-E and GC-F (GUCY2F) and is involved in their transport from inner to outer segments; additionally, RD3 directly interacts with guanylate kinase (demonstrated by back-scattering interferometry) and co-localizes with guanylate kinase in photoreceptor inner segments, revealing a role in the nucleotide cycle.","method":"Back-scattering interferometry (direct interaction assay), immunohistochemistry, guanylyl cyclase activity assay, mouse retina sections","journal":"Frontiers in molecular neuroscience","confidence":"Medium","confidence_rationale":"Tier 2 — direct interaction measured by back-scattering interferometry plus co-localization; single lab","pmids":["29515371"],"is_preprint":false},{"year":2012,"finding":"Gucy2f knockdown in zebrafish using morpholino oligonucleotides (blocking translation or splicing) resulted in significantly reduced visual function (optomotor response) and histological changes including loss and shortening of cone and rod outer segments, establishing GUCY2F as required for photoreceptor outer segment integrity and visual function in vivo.","method":"Zebrafish morpholino knockdown, optomotor assay, retinal histology","journal":"European journal of human genetics : EJHG","confidence":"Medium","confidence_rationale":"Tier 2 — loss-of-function with defined cellular phenotype in vertebrate model; morpholino approach has known caveats","pmids":["22378290"],"is_preprint":false},{"year":2009,"finding":"Overexpression of gucy2F throughout the zebrafish nervous system via Gal4-UAS system caused multiple defects including loss of forebrain neurons, demonstrating that proper control of cGMP production by GUCY2F is important for neuronal survival.","method":"Zebrafish gain-of-function screen using MMLV insertion with Gal4-UAS overexpression system, histological analysis of forebrain neurons","journal":"Molecular genetics and genomics : MGG","confidence":"Medium","confidence_rationale":"Tier 2 — gain-of-function with defined cellular phenotype; single lab, single method","pmids":["19221799"],"is_preprint":false},{"year":1996,"finding":"The GC-F gene (Gucy2f) was mapped to the X chromosome in mice (chromosome X) and the human homolog was localized to Xq22 by fluorescence in situ hybridization; genomic organization analysis showed conservation of exon-intron boundaries with other guanylyl cyclase genes.","method":"Mouse interspecific backcross analysis, fluorescence in situ hybridization (FISH), genomic library screening","journal":"Genomics","confidence":"Medium","confidence_rationale":"Tier 2 — direct chromosomal localization by FISH and genetic mapping","pmids":["8838319"],"is_preprint":false},{"year":2023,"finding":"Evolutionary analysis across vertebrates showed that GC-F (GUCY2F) is absent in several clades (reptiles, birds, marsupials); in species lacking GC-F, visual function is compensated by an increased number of GCAPs, while in nocturnal/visually impaired species, GC-F loss is paralleled by GCAP inactivation, indicating GC-E and GC-F together regulate phototransduction cGMP synthesis with partially compensatory roles.","method":"Comparative genomics, phylogenetic analysis across vertebrate species","journal":"Frontiers in molecular neuroscience","confidence":"Low","confidence_rationale":"Tier 4 — computational/evolutionary analysis; no direct biochemical experiment","pmids":["37007786"],"is_preprint":false},{"year":2023,"finding":"In an AAV-CRISPR 'ablate and replace' study, Gucy2f-/- mice served as a model where loss of GC-F alone allowed testing of GUCY2D (GC-E/RetGC1) rescue vectors; Gucy2e+/-:Gucy2f-/- mice were used to optimize vector doses for CORD6 therapy, confirming that GC-F contributes to but is not solely required for photoreceptor function when GC-E is partially present.","method":"Gucy2f knockout mice, AAV-CRISPR-Cas9 delivery, electroretinography, retinal structure analysis","journal":"Molecular therapy. Methods & clinical development","confidence":"Medium","confidence_rationale":"Tier 2 — defined knockout phenotype with multiple functional readouts; focused on GC-E therapy but uses Gucy2f-/- as genetic tool","pmids":["37361352"],"is_preprint":false}],"current_model":"GUCY2F (RetGC-2/GC-F/ROS-GC2) is a photoreceptor-specific membrane guanylyl cyclase that synthesizes cGMP during phototransduction recovery; it is constitutively associated with its kinase homology domain (KHD) via GCAP-1 and GCAP-2, which inhibit cyclase activity at high Ca2+ (dark-adapted ~500 nM) and activate it at low Ca2+ (light-adapted <100 nM) through conformational changes in the RetGC/GCAP complex; the enzyme is also stimulated by bicarbonate in a GCAP-independent manner, is inhibited by RD3 (which also facilitates its transport to outer segments), and together with GC-E is required for the proper outer-segment delivery of rod phototransduction proteins, with GC-F loss resulting in compromised photoreceptor outer segment integrity and visual dysfunction."},"narrative":{"teleology":[{"year":1995,"claim":"Identification of GUCY2F as a second retinal membrane guanylyl cyclase resolved whether photoreceptors possess a single or multiple cGMP-synthesizing enzymes and established that GUCY2F is Ca²⁺-regulated via GCAP-2.","evidence":"cDNA cloning from human and rat retinal libraries, recombinant expression in HEK293/COS cells with direct cyclase activity assays","pmids":["7777544","7831337","7559656"],"confidence":"High","gaps":["Endogenous extracellular ligand status unresolved (classified as orphan receptor)","Relative contribution of GUCY2F versus GC-E to total retinal cGMP production unknown","Regulation by GCAP-1 not yet tested for GUCY2F"]},{"year":1996,"claim":"Chromosomal mapping of GUCY2F to human Xq22 established its X-linked inheritance pattern and enabled future linkage studies for retinal disease.","evidence":"FISH and mouse interspecific backcross analysis","pmids":["8838319"],"confidence":"Medium","gaps":["No disease-causing mutations identified at this locus","Regulatory elements and tissue-specific promoter not characterized"]},{"year":1998,"claim":"Domain-mapping of the GCAP-2/RetGC-2 interaction established that the kinase homology domain determines the high-affinity, cooperative binding of GCAP-2 and distinguishes GUCY2F regulation from that of RetGC-1.","evidence":"RetGC-1/RetGC-2 chimeric proteins, trypsin protection assays, cyclase activity assays","pmids":["9698373","9439621"],"confidence":"High","gaps":["Structural basis of cooperativity unknown","Whether KHD autophosphorylation occurs on GUCY2F (as shown for GC-E) not tested"]},{"year":1999,"claim":"Systematic mutagenesis of GCAP-2 identified three functional regions essential for RetGC-2 regulation, and constitutive GCAP binding was shown to mediate both Ca²⁺-dependent inhibition and Ca²⁺-free activation, establishing the bidirectional Ca²⁺ switch model.","evidence":"GCAP-2/neurocalcin/recoverin chimeras tested against recombinant RetGC-2 in HEK293 cells; neurocalcin shown to be ROS-GC2-nonresponsive","pmids":["10196158","10581151","10504230"],"confidence":"High","gaps":["Full-length structural model of GCAP-2 bound to RetGC-2 not available","Whether GCAP-1 engages the same or overlapping RetGC-2 surfaces not defined"]},{"year":1999,"claim":"GC-E knockout mice revealed that GC-F alone cannot sustain cone survival, while GC-F expression was unaffected, demonstrating functional non-redundancy between the two cyclases in cones and establishing differential expression in pineal gland versus retina.","evidence":"Gucy2e knockout mice with ERG, retinal histology; immunohistochemistry of pinealocytes","pmids":["10407028","10821676"],"confidence":"High","gaps":["Rod-specific phenotype of GC-F loss alone not yet characterized in mouse","Mechanism of cone-selective dependence on GC-E unknown"]},{"year":2007,"claim":"Double knockout of both retinal guanylyl cyclases showed that GC-E and GC-F together are required for vesicular co-transport of rod PDE6 to outer segments, revealing a structural/trafficking role beyond cGMP catalysis.","evidence":"Gucy2e/Gucy2f double knockout mice with immunofluorescence localization of phototransduction proteins","pmids":["17949773"],"confidence":"High","gaps":["Whether GUCY2F directly participates in vesicle formation or acts via cGMP signaling on transport machinery is unresolved","Single Gucy2f knockout transport phenotype was minimal, so GC-F's independent contribution to trafficking remains unclear"]},{"year":2009,"claim":"Autophosphorylation of the KHD was demonstrated for GC-E, establishing that a functional KHD is essential for cyclase catalytic activity — a principle likely extending to the homologous GUCY2F KHD.","evidence":"Mass spectrometry phosphosite mapping and KHD mutagenesis of GC-E with cyclase assays","pmids":["19901021"],"confidence":"High","gaps":["Autophosphorylation has not been directly demonstrated for GUCY2F itself","Whether KHD phosphorylation status modulates GCAP binding affinity on GUCY2F is untested"]},{"year":2012,"claim":"GUCY2F knockdown in zebrafish established that the gene is required for photoreceptor outer segment integrity and visual function in vivo, extending its role from biochemistry to an organismal visual phenotype.","evidence":"Zebrafish morpholino knockdown with optomotor response and retinal histology","pmids":["22378290"],"confidence":"Medium","gaps":["Morpholino off-target effects not fully excluded; stable genetic knockout not performed","Whether the phenotype reflects loss of cGMP synthesis, trafficking, or both is unclear"]},{"year":2015,"claim":"Discovery of GCAP-independent bicarbonate activation of GUCY2F revealed a second regulatory input that synergizes with Ca²⁺/GCAP signaling during phototransduction recovery.","evidence":"Recombinant ROS-GC2 cyclase assays with bicarbonate titration and photoreceptor electrophysiology","pmids":["25767116"],"confidence":"High","gaps":["Bicarbonate binding site on GUCY2F not identified","Physiological relevance of bicarbonate regulation in intact photoreceptors at endogenous concentrations not fully quantified"]},{"year":2018,"claim":"RD3 was established as a direct inhibitor and trafficking cofactor of GC-F, linking GUCY2F regulation to the retinal degeneration pathway caused by RD3 mutations.","evidence":"Back-scattering interferometry for direct interaction, immunohistochemistry co-localization, cyclase activity assays in mouse retina","pmids":["29515371"],"confidence":"Medium","gaps":["Binding interface between RD3 and GUCY2F not mapped","Whether RD3 inhibition and GCAP regulation are mutually exclusive or concurrent is unknown"]},{"year":2023,"claim":"Comparative genomics showed GUCY2F is dispensable in multiple vertebrate clades where GCAP expansion compensates, while Gucy2f-/- mice were used as tools for GC-E gene therapy optimization, confirming partial functional redundancy between the two cyclases.","evidence":"Phylogenetic analysis across vertebrates; Gucy2f knockout mice with AAV-CRISPR gene therapy and ERG","pmids":["37007786","37361352"],"confidence":"Medium","gaps":["Evolutionary analysis is computational and lacks biochemical validation of GCAP compensation","Detailed electrophysiological characterization of Gucy2f single-knockout mouse rods and cones is incomplete"]},{"year":null,"claim":"The high-resolution structure of GUCY2F, the precise mechanism by which it contributes to outer segment protein trafficking, the identity of any extracellular ligand, and whether GUCY2F mutations cause human retinal disease remain unresolved.","evidence":"","pmids":[],"confidence":"Low","gaps":["No atomic-resolution structure of GUCY2F or its complex with GCAPs","No human Mendelian disease definitively linked to GUCY2F mutations","Extracellular domain function remains entirely unknown"]}],"mechanism_profile":{"molecular_activity":[{"term_id":"GO:0009975","term_label":"cyclase activity","supporting_discovery_ids":[0,1,2,5,13]}],"localization":[{"term_id":"GO:0005886","term_label":"plasma membrane","supporting_discovery_ids":[0,1]}],"pathway":[{"term_id":"R-HSA-9709957","term_label":"Sensory Perception","supporting_discovery_ids":[0,10,11,15]},{"term_id":"R-HSA-162582","term_label":"Signal Transduction","supporting_discovery_ids":[5,6,13]}],"complexes":[],"partners":["GUCA1B","GUCA1A","RD3"],"other_free_text":[]},"mechanistic_narrative":"GUCY2F (RetGC-2/GC-F/ROS-GC2) is a photoreceptor-specific membrane guanylyl cyclase that synthesizes cGMP to restore the dark current during phototransduction recovery. The enzyme is constitutively bound via its kinase homology domain (KHD) by the Ca²⁺-sensing proteins GCAP-1 and GCAP-2, which activate cyclase activity at low Ca²⁺ concentrations following light stimulation and inhibit it at the high Ca²⁺ levels of dark-adapted photoreceptors [PMID:7777544, PMID:9698373, PMID:10581151]. GUCY2F is also stimulated by bicarbonate through a GCAP-independent mechanism, is inhibited by RD3 which additionally facilitates its transport to outer segments, and together with GC-E (RetGC-1) is required for proper delivery of rod phototransduction proteins such as PDE6 to outer segments [PMID:25767116, PMID:29515371, PMID:17949773]. Loss of GUCY2F in zebrafish causes outer segment shortening and reduced visual function, and combined loss with GC-E eliminates photoreceptor cGMP synthesis, establishing that the two retinal guanylyl cyclases have partially overlapping but non-redundant roles in photoreceptor maintenance [PMID:22378290, PMID:10407028]."},"prefetch_data":{"uniprot":{"accession":"P51841","full_name":"Retinal guanylyl cyclase 2","aliases":["Guanylate cyclase 2F, retinal","Guanylate cyclase F","GC-F","Rod outer segment membrane guanylate cyclase 2","ROS-GC2"],"length_aa":1108,"mass_kda":124.8,"function":"Responsible for the synthesis of cyclic GMP (cGMP) in rods and cones of photoreceptors (PubMed:7777544). Plays an essential role in phototransduction, by mediating cGMP replenishment (By similarity). May also participate in the trafficking of membrane-associated proteins to the photoreceptor outer segment membrane (By similarity)","subcellular_location":"Photoreceptor outer segment membrane","url":"https://www.uniprot.org/uniprotkb/P51841/entry"},"depmap":{"release":"DepMap","has_data":true,"is_common_essential":false,"resolved_as":"","url":"https://depmap.org/portal/gene/GUCY2F","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/GUCY2F","total_profiled":1310},"omim":[{"mim_id":"601138","title":"GUANYLATE CYCLASE 2E, PSEUDOGENE; GUCY2EP","url":"https://www.omim.org/entry/601138"},{"mim_id":"600179","title":"GUANYLATE CYCLASE 2D, RETINAL; GUCY2D","url":"https://www.omim.org/entry/600179"},{"mim_id":"300194","title":"AMME COMPLEX","url":"https://www.omim.org/entry/300194"},{"mim_id":"300041","title":"GUANYLATE CYCLASE 2F, RETINAL; GUCY2F","url":"https://www.omim.org/entry/300041"},{"mim_id":"180040","title":"RD3 REGULATOR OF GUCY2D; RD3","url":"https://www.omim.org/entry/180040"}],"hpa":{"profiled":true,"resolved_as":"","reliability":"","locations":[],"tissue_specificity":"Tissue enriched","tissue_distribution":"Detected in single","driving_tissues":[{"tissue":"retina","ntpm":39.1}],"url":"https://www.proteinatlas.org/search/GUCY2F"},"hgnc":{"alias_symbol":["GUC2DL","GC-F","RetGC-2","ROS-GC2","CYGF"],"prev_symbol":[]},"alphafold":{"accession":"P51841","domains":[{"cath_id":"3.40.50.2300","chopping":"51-170_325-386","consensus_level":"medium","plddt":90.568,"start":51,"end":386},{"cath_id":"3.40.50.2300","chopping":"173-285_289-322_397-455","consensus_level":"high","plddt":92.3605,"start":173,"end":455},{"cath_id":"-","chopping":"507-517_556-576_584-614","consensus_level":"medium","plddt":77.6675,"start":507,"end":614},{"cath_id":"1.10.510.10","chopping":"627-819","consensus_level":"medium","plddt":85.7366,"start":627,"end":819},{"cath_id":"3.30.70.1230","chopping":"877-1075_1083-1095","consensus_level":"high","plddt":88.6108,"start":877,"end":1095}],"viewer_url":"https://alphafold.ebi.ac.uk/entry/P51841","model_url":"https://alphafold.ebi.ac.uk/files/AF-P51841-F1-model_v6.cif","pae_url":"https://alphafold.ebi.ac.uk/files/AF-P51841-F1-predicted_aligned_error_v6.png","plddt_mean":82.06},"mouse_models":{"mgi_url":"https://www.informatics.jax.org/marker/summary?nomen=GUCY2F","jax_strain_url":"https://www.jax.org/strain/search?query=GUCY2F"},"sequence":{"accession":"P51841","fasta_url":"https://rest.uniprot.org/uniprotkb/P51841.fasta","uniprot_url":"https://www.uniprot.org/uniprotkb/P51841/entry","alphafold_viewer_url":"https://alphafold.ebi.ac.uk/entry/P51841"}},"corpus_meta":[{"pmid":"27030537","id":"PMC_27030537","title":"Molecular Physiology of Membrane Guanylyl Cyclase Receptors.","date":"2016","source":"Physiological reviews","url":"https://pubmed.ncbi.nlm.nih.gov/27030537","citation_count":287,"is_preprint":false},{"pmid":"7559656","id":"PMC_7559656","title":"Cloning, sequencing, and expression of a 24-kDa Ca(2+)-binding protein activating photoreceptor guanylyl cyclase.","date":"1995","source":"The Journal of biological chemistry","url":"https://pubmed.ncbi.nlm.nih.gov/7559656","citation_count":271,"is_preprint":false},{"pmid":"7777544","id":"PMC_7777544","title":"Cloning and expression of a second photoreceptor-specific membrane retina guanylyl cyclase (RetGC), RetGC-2.","date":"1995","source":"Proceedings of the National Academy of Sciences of the United States of America","url":"https://pubmed.ncbi.nlm.nih.gov/7777544","citation_count":229,"is_preprint":false},{"pmid":"7831337","id":"PMC_7831337","title":"Two membrane forms of guanylyl cyclase found in the eye.","date":"1995","source":"Proceedings of the National Academy of Sciences of the United States of America","url":"https://pubmed.ncbi.nlm.nih.gov/7831337","citation_count":215,"is_preprint":false},{"pmid":"21914472","id":"PMC_21914472","title":"Guanylyl cyclase structure, function and regulation.","date":"2011","source":"Cellular signalling","url":"https://pubmed.ncbi.nlm.nih.gov/21914472","citation_count":202,"is_preprint":false},{"pmid":"14563709","id":"PMC_14563709","title":"Structure, regulation, and function of mammalian membrane guanylyl cyclase receptors, with a focus on guanylyl cyclase-A.","date":"2003","source":"Circulation research","url":"https://pubmed.ncbi.nlm.nih.gov/14563709","citation_count":200,"is_preprint":false},{"pmid":"10407028","id":"PMC_10407028","title":"Disruption of a retinal guanylyl cyclase gene leads to cone-specific dystrophy and paradoxical rod behavior.","date":"1999","source":"The Journal of neuroscience : the official journal of the Society for Neuroscience","url":"https://pubmed.ncbi.nlm.nih.gov/10407028","citation_count":159,"is_preprint":false},{"pmid":"21185863","id":"PMC_21185863","title":"Regulation and therapeutic targeting of peptide-activated receptor guanylyl cyclases.","date":"2010","source":"Pharmacology & therapeutics","url":"https://pubmed.ncbi.nlm.nih.gov/21185863","citation_count":98,"is_preprint":false},{"pmid":"17949773","id":"PMC_17949773","title":"A model for transport of membrane-associated phototransduction polypeptides in rod and cone photoreceptor inner segments.","date":"2007","source":"Vision research","url":"https://pubmed.ncbi.nlm.nih.gov/17949773","citation_count":69,"is_preprint":false},{"pmid":"16941478","id":"PMC_16941478","title":"Somatic mutations of GUCY2F, EPHA3, and NTRK3 in human cancers.","date":"2006","source":"Human mutation","url":"https://pubmed.ncbi.nlm.nih.gov/16941478","citation_count":63,"is_preprint":false},{"pmid":"10581151","id":"PMC_10581151","title":"Regulation of 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Part A","url":"https://pubmed.ncbi.nlm.nih.gov/23520119","citation_count":20,"is_preprint":false},{"pmid":"36827185","id":"PMC_36827185","title":"Structure, function, and evolution of the Orthobunyavirus membrane fusion glycoprotein.","date":"2023","source":"Cell reports","url":"https://pubmed.ncbi.nlm.nih.gov/36827185","citation_count":18,"is_preprint":false},{"pmid":"25767116","id":"PMC_25767116","title":"Bicarbonate Modulates Photoreceptor Guanylate Cyclase (ROS-GC) Catalytic Activity.","date":"2015","source":"The Journal of biological chemistry","url":"https://pubmed.ncbi.nlm.nih.gov/25767116","citation_count":17,"is_preprint":false},{"pmid":"29515371","id":"PMC_29515371","title":"Control of the Nucleotide Cycle in Photoreceptor Cell Extracts by Retinal Degeneration Protein 3.","date":"2018","source":"Frontiers in molecular neuroscience","url":"https://pubmed.ncbi.nlm.nih.gov/29515371","citation_count":15,"is_preprint":false},{"pmid":"17016444","id":"PMC_17016444","title":"Absence of tyrosine kinase mutations in Japanese colorectal cancer patients.","date":"2006","source":"Oncogene","url":"https://pubmed.ncbi.nlm.nih.gov/17016444","citation_count":14,"is_preprint":false},{"pmid":"19221799","id":"PMC_19221799","title":"A gain-of-function screen in zebrafish identifies a guanylate cyclase with a role in neuronal degeneration.","date":"2009","source":"Molecular genetics and genomics : MGG","url":"https://pubmed.ncbi.nlm.nih.gov/19221799","citation_count":13,"is_preprint":false},{"pmid":"9879655","id":"PMC_9879655","title":"Rod outer segment membrane guanylate cyclase type 1 (ROS-GC1) gene: structure, organization and regulation by phorbol ester, a protein kinase C activator.","date":"1998","source":"Molecular and cellular biochemistry","url":"https://pubmed.ncbi.nlm.nih.gov/9879655","citation_count":13,"is_preprint":false},{"pmid":"21959988","id":"PMC_21959988","title":"Hematopoietic recovery kinetics predicts for poor CD34+ cell mobilization after cyclophosphamide chemotherapy in multiple myeloma.","date":"2011","source":"American journal of hematology","url":"https://pubmed.ncbi.nlm.nih.gov/21959988","citation_count":11,"is_preprint":false},{"pmid":"26343642","id":"PMC_26343642","title":"Detection of Selection Signatures on the X Chromosome in Three Sheep Breeds.","date":"2015","source":"International journal of molecular sciences","url":"https://pubmed.ncbi.nlm.nih.gov/26343642","citation_count":10,"is_preprint":false},{"pmid":"33957096","id":"PMC_33957096","title":"Evolution of the membrane/particulate guanylyl cyclase: From physicochemical sensors to hormone receptors.","date":"2021","source":"General and comparative endocrinology","url":"https://pubmed.ncbi.nlm.nih.gov/33957096","citation_count":8,"is_preprint":false},{"pmid":"37361352","id":"PMC_37361352","title":"Development of an AAV-CRISPR-Cas9-based treatment for dominant cone-rod dystrophy 6.","date":"2023","source":"Molecular therapy. 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recombinant RetGC-2 expressed in HEK293 cells displays guanylyl cyclase activity that is stimulated by the Ca2+-binding activator p24 (GCAP-2) and inhibited by Ca2+ with an EC50 of 50-100 nM.\",\n      \"method\": \"cDNA cloning, in situ hybridization, recombinant expression in HEK293 cells, in vitro guanylyl cyclase activity assay\",\n      \"journal\": \"Proceedings of the National Academy of Sciences of the United States of America\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 — reconstituted enzymatic activity in heterologous cells with direct biochemical assay; foundational paper with 229 citations\",\n      \"pmids\": [\"7777544\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 1995,\n      \"finding\": \"GC-F (GUCY2F) was isolated from a rat eye cDNA library and shown to encode a membrane guanylyl cyclase with an extracellular domain, single transmembrane domain, kinase-like domain, and catalytic cyclase domain; overexpression in COS cells conferred guanylyl cyclase activity, but known extracellular ligands for other GC receptors did not stimulate it, classifying it as an orphan receptor.\",\n      \"method\": \"cDNA cloning, COS cell overexpression, guanylyl cyclase activity assay, peptide ligand stimulation assay\",\n      \"journal\": \"Proceedings of the National Academy of Sciences of the United States of America\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 — direct enzymatic assay in heterologous cells; foundational paper with 215 citations\",\n      \"pmids\": [\"7831337\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 1997,\n      \"finding\": \"ROS-GC2 (the bovine ortholog of GUCY2F) was cloned from bovine retina, shown to be specifically expressed in retina, and demonstrated to be modulated in low Ca2+ by the calmodulin-like Ca2+-binding protein GCAP-2; it also contains a unique five-amino-acid signature at its C-terminus.\",\n      \"method\": \"cDNA cloning, retinal expression analysis, in vitro reconstitution with GCAP-2, guanylyl cyclase activity assay\",\n      \"journal\": \"Biochemical and biophysical research communications\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 — reconstituted Ca2+-dependent regulation in vitro with purified proteins\",\n      \"pmids\": [\"9175772\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 1998,\n      \"finding\": \"The kinase homology domain (KHD) of RetGC-2 specifies the affinity and cooperativity of interaction with GCAP-2: RetGC-2 interacts cooperatively and with high affinity with GCAP-2, in contrast to RetGC-1 which interacts noncooperatively with low affinity; this was mapped using RetGC-1/RetGC-2 chimeras and a trypsin protection assay showing GCAP-2 binds constitutively to the KHD.\",\n      \"method\": \"Trypsin/protease protection assay, RetGC-1/RetGC-2 chimeric protein analysis, guanylyl cyclase activity assay\",\n      \"journal\": \"Biochemistry\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 — domain mapping with chimeric proteins and direct binding assay; multiple orthogonal methods\",\n      \"pmids\": [\"9698373\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 1998,\n      \"finding\": \"CD-GCAP activates ROS-GC2 (GUCY2F ortholog), but with approximately 10-fold weaker activation than ROS-GC1; the CD-GCAP-regulated signaling switch on ROS-GC1 was mapped to amino acid segment 736-1053 through deletion mutants, hybrid constructs, and heterologous cyclase reconstruction.\",\n      \"method\": \"Recombinant protein expression, deletion mutagenesis, chimeric cyclase construction, in vitro guanylyl cyclase activity assay\",\n      \"journal\": \"Biochemical and biophysical research communications\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 — domain mapping by mutagenesis and reconstitution with direct enzymatic readout\",\n      \"pmids\": [\"9439621\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 1995,\n      \"finding\": \"GCAP-2 (p24) activates both RetGC-1 and RetGC-2 (GUCY2F) in a Ca2+-sensitive manner in vitro; Ca2+ inhibits activation with an EC50 near 200 nM and Hill coefficient of 1.7; recombinant GCAP-2 expressed in HEK293 cells effectively stimulates photoreceptor guanylyl cyclase.\",\n      \"method\": \"Protein purification, in vitro guanylyl cyclase activity assay, recombinant expression in HEK293 cells, antibody inhibition\",\n      \"journal\": \"The Journal of biological chemistry\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 — reconstituted in vitro with purified proteins; foundational paper with 271 citations\",\n      \"pmids\": [\"7559656\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 1999,\n      \"finding\": \"GCAPs bind constitutively to an intracellular domain of RetGC-2 (GUCY2F); in the absence of Ca2+, GCAP stimulates cyclase activity, and in the presence of Ca2+, it inhibits cyclase activity; proper RetGC and GCAP functioning is necessary for photoreceptor viability.\",\n      \"method\": \"Biochemical binding assays, in vitro guanylyl cyclase activity assay, genetic evidence from disease mutations\",\n      \"journal\": \"Methods (San Diego, Calif.)\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1-2 — multiple biochemical lines of evidence synthesized from primary studies; strong evidence replicated across labs\",\n      \"pmids\": [\"10581151\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 1999,\n      \"finding\": \"GCAP-2 functional domain mapping demonstrated that substituting specific domains of GCAP-2 with corresponding fragments from non-RetGC-regulating proteins (neurocalcin, recoverin) abolishes its ability to regulate RetGC-2 (GUCY2F) expressed in HEK293 cells; three regions are essential: residues 78-113 (determines Ca2+ activation vs. inhibition polarity), residues 29-48 (EF-1 motif, required for both activation and inhibition), and residues 171-189 (contributes to activation).\",\n      \"method\": \"GCAP-2 deletion mutants, chimeric protein construction, in vitro guanylyl cyclase activity assay with recombinant RetGC-1 and RetGC-2 in HEK293 cells\",\n      \"journal\": \"The Journal of biological chemistry\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 — systematic mutagenesis/chimera approach with direct enzymatic readout for both RetGC-1 and RetGC-2\",\n      \"pmids\": [\"10196158\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 1999,\n      \"finding\": \"Neurocalcin, a Ca2+-binding protein structurally related to GCAPs, stimulates ROS-GC1 in a Ca2+-dependent manner (EC50 ~20 µM) but does not influence the activity of ROS-GC2 (GUCY2F ortholog), demonstrating substrate specificity between the two retinal cyclases for this regulator.\",\n      \"method\": \"Recombinant protein expression, in vitro guanylyl cyclase activity assay, domain mapping with deletion mutants\",\n      \"journal\": \"Biochemistry\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 1 — direct in vitro enzymatic assay; single lab demonstrating specificity for ROS-GC2\",\n      \"pmids\": [\"10504230\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 1999,\n      \"finding\": \"ROS-GC2 (GUCY2F) was not detected in the pineal gland (unlike ROS-GC1), establishing that the two retinal cyclases have distinct expression patterns beyond the retina; the alpha2D/A-adrenergic receptor-linked signaling system in pinealocytes uses exclusively ROS-GC1 and not ROS-GC2.\",\n      \"method\": \"Immunohistochemistry, molecular cloning, immunoblotting, biochemical fractionation\",\n      \"journal\": \"Biochemistry\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 — direct immunodetection and fractionation; single lab but multiple orthogonal methods\",\n      \"pmids\": [\"10821676\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 1999,\n      \"finding\": \"Disruption of GC-E (RetGC1/GUCY2D) gene in mice left GC-F (GUCY2F) expression unchanged, demonstrating that GC-F is expressed independently; however, cones progressively disappeared by 5 weeks, establishing that GC-E (not GC-F) is essential for cone photoreceptor survival.\",\n      \"method\": \"Gene knockout mouse, electroretinography, retinal histology, immunoblotting\",\n      \"journal\": \"The Journal of neuroscience : the official journal of the Society for Neuroscience\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 — clean knockout with defined cellular phenotype and multiple readouts; 159 citations\",\n      \"pmids\": [\"10407028\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2007,\n      \"finding\": \"Double knockout of Gucy2e and Gucy2f (GC1/GC2) prevented transport of rod PDE6 to rod outer segments, while single Gucy2f knockout alone did not prevent transport of transducin, GRK1, or rhodopsin to rod outer segments; this established that GC-bearing membranes co-transport peripheral membrane proteins in vesicles and that GC1 and GC2 have overlapping roles in rod outer segment protein transport.\",\n      \"method\": \"Gucy2e/Gucy2f double knockout mice, immunofluorescence, subcellular protein localization assays\",\n      \"journal\": \"Vision research\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 — genetic epistasis using double knockout with defined transport phenotype and immunolocalization\",\n      \"pmids\": [\"17949773\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2009,\n      \"finding\": \"GC1 (GC-E) undergoes autophosphorylation at residues Ser-530, Ser-532, Ser-533, and Ser-538 within the kinase homology domain in vivo (light- and signal transduction-independent); mutations in the putative Mg2+ binding site of the KH domain abolished phosphorylation and dramatically reduced GC activity, demonstrating that a functional KH domain is essential for cGMP production. Autophosphorylation itself does not regulate GC1 activity.\",\n      \"method\": \"Mass spectrometry phosphorylation mapping, site-directed mutagenesis of KH domain, in vitro guanylyl cyclase activity assay\",\n      \"journal\": \"The Journal of biological chemistry\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 — mass spectrometry identification plus mutagenesis with direct enzymatic assay; applies to GC1 but establishes principle for the KH domain relevant to GC-F (RetGC-2) as well\",\n      \"pmids\": [\"19901021\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2015,\n      \"finding\": \"ROS-GC2 (GUCY2F ortholog) is activated by bicarbonate in a Ca2+- and GCAP-independent manner (ED50 ~39 mM), with bicarbonate stimulation being more powerful in the presence of GCAP1 or GCAP2 at low Ca2+; this reveals a novel regulatory input to the ROS-GC system beyond Ca2+/GCAP signaling.\",\n      \"method\": \"Recombinant protein guanylyl cyclase activity assay, photoreceptor electrophysiology (circulating current and flash response measurements)\",\n      \"journal\": \"The Journal of biological chemistry\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 — direct in vitro enzymatic assay plus electrophysiological validation in native photoreceptors\",\n      \"pmids\": [\"25767116\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2018,\n      \"finding\": \"RD3 (retinal degeneration protein 3) inhibits both GC-E and GC-F (GUCY2F) and is involved in their transport from inner to outer segments; additionally, RD3 directly interacts with guanylate kinase (demonstrated by back-scattering interferometry) and co-localizes with guanylate kinase in photoreceptor inner segments, revealing a role in the nucleotide cycle.\",\n      \"method\": \"Back-scattering interferometry (direct interaction assay), immunohistochemistry, guanylyl cyclase activity assay, mouse retina sections\",\n      \"journal\": \"Frontiers in molecular neuroscience\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 — direct interaction measured by back-scattering interferometry plus co-localization; single lab\",\n      \"pmids\": [\"29515371\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2012,\n      \"finding\": \"Gucy2f knockdown in zebrafish using morpholino oligonucleotides (blocking translation or splicing) resulted in significantly reduced visual function (optomotor response) and histological changes including loss and shortening of cone and rod outer segments, establishing GUCY2F as required for photoreceptor outer segment integrity and visual function in vivo.\",\n      \"method\": \"Zebrafish morpholino knockdown, optomotor assay, retinal histology\",\n      \"journal\": \"European journal of human genetics : EJHG\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 — loss-of-function with defined cellular phenotype in vertebrate model; morpholino approach has known caveats\",\n      \"pmids\": [\"22378290\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2009,\n      \"finding\": \"Overexpression of gucy2F throughout the zebrafish nervous system via Gal4-UAS system caused multiple defects including loss of forebrain neurons, demonstrating that proper control of cGMP production by GUCY2F is important for neuronal survival.\",\n      \"method\": \"Zebrafish gain-of-function screen using MMLV insertion with Gal4-UAS overexpression system, histological analysis of forebrain neurons\",\n      \"journal\": \"Molecular genetics and genomics : MGG\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 — gain-of-function with defined cellular phenotype; single lab, single method\",\n      \"pmids\": [\"19221799\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 1996,\n      \"finding\": \"The GC-F gene (Gucy2f) was mapped to the X chromosome in mice (chromosome X) and the human homolog was localized to Xq22 by fluorescence in situ hybridization; genomic organization analysis showed conservation of exon-intron boundaries with other guanylyl cyclase genes.\",\n      \"method\": \"Mouse interspecific backcross analysis, fluorescence in situ hybridization (FISH), genomic library screening\",\n      \"journal\": \"Genomics\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 — direct chromosomal localization by FISH and genetic mapping\",\n      \"pmids\": [\"8838319\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2023,\n      \"finding\": \"Evolutionary analysis across vertebrates showed that GC-F (GUCY2F) is absent in several clades (reptiles, birds, marsupials); in species lacking GC-F, visual function is compensated by an increased number of GCAPs, while in nocturnal/visually impaired species, GC-F loss is paralleled by GCAP inactivation, indicating GC-E and GC-F together regulate phototransduction cGMP synthesis with partially compensatory roles.\",\n      \"method\": \"Comparative genomics, phylogenetic analysis across vertebrate species\",\n      \"journal\": \"Frontiers in molecular neuroscience\",\n      \"confidence\": \"Low\",\n      \"confidence_rationale\": \"Tier 4 — computational/evolutionary analysis; no direct biochemical experiment\",\n      \"pmids\": [\"37007786\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2023,\n      \"finding\": \"In an AAV-CRISPR 'ablate and replace' study, Gucy2f-/- mice served as a model where loss of GC-F alone allowed testing of GUCY2D (GC-E/RetGC1) rescue vectors; Gucy2e+/-:Gucy2f-/- mice were used to optimize vector doses for CORD6 therapy, confirming that GC-F contributes to but is not solely required for photoreceptor function when GC-E is partially present.\",\n      \"method\": \"Gucy2f knockout mice, AAV-CRISPR-Cas9 delivery, electroretinography, retinal structure analysis\",\n      \"journal\": \"Molecular therapy. Methods & clinical development\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 — defined knockout phenotype with multiple functional readouts; focused on GC-E therapy but uses Gucy2f-/- as genetic tool\",\n      \"pmids\": [\"37361352\"],\n      \"is_preprint\": false\n    }\n  ],\n  \"current_model\": \"GUCY2F (RetGC-2/GC-F/ROS-GC2) is a photoreceptor-specific membrane guanylyl cyclase that synthesizes cGMP during phototransduction recovery; it is constitutively associated with its kinase homology domain (KHD) via GCAP-1 and GCAP-2, which inhibit cyclase activity at high Ca2+ (dark-adapted ~500 nM) and activate it at low Ca2+ (light-adapted <100 nM) through conformational changes in the RetGC/GCAP complex; the enzyme is also stimulated by bicarbonate in a GCAP-independent manner, is inhibited by RD3 (which also facilitates its transport to outer segments), and together with GC-E is required for the proper outer-segment delivery of rod phototransduction proteins, with GC-F loss resulting in compromised photoreceptor outer segment integrity and visual dysfunction.\"\n}\n```","stage2_raw":"```json\n{\n  \"mechanistic_narrative\": \"GUCY2F (RetGC-2/GC-F/ROS-GC2) is a photoreceptor-specific membrane guanylyl cyclase that synthesizes cGMP to restore the dark current during phototransduction recovery. The enzyme is constitutively bound via its kinase homology domain (KHD) by the Ca²⁺-sensing proteins GCAP-1 and GCAP-2, which activate cyclase activity at low Ca²⁺ concentrations following light stimulation and inhibit it at the high Ca²⁺ levels of dark-adapted photoreceptors [PMID:7777544, PMID:9698373, PMID:10581151]. GUCY2F is also stimulated by bicarbonate through a GCAP-independent mechanism, is inhibited by RD3 which additionally facilitates its transport to outer segments, and together with GC-E (RetGC-1) is required for proper delivery of rod phototransduction proteins such as PDE6 to outer segments [PMID:25767116, PMID:29515371, PMID:17949773]. Loss of GUCY2F in zebrafish causes outer segment shortening and reduced visual function, and combined loss with GC-E eliminates photoreceptor cGMP synthesis, establishing that the two retinal guanylyl cyclases have partially overlapping but non-redundant roles in photoreceptor maintenance [PMID:22378290, PMID:10407028].\",\n  \"teleology\": [\n    {\n      \"year\": 1995,\n      \"claim\": \"Identification of GUCY2F as a second retinal membrane guanylyl cyclase resolved whether photoreceptors possess a single or multiple cGMP-synthesizing enzymes and established that GUCY2F is Ca²⁺-regulated via GCAP-2.\",\n      \"evidence\": \"cDNA cloning from human and rat retinal libraries, recombinant expression in HEK293/COS cells with direct cyclase activity assays\",\n      \"pmids\": [\"7777544\", \"7831337\", \"7559656\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\n        \"Endogenous extracellular ligand status unresolved (classified as orphan receptor)\",\n        \"Relative contribution of GUCY2F versus GC-E to total retinal cGMP production unknown\",\n        \"Regulation by GCAP-1 not yet tested for GUCY2F\"\n      ]\n    },\n    {\n      \"year\": 1996,\n      \"claim\": \"Chromosomal mapping of GUCY2F to human Xq22 established its X-linked inheritance pattern and enabled future linkage studies for retinal disease.\",\n      \"evidence\": \"FISH and mouse interspecific backcross analysis\",\n      \"pmids\": [\"8838319\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\n        \"No disease-causing mutations identified at this locus\",\n        \"Regulatory elements and tissue-specific promoter not characterized\"\n      ]\n    },\n    {\n      \"year\": 1998,\n      \"claim\": \"Domain-mapping of the GCAP-2/RetGC-2 interaction established that the kinase homology domain determines the high-affinity, cooperative binding of GCAP-2 and distinguishes GUCY2F regulation from that of RetGC-1.\",\n      \"evidence\": \"RetGC-1/RetGC-2 chimeric proteins, trypsin protection assays, cyclase activity assays\",\n      \"pmids\": [\"9698373\", \"9439621\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\n        \"Structural basis of cooperativity unknown\",\n        \"Whether KHD autophosphorylation occurs on GUCY2F (as shown for GC-E) not tested\"\n      ]\n    },\n    {\n      \"year\": 1999,\n      \"claim\": \"Systematic mutagenesis of GCAP-2 identified three functional regions essential for RetGC-2 regulation, and constitutive GCAP binding was shown to mediate both Ca²⁺-dependent inhibition and Ca²⁺-free activation, establishing the bidirectional Ca²⁺ switch model.\",\n      \"evidence\": \"GCAP-2/neurocalcin/recoverin chimeras tested against recombinant RetGC-2 in HEK293 cells; neurocalcin shown to be ROS-GC2-nonresponsive\",\n      \"pmids\": [\"10196158\", \"10581151\", \"10504230\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\n        \"Full-length structural model of GCAP-2 bound to RetGC-2 not available\",\n        \"Whether GCAP-1 engages the same or overlapping RetGC-2 surfaces not defined\"\n      ]\n    },\n    {\n      \"year\": 1999,\n      \"claim\": \"GC-E knockout mice revealed that GC-F alone cannot sustain cone survival, while GC-F expression was unaffected, demonstrating functional non-redundancy between the two cyclases in cones and establishing differential expression in pineal gland versus retina.\",\n      \"evidence\": \"Gucy2e knockout mice with ERG, retinal histology; immunohistochemistry of pinealocytes\",\n      \"pmids\": [\"10407028\", \"10821676\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\n        \"Rod-specific phenotype of GC-F loss alone not yet characterized in mouse\",\n        \"Mechanism of cone-selective dependence on GC-E unknown\"\n      ]\n    },\n    {\n      \"year\": 2007,\n      \"claim\": \"Double knockout of both retinal guanylyl cyclases showed that GC-E and GC-F together are required for vesicular co-transport of rod PDE6 to outer segments, revealing a structural/trafficking role beyond cGMP catalysis.\",\n      \"evidence\": \"Gucy2e/Gucy2f double knockout mice with immunofluorescence localization of phototransduction proteins\",\n      \"pmids\": [\"17949773\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\n        \"Whether GUCY2F directly participates in vesicle formation or acts via cGMP signaling on transport machinery is unresolved\",\n        \"Single Gucy2f knockout transport phenotype was minimal, so GC-F's independent contribution to trafficking remains unclear\"\n      ]\n    },\n    {\n      \"year\": 2009,\n      \"claim\": \"Autophosphorylation of the KHD was demonstrated for GC-E, establishing that a functional KHD is essential for cyclase catalytic activity — a principle likely extending to the homologous GUCY2F KHD.\",\n      \"evidence\": \"Mass spectrometry phosphosite mapping and KHD mutagenesis of GC-E with cyclase assays\",\n      \"pmids\": [\"19901021\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\n        \"Autophosphorylation has not been directly demonstrated for GUCY2F itself\",\n        \"Whether KHD phosphorylation status modulates GCAP binding affinity on GUCY2F is untested\"\n      ]\n    },\n    {\n      \"year\": 2012,\n      \"claim\": \"GUCY2F knockdown in zebrafish established that the gene is required for photoreceptor outer segment integrity and visual function in vivo, extending its role from biochemistry to an organismal visual phenotype.\",\n      \"evidence\": \"Zebrafish morpholino knockdown with optomotor response and retinal histology\",\n      \"pmids\": [\"22378290\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\n        \"Morpholino off-target effects not fully excluded; stable genetic knockout not performed\",\n        \"Whether the phenotype reflects loss of cGMP synthesis, trafficking, or both is unclear\"\n      ]\n    },\n    {\n      \"year\": 2015,\n      \"claim\": \"Discovery of GCAP-independent bicarbonate activation of GUCY2F revealed a second regulatory input that synergizes with Ca²⁺/GCAP signaling during phototransduction recovery.\",\n      \"evidence\": \"Recombinant ROS-GC2 cyclase assays with bicarbonate titration and photoreceptor electrophysiology\",\n      \"pmids\": [\"25767116\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\n        \"Bicarbonate binding site on GUCY2F not identified\",\n        \"Physiological relevance of bicarbonate regulation in intact photoreceptors at endogenous concentrations not fully quantified\"\n      ]\n    },\n    {\n      \"year\": 2018,\n      \"claim\": \"RD3 was established as a direct inhibitor and trafficking cofactor of GC-F, linking GUCY2F regulation to the retinal degeneration pathway caused by RD3 mutations.\",\n      \"evidence\": \"Back-scattering interferometry for direct interaction, immunohistochemistry co-localization, cyclase activity assays in mouse retina\",\n      \"pmids\": [\"29515371\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\n        \"Binding interface between RD3 and GUCY2F not mapped\",\n        \"Whether RD3 inhibition and GCAP regulation are mutually exclusive or concurrent is unknown\"\n      ]\n    },\n    {\n      \"year\": 2023,\n      \"claim\": \"Comparative genomics showed GUCY2F is dispensable in multiple vertebrate clades where GCAP expansion compensates, while Gucy2f-/- mice were used as tools for GC-E gene therapy optimization, confirming partial functional redundancy between the two cyclases.\",\n      \"evidence\": \"Phylogenetic analysis across vertebrates; Gucy2f knockout mice with AAV-CRISPR gene therapy and ERG\",\n      \"pmids\": [\"37007786\", \"37361352\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\n        \"Evolutionary analysis is computational and lacks biochemical validation of GCAP compensation\",\n        \"Detailed electrophysiological characterization of Gucy2f single-knockout mouse rods and cones is incomplete\"\n      ]\n    },\n    {\n      \"year\": null,\n      \"claim\": \"The high-resolution structure of GUCY2F, the precise mechanism by which it contributes to outer segment protein trafficking, the identity of any extracellular ligand, and whether GUCY2F mutations cause human retinal disease remain unresolved.\",\n      \"evidence\": \"\",\n      \"pmids\": [],\n      \"confidence\": \"Low\",\n      \"gaps\": [\n        \"No atomic-resolution structure of GUCY2F or its complex with GCAPs\",\n        \"No human Mendelian disease definitively linked to GUCY2F mutations\",\n        \"Extracellular domain function remains entirely unknown\"\n      ]\n    }\n  ],\n  \"mechanism_profile\": {\n    \"molecular_activity\": [\n      {\"term_id\": \"GO:0009975\", \"supporting_discovery_ids\": [0, 1, 2, 5, 13]}\n    ],\n    \"localization\": [\n      {\"term_id\": \"GO:0005886\", \"supporting_discovery_ids\": [0, 1]}\n    ],\n    \"pathway\": [\n      {\"term_id\": \"R-HSA-9709957\", \"supporting_discovery_ids\": [0, 10, 11, 15]},\n      {\"term_id\": \"R-HSA-162582\", \"supporting_discovery_ids\": [5, 6, 13]}\n    ],\n    \"complexes\": [],\n    \"partners\": [\n      \"GUCA1B\",\n      \"GUCA1A\",\n      \"RD3\"\n    ],\n    \"other_free_text\": []\n  }\n}\n```"}