{"gene":"LRRC8D","run_date":"2026-06-10T02:59:50","timeline":{"discoveries":[{"year":2014,"finding":"LRRC8D is required for import of the antibiotic blasticidin S into mammalian cells, demonstrating a transport function. LRRC8D localizes to the plasma membrane and physically interacts with LRRC8A, LRRC8B, and LRRC8C.","method":"Genetic screen/loss-of-function, co-immunoprecipitation, localization and topology assays","journal":"The Journal of biological chemistry","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — functional KO phenotype with specific substrate readout plus co-IP for interactions, single lab but orthogonal methods","pmids":["24782309"],"is_preprint":false},{"year":2015,"finding":"LRRC8D is a subunit of heteromeric volume-regulated anion channels (VRACs). LRRC8A/LRRC8D-containing VRACs mediate ~50% of cellular cisplatin/carboplatin uptake under isotonic conditions, and cell swelling strongly enhances this uptake. Incorporation of LRRC8D increases VRAC permeability for cisplatin and taurine relative to channels lacking LRRC8D, indicating LRRC8D contributes to pore properties.","method":"CRISPR/siRNA knockout, isotopic drug uptake assays, electrophysiology, genetic rescue","journal":"The EMBO journal","confidence":"High","confidence_rationale":"Tier 2 / Strong — multiple orthogonal methods (KO, uptake assays, electrophysiology), replicated by subsequent studies","pmids":["26530471"],"is_preprint":false},{"year":2017,"finding":"In primary rat astrocytes, LRRC8A/D-containing heteromeric VRACs preferentially mediate swelling-activated release of uncharged osmolytes (taurine, myo-inositol), whereas charged osmolyte (d-aspartate) release depends on LRRC8A/C/E-containing channels. LRRC8D knockdown strongly inhibits taurine and myo-inositol efflux without major effect on d-aspartate efflux, establishing subunit-specific substrate selectivity.","method":"RNAi knockdown, radiotracer flux assays with hypoosmotic challenge","journal":"The Journal of physiology","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — multiple substrates measured simultaneously with RNAi, single lab, replicated conceptually by other groups","pmids":["28833202"],"is_preprint":false},{"year":2017,"finding":"LRRC8A/D heteromeric channels are strongly inhibited by oxidation of intracellular cysteine residues (by chloramine-T or tert-butyl hydroperoxide), in contrast to LRRC8A/E heteromers which are potentiated. The extracellular loop domains (EL1, EL2) of LRRC8D contain external oxidation sites that confer susceptibility to oxidant-mediated inhibition.","method":"Electrophysiology of heterologously expressed fluorescently tagged LRRC8 heteromers, pharmacological oxidation, domain-swap chimeras","journal":"The Journal of physiology","confidence":"Medium","confidence_rationale":"Tier 1-2 / Moderate — direct electrophysiological measurement on defined heteromers plus chimera mutagenesis, single lab","pmids":["28841766","33932953"],"is_preprint":false},{"year":2018,"finding":"Pancreatic islets prominently express LRRC8D alongside LRRC8A; LRRC8D-containing VRACs may mediate neurotransmitter-permeable channel activity in beta-cells, contributing to autocrine/paracrine signaling within islets. Loss of the essential subunit LRRC8A (which partners with LRRC8D) reduces first-phase glucose-induced insulin secretion and impairs glucose tolerance.","method":"Conditional knockout mice, patch-clamp electrophysiology, insulin secretion assays, glucose tolerance tests","journal":"Nature communications","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — conditional KO with defined cellular phenotype and multiple readouts; LRRC8D role inferred from expression and subunit composition, not direct KO of LRRC8D alone","pmids":["29773801"],"is_preprint":false},{"year":2018,"finding":"The first extracellular loop (EL1) of LRRC8D (connecting transmembrane domains 1 and 2) is essential for VRAC activity. Chimeric channels in which EL1 of LRRC8A is replaced with that of LRRC8D confer functional VRAC activity and volume-dependent regulation.","method":"Chimeric channel mutagenesis, whole-cell patch clamp, hypotonicity-induced current recording","journal":"The Journal of general physiology","confidence":"Medium","confidence_rationale":"Tier 1 / Moderate — mutagenesis with functional electrophysiological readout, single lab","pmids":["29853476"],"is_preprint":false},{"year":2020,"finding":"Cryo-EM structure of the human LRRC8D homo-hexamer reveals a two-fold symmetric arrangement. The extracellular pore constriction of LRRC8D is wider than in LRRC8A structures, explaining increased permeability for organic substrates. An N-terminal helix protrudes into the pore from the intracellular side and is implicated in gating. Structure-based electrophysiological mutagenesis confirmed these functional features.","method":"Cryo-EM structure determination, electrophysiology (structure-based mutagenesis)","journal":"Communications biology","confidence":"High","confidence_rationale":"Tier 1 / Moderate — high-resolution cryo-EM structure combined with electrophysiological validation of structural features, single lab but orthogonal methods","pmids":["32415200"],"is_preprint":false},{"year":2020,"finding":"LRRC8D inhibits cGAMP transport through VRAC. LRRC8A:C and LRRC8A:E heteromers transport cGAMP and other 2'3'-cyclic dinucleotides, whereas LRRC8D incorporation into the complex reduces this transport activity.","method":"CRISPR knockout screens, genetic rescue with individual subunits, cGAMP transport assays, STING pathway activation readout","journal":"Molecular cell","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — functional assay with multiple subunit knockouts and defined transport readout, single lab","pmids":["33171122"],"is_preprint":false},{"year":2021,"finding":"LRRC8D co-immunoprecipitates with NADPH oxidase 1 (Nox1) in vascular smooth muscle cells and co-localizes with Nox1 at the plasma membrane and in vesicles. LRRC8D knockdown potentiates NF-κB activation in response to TNFα, suggesting LRRC8D-containing VRACs negatively modulate this inflammatory signaling pathway.","method":"Co-immunoprecipitation, siRNA knockdown, NF-κB reporter assay, superoxide production assay","journal":"The Journal of physiology","confidence":"Medium","confidence_rationale":"Tier 2-3 / Moderate — co-IP plus functional knockdown with multiple readouts, single lab","pmids":["33932953"],"is_preprint":false},{"year":2022,"finding":"LRRC8D localizes to basolateral membranes of proximal tubules in the kidney. Constitutive deletion of Lrrc8d in mice causes proximal tubular injury, increased diuresis, and mild Fanconi-like symptoms, establishing LRRC8A/D channels as required for basolateral exit of organic compounds including metabolites in proximal tubules.","method":"Epitope-tagged knock-in mice for localization, constitutive Lrrc8d knockout mice, histology, urine/serum metabolomics","journal":"Journal of the American Society of Nephrology : JASN","confidence":"High","confidence_rationale":"Tier 2 / Strong — direct localization by knock-in tags plus KO with multiple functional readouts (histology, metabolomics, urine analysis), replicated across subunit deletions","pmids":["35777784"],"is_preprint":false},{"year":2022,"finding":"Loss of LRRC8D significantly reduces cis- and carboplatin uptake in BRCA1;p53-deficient mouse mammary tumor cells, resulting in reduced DNA damage and in vivo drug resistance. Lrrc8d deletion in mice does not affect viability or fertility, unlike Lrrc8a deletion.","method":"Conditional/constitutive gene knockout in mice, platinum drug uptake measurement, DNA damage assays, in vivo tumor treatment","journal":"Cancer research communications","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — in vivo KO with multiple mechanistic readouts, single lab, confirms prior in vitro findings","pmids":["36467895"],"is_preprint":false},{"year":2024,"finding":"In the cochlea, LRRC8D is expressed in sensory hair cells and in the stria vascularis (alongside LRRC8A and LRRC8E). Combined ablation of LRRC8D and LRRC8E results in cochlear degeneration and congenital deafness, with severe reduction in the endocochlear potential and loss of Kir4.1 (KCNJ10) expression, suggesting LRRC8D-containing VRACs transport metabolites (e.g., glutathione) needed to maintain inner ear redox potential.","method":"Conditional knockout mice, auditory brainstem response, endocochlear potential measurement, immunohistochemistry, Western blot","journal":"The Journal of biological chemistry","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — conditional double KO with defined electrophysiological and molecular phenotypes, single lab","pmids":["38838775"],"is_preprint":false},{"year":2025,"finding":"NAA60, an N-terminal acetyltransferase localized to the Golgi apparatus, acetylates the N-termini of LRRC8A and LRRC8D. Loss of NAA60 decreases cis- and carboplatin uptake; introduction of positively charged amino acids at the LRRC8A/D N-termini (mimicking absence of acetylation) similarly decreases platinum drug sensitivity, establishing N-terminal acetylation as a post-translational modification required for LRRC8A/D-mediated drug uptake.","method":"Co-immunoprecipitation/mass spectrometry, N-terminal acetylation assays, site-directed mutagenesis (charge substitution), drug uptake assays, in vivo tumor models","journal":"Communications biology","confidence":"Medium","confidence_rationale":"Tier 1-2 / Moderate — mutagenesis combined with functional uptake assay and in vivo validation, single lab, multiple orthogonal methods","pmids":["41053424"],"is_preprint":false},{"year":2024,"finding":"Cryo-EM structures of a native-like LRRC8A:D VRAC with 4:2 stoichiometry reveal that LRRC8D subunits increase hydrophobicity and widen the selectivity filter compared to LRRC8A homomers, explaining LRRC8D-unique substrate selectivity. Lipids occupy the pore in the closed state (as in LRRC8A:C VRACs). Incorporation of LRRC8D disrupts packing of cytoplasmic LRR domains, increases channel dynamics, and opens lateral fenestrations proposed to be necessary for pore lipid evacuation and channel activation. Electrophysiology confirmed lipid-gating is a general VRAC property.","method":"Cryo-EM structure determination (4:2 stoichiometry), electrophysiology","journal":"bioRxiv","confidence":"Medium","confidence_rationale":"Tier 1 / Weak — cryo-EM structure with electrophysiological validation but preprint, single lab","pmids":["bio_10.1101_2024.11.24.625074"],"is_preprint":true},{"year":2025,"finding":"In lateral ventricular neural stem cells, LRRC8D (a core VRAC component) participates in GABA transport/signaling and provides negative feedback to ChAT+ neurons in the ACC-subependymal circuit, with evidence that LRRC8D-regulated chloride and GABA transport modulates neural stem cell proliferation.","method":"Immunohistochemistry, in vivo circuit activation, functional behavioral/proliferation readouts","journal":"Cells","confidence":"Low","confidence_rationale":"Tier 3 / Weak — single study, correlative localization and circuit manipulation without direct biochemical mechanistic dissection of LRRC8D's molecular role","pmids":["40136675"],"is_preprint":false}],"current_model":"LRRC8D is a facultative pore-forming subunit of heteromeric volume-regulated anion channels (VRACs), assembling with the obligatory LRRC8A subunit (and sometimes LRRC8B/C/E) as a hexamer with 4:2 stoichiometry; its incorporation widens the selectivity filter and increases pore hydrophobicity to confer elevated permeability for organic osmolytes (taurine, myo-inositol), platinum-based drugs (cisplatin, carboplatin), and blasticidin S, while inhibiting cGAMP transport; LRRC8D-containing channels are directly inhibited by oxidation of their extracellular loop cysteines, negatively modulate Nox1/NF-κB inflammatory signaling, and are functionally regulated by N-terminal acetylation by NAA60; in vivo, LRRC8A/D channels are required at the basolateral membrane of renal proximal tubules for metabolite export, and combined loss of LRRC8D/E causes cochlear degeneration and deafness."},"narrative":{"mechanistic_narrative":"LRRC8D is a facultative pore-forming subunit of heteromeric volume-regulated anion channels (VRACs), which it forms by assembling at the plasma membrane with the obligatory LRRC8A subunit (and LRRC8B/C) [PMID:24782309, PMID:26530471]. Cryo-EM of a native-like LRRC8A:D channel with 4:2 stoichiometry shows that incorporation of LRRC8D widens the extracellular selectivity-filter constriction and increases pore hydrophobicity relative to LRRC8A homomers, the structural basis for its distinctive substrate profile [PMID:32415200, PMID:bio_10.1101_2024.11.24.625074]. Functionally, LRRC8D-containing channels confer elevated permeability to uncharged organic osmolytes such as taurine and myo-inositol and to platinum-based drugs (cisplatin, carboplatin), while charged osmolyte release depends on LRRC8A/C/E channels [PMID:26530471, PMID:28833202]. This drug-permeation activity is physiologically and clinically consequential: LRRC8D-mediated platinum uptake drives DNA damage in tumor cells, and its loss confers in vivo platinum resistance [PMID:36467895], and the activity requires N-terminal acetylation of LRRC8A/D by the Golgi acetyltransferase NAA60 [PMID:41053424]. LRRC8D incorporation conversely suppresses channel transport of the immune second messenger cGAMP [PMID:33171122]. LRRC8D-containing channels are gated and modulated by their environment — directly inhibited by oxidation of extracellular-loop cysteines [PMID:28841766, PMID:33932953], and lipid-occupied in the closed state with LRRC8D-induced lateral fenestrations proposed to permit pore lipid evacuation upon activation [PMID:bio_10.1101_2024.11.24.625074]. In vivo, LRRC8A/D channels reside at the basolateral membrane of renal proximal tubules where they are required for export of organic metabolites [PMID:35777784], and combined loss of LRRC8D and LRRC8E causes cochlear degeneration and congenital deafness with collapse of the endocochlear potential [PMID:38838775].","teleology":[{"year":2014,"claim":"Established that LRRC8D has a transport function and physically assembles with other LRRC8 family members, answering whether LRRC8D is a plasma-membrane transport component.","evidence":"Genetic loss-of-function screen for blasticidin S import plus co-immunoprecipitation and topology assays in mammalian cells","pmids":["24782309"],"confidence":"Medium","gaps":["Did not define the channel identity or that VRAC is the relevant conductance","Substrate scope limited to blasticidin S at this stage"]},{"year":2015,"claim":"Identified LRRC8D as a VRAC subunit that shapes pore permeability, answering how LRRC8D contributes to anion-channel function and explaining its role in chemotherapy uptake.","evidence":"CRISPR/siRNA knockout, isotopic platinum-drug uptake assays, electrophysiology and genetic rescue","pmids":["26530471"],"confidence":"High","gaps":["Structural basis of altered permeability not yet resolved","Stoichiometry of LRRC8D within the hexamer undefined"]},{"year":2017,"claim":"Showed subunit-specific substrate selectivity, answering which osmolytes LRRC8D-containing channels preferentially conduct versus other heteromers.","evidence":"RNAi knockdown with radiotracer flux of taurine, myo-inositol and d-aspartate under hypoosmotic challenge in primary astrocytes","pmids":["28833202"],"confidence":"Medium","gaps":["Selectivity inferred from knockdown rather than reconstituted defined channels","Single cell type"]},{"year":2017,"claim":"Defined redox regulation of LRRC8D channels, answering how oxidation differentially modulates VRAC heteromers and localizing the oxidation-sensitive sites.","evidence":"Electrophysiology of defined fluorescently tagged heteromers with pharmacological oxidation and domain-swap chimeras","pmids":["28841766","33932953"],"confidence":"Medium","gaps":["Specific cysteine residues conferring inhibition not fully enumerated","Physiological oxidant relevant in vivo not established"]},{"year":2018,"claim":"Mapped the first extracellular loop as essential for channel activity, answering which LRRC8D domain enables functional VRAC assembly and gating.","evidence":"Chimeric channel mutagenesis with whole-cell patch clamp under hypotonicity","pmids":["29853476"],"confidence":"Medium","gaps":["Mechanism by which EL1 confers volume sensing unresolved","Single-subunit chimera context"]},{"year":2018,"claim":"Implicated LRRC8D channels in pancreatic islet signaling, addressing a physiological context for LRRC8D-containing VRAC activity.","evidence":"Conditional LRRC8A knockout mice with patch-clamp, insulin secretion and glucose tolerance tests","pmids":["29773801"],"confidence":"Medium","gaps":["LRRC8D role inferred from expression and partnership, not direct LRRC8D knockout","Permeant signaling molecule not directly identified"]},{"year":2020,"claim":"Provided a structural explanation for LRRC8D permeability, answering why LRRC8D channels conduct large organic substrates.","evidence":"Cryo-EM of the human LRRC8D homo-hexamer with structure-based electrophysiological mutagenesis","pmids":["32415200"],"confidence":"High","gaps":["Homo-hexamer is not the physiological heteromeric assembly","Gating transitions not captured"]},{"year":2020,"claim":"Showed LRRC8D inhibits cGAMP transport, answering how subunit composition selects against immune second-messenger permeation.","evidence":"CRISPR knockout screens, subunit rescue, cGAMP transport assays and STING activation readouts","pmids":["33171122"],"confidence":"Medium","gaps":["Structural basis for cGAMP exclusion by LRRC8D not defined","Single-lab functional inference"]},{"year":2021,"claim":"Linked LRRC8D to inflammatory signaling, answering whether LRRC8D-containing channels modulate Nox1/NF-κB pathways.","evidence":"Co-immunoprecipitation with Nox1, siRNA knockdown, NF-κB reporter and superoxide assays in vascular smooth muscle cells","pmids":["33932953"],"confidence":"Medium","gaps":["Co-IP without reciprocal validation","Direct versus indirect modulation of NF-κB unresolved"]},{"year":2022,"claim":"Established an in vivo physiological role in the kidney, answering whether LRRC8A/D channels export metabolites at the basolateral proximal tubule membrane.","evidence":"Epitope-tagged knock-in localization and constitutive Lrrc8d knockout mice with histology and urine/serum metabolomics","pmids":["35777784"],"confidence":"High","gaps":["Specific exported metabolites only partially defined","Tissue-specific contribution versus systemic effects not fully separated"]},{"year":2022,"claim":"Demonstrated in vivo chemotherapy relevance, answering whether LRRC8D loss confers platinum resistance and whether LRRC8D is dispensable for organismal viability.","evidence":"Conditional/constitutive knockout mice with platinum uptake, DNA damage assays and tumor treatment","pmids":["36467895"],"confidence":"Medium","gaps":["Mechanism of residual platinum uptake via other subunits not quantified","Clinical translation untested"]},{"year":2024,"claim":"Defined a cochlear requirement, answering the in vivo consequence of losing LRRC8D-containing channels in the inner ear.","evidence":"Conditional LRRC8D/E double knockout mice with auditory brainstem response, endocochlear potential measurement and immunohistochemistry","pmids":["38838775"],"confidence":"Medium","gaps":["Glutathione as the critical transported metabolite is inferential","LRRC8D-specific contribution versus LRRC8E not separated"]},{"year":2024,"claim":"Resolved the native-like heteromeric architecture and lipid-gating, answering how LRRC8D incorporation alters the pore and channel dynamics in the physiological 4:2 assembly.","evidence":"Cryo-EM of LRRC8A:D VRAC at 4:2 stoichiometry with electrophysiology (preprint)","pmids":["bio_10.1101_2024.11.24.625074"],"confidence":"Medium","gaps":["Preprint, not peer-reviewed","Lipid-evacuation gating model awaits independent confirmation"]},{"year":2025,"claim":"Identified N-terminal acetylation as a required post-translational modification, answering how LRRC8A/D channels are regulated for drug uptake.","evidence":"Co-IP/mass spectrometry, acetylation and charge-substitution mutagenesis, drug uptake assays and in vivo tumor models","pmids":["41053424"],"confidence":"Medium","gaps":["Mechanism by which N-terminal charge affects gating/permeation not structurally defined","Single-lab finding"]},{"year":2025,"claim":"Proposed a neural stem cell signaling role, addressing whether LRRC8D modulates GABA/chloride transport in a defined circuit.","evidence":"Immunohistochemistry, in vivo circuit activation and proliferation/behavioral readouts","pmids":["40136675"],"confidence":"Low","gaps":["Correlative localization without direct biochemical dissection of LRRC8D's molecular role","GABA permeation through LRRC8D channels not directly measured"]},{"year":null,"claim":"How LRRC8D subunit incorporation is regulated to tune channel composition and substrate selectivity across tissues, and the gating transitions linking volume sensing, lipid evacuation and pore opening, remain unresolved.","evidence":"","pmids":[],"confidence":"Medium","gaps":["No structure of an open/activated heteromeric channel","Signals controlling tissue-specific subunit assembly unknown","Full physiological substrate repertoire incomplete"]}],"mechanism_profile":{"molecular_activity":[{"term_id":"GO:0005215","term_label":"transporter activity","supporting_discovery_ids":[0,1,2,7,9]},{"term_id":"GO:0005198","term_label":"structural molecule activity","supporting_discovery_ids":[6,13]}],"localization":[{"term_id":"GO:0005886","term_label":"plasma membrane","supporting_discovery_ids":[0,8,9]},{"term_id":"GO:0031410","term_label":"cytoplasmic vesicle","supporting_discovery_ids":[8]}],"pathway":[{"term_id":"R-HSA-382551","term_label":"Transport of small molecules","supporting_discovery_ids":[1,2,9,10]},{"term_id":"R-HSA-8953897","term_label":"Cellular responses to stimuli","supporting_discovery_ids":[1,2,3]}],"complexes":["Volume-regulated anion channel (VRAC) LRRC8A:D heteromer"],"partners":["LRRC8A","LRRC8B","LRRC8C","LRRC8E","NOX1","NAA60"],"other_free_text":[]}},"prefetch_data":{"uniprot":{"accession":"Q7L1W4","full_name":"Volume-regulated anion channel subunit LRRC8D","aliases":["Leucine-rich repeat-containing protein 5","Leucine-rich repeat-containing protein 8D","HsLRRC8D"],"length_aa":858,"mass_kda":98.2,"function":"Non-essential component of the volume-regulated anion channel (VRAC, also named VSOAC channel), an anion channel required to maintain a constant cell volume in response to extracellular or intracellular osmotic changes (PubMed:24790029, PubMed:26530471, PubMed:26824658, PubMed:28193731, PubMed:32415200). The VRAC channel conducts iodide better than chloride and can also conduct organic osmolytes like taurine (PubMed:24790029, PubMed:26824658, PubMed:28193731). Plays a redundant role in the efflux of amino acids, such as aspartate, in response to osmotic stress (PubMed:28193731). LRRC8A and LRRC8D are required for the uptake of the drug cisplatin (PubMed:26530471). Channel activity requires LRRC8A plus at least one other family member (LRRC8B, LRRC8C, LRRC8D or LRRC8E); channel characteristics depend on the precise subunit composition (PubMed:24782309, PubMed:24790029, PubMed:26824658, PubMed:28193731). Also acts as a regulator of glucose-sensing in pancreatic beta cells: VRAC currents, generated in response to hypotonicity- or glucose-induced beta cell swelling, depolarize cells, thereby causing electrical excitation, leading to increase glucose sensitivity and insulin secretion (By similarity). VRAC channels containing LRRC8D inhibit transport of immunoreactive cyclic dinucleotide GMP-AMP (2'-3'-cGAMP), an immune messenger produced in response to DNA virus in the cytosol (PubMed:33171122). Mediates the import of the antibiotic blasticidin-S into the cell (PubMed:24782309)","subcellular_location":"Cell membrane; Endoplasmic reticulum membrane","url":"https://www.uniprot.org/uniprotkb/Q7L1W4/entry"},"depmap":{"release":"DepMap","has_data":true,"is_common_essential":false,"resolved_as":"","url":"https://depmap.org/portal/gene/LRRC8D","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/LRRC8D","total_profiled":1310},"omim":[{"mim_id":"612890","title":"LEUCINE-RICH REPEAT-CONTAINING PROTEIN 8D; LRRC8D","url":"https://www.omim.org/entry/612890"}],"hpa":{"profiled":true,"resolved_as":"","reliability":"","locations":[],"tissue_specificity":"Low tissue specificity","tissue_distribution":"Detected in all","driving_tissues":[],"url":"https://www.proteinatlas.org/search/LRRC8D"},"hgnc":{"alias_symbol":["FLJ10470"],"prev_symbol":["LRRC5"]},"alphafold":{"accession":"Q7L1W4","domains":[{"cath_id":"1.20.1440","chopping":"5-58_161-187_308-392","consensus_level":"medium","plddt":89.0257,"start":5,"end":392},{"cath_id":"3.80.10.10","chopping":"671-851","consensus_level":"medium","plddt":93.0822,"start":671,"end":851}],"viewer_url":"https://alphafold.ebi.ac.uk/entry/Q7L1W4","model_url":"https://alphafold.ebi.ac.uk/files/AF-Q7L1W4-F1-model_v6.cif","pae_url":"https://alphafold.ebi.ac.uk/files/AF-Q7L1W4-F1-predicted_aligned_error_v6.png","plddt_mean":80.75},"mouse_models":{"mgi_url":"https://www.informatics.jax.org/marker/summary?nomen=LRRC8D","jax_strain_url":"https://www.jax.org/strain/search?query=LRRC8D"},"sequence":{"accession":"Q7L1W4","fasta_url":"https://rest.uniprot.org/uniprotkb/Q7L1W4.fasta","uniprot_url":"https://www.uniprot.org/uniprotkb/Q7L1W4/entry","alphafold_viewer_url":"https://alphafold.ebi.ac.uk/entry/Q7L1W4"}},"corpus_meta":[{"pmid":"26530471","id":"PMC_26530471","title":"Subunit composition of VRAC channels determines substrate specificity and cellular resistance to Pt-based anti-cancer drugs.","date":"2015","source":"The EMBO journal","url":"https://pubmed.ncbi.nlm.nih.gov/26530471","citation_count":225,"is_preprint":false},{"pmid":"33171122","id":"PMC_33171122","title":"LRRC8A:C/E Heteromeric Channels Are Ubiquitous Transporters of cGAMP.","date":"2020","source":"Molecular cell","url":"https://pubmed.ncbi.nlm.nih.gov/33171122","citation_count":155,"is_preprint":false},{"pmid":"28833202","id":"PMC_28833202","title":"Molecular composition and heterogeneity of the LRRC8-containing swelling-activated osmolyte channels in primary rat astrocytes.","date":"2017","source":"The Journal of physiology","url":"https://pubmed.ncbi.nlm.nih.gov/28833202","citation_count":85,"is_preprint":false},{"pmid":"29773801","id":"PMC_29773801","title":"LRRC8/VRAC anion channels enhance β-cell glucose sensing and insulin secretion.","date":"2018","source":"Nature communications","url":"https://pubmed.ncbi.nlm.nih.gov/29773801","citation_count":85,"is_preprint":false},{"pmid":"26635246","id":"PMC_26635246","title":"VRAC: molecular identification as LRRC8 heteromers with differential functions.","date":"2015","source":"Pflugers Archiv : European journal of physiology","url":"https://pubmed.ncbi.nlm.nih.gov/26635246","citation_count":70,"is_preprint":false},{"pmid":"24782309","id":"PMC_24782309","title":"The protein synthesis inhibitor blasticidin s enters mammalian cells via leucine-rich repeat-containing protein 8D.","date":"2014","source":"The Journal of biological chemistry","url":"https://pubmed.ncbi.nlm.nih.gov/24782309","citation_count":67,"is_preprint":false},{"pmid":"28841766","id":"PMC_28841766","title":"Subunit-dependent oxidative stress sensitivity of LRRC8 volume-regulated anion channels.","date":"2017","source":"The Journal of physiology","url":"https://pubmed.ncbi.nlm.nih.gov/28841766","citation_count":48,"is_preprint":false},{"pmid":"15094057","id":"PMC_15094057","title":"LRRC8 involved in B cell development belongs to a novel family of leucine-rich repeat proteins.","date":"2004","source":"FEBS letters","url":"https://pubmed.ncbi.nlm.nih.gov/15094057","citation_count":47,"is_preprint":false},{"pmid":"32415200","id":"PMC_32415200","title":"Cryo-EM structure of the volume-regulated anion channel LRRC8D isoform identifies features important for substrate permeation.","date":"2020","source":"Communications biology","url":"https://pubmed.ncbi.nlm.nih.gov/32415200","citation_count":44,"is_preprint":false},{"pmid":"29853476","id":"PMC_29853476","title":"Intracellular and extracellular loops of LRRC8 are essential for volume-regulated anion channel function.","date":"2018","source":"The Journal of general physiology","url":"https://pubmed.ncbi.nlm.nih.gov/29853476","citation_count":39,"is_preprint":false},{"pmid":"27764579","id":"PMC_27764579","title":"Specific and essential but not sufficient roles of LRRC8A in the activity of volume-sensitive outwardly rectifying anion channel (VSOR).","date":"2016","source":"Channels (Austin, Tex.)","url":"https://pubmed.ncbi.nlm.nih.gov/27764579","citation_count":33,"is_preprint":false},{"pmid":"33577730","id":"PMC_33577730","title":"Mechanisms of Activation of LRRC8 Volume Regulated Anion Channels.","date":"2021","source":"Cellular physiology and biochemistry : international journal of experimental cellular physiology, biochemistry, and pharmacology","url":"https://pubmed.ncbi.nlm.nih.gov/33577730","citation_count":29,"is_preprint":false},{"pmid":"32442571","id":"PMC_32442571","title":"Volume-regulated anion channel as a novel cancer therapeutic target.","date":"2020","source":"International journal of biological 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Cell physiology","url":"https://pubmed.ncbi.nlm.nih.gov/33356947","citation_count":23,"is_preprint":false},{"pmid":"33932953","id":"PMC_33932953","title":"Oxidant-resistant LRRC8A/C anion channels support superoxide production by NADPH oxidase 1.","date":"2021","source":"The Journal of physiology","url":"https://pubmed.ncbi.nlm.nih.gov/33932953","citation_count":22,"is_preprint":false},{"pmid":"37467717","id":"PMC_37467717","title":"A target discovery pipeline identified ILT3 as a target for immunotherapy of multiple myeloma.","date":"2023","source":"Cell reports. 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Switzerland)","url":"https://pubmed.ncbi.nlm.nih.gov/32485798","citation_count":9,"is_preprint":false},{"pmid":"36719767","id":"PMC_36719767","title":"Epigenome-Wide Association Study Reveals CpG Sites Associated with Thyroid Function and Regulatory Effects on KLF9.","date":"2023","source":"Thyroid : official journal of the American Thyroid Association","url":"https://pubmed.ncbi.nlm.nih.gov/36719767","citation_count":8,"is_preprint":false},{"pmid":"26564094","id":"PMC_26564094","title":"VRACs swallow platinum drugs.","date":"2015","source":"The EMBO journal","url":"https://pubmed.ncbi.nlm.nih.gov/26564094","citation_count":8,"is_preprint":false},{"pmid":"36162674","id":"PMC_36162674","title":"Circuit-wide proteomics profiling reveals brain region-specific protein signatures in the male WKY rats with endogenous depression.","date":"2022","source":"Journal of affective 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Cell physiology","url":"https://pubmed.ncbi.nlm.nih.gov/41740631","citation_count":0,"is_preprint":false},{"pmid":"40136675","id":"PMC_40136675","title":"Lateral Ventricular Neural Stem Cells Provide Negative Feedback to Circuit Activation Through GABAergic Signaling.","date":"2025","source":"Cells","url":"https://pubmed.ncbi.nlm.nih.gov/40136675","citation_count":0,"is_preprint":false},{"pmid":"41300786","id":"PMC_41300786","title":"Chromothriptic Translocation t(1;18): A Paradigm of Genomic Complexity in a Child with Normal Intellectual Development and Pyridoxine-Dependent Epilepsy.","date":"2025","source":"Genes","url":"https://pubmed.ncbi.nlm.nih.gov/41300786","citation_count":0,"is_preprint":false},{"pmid":null,"id":"bio_10.1101_2024.11.24.625074","title":"Assembly and lipid-gating of LRRC8A:D volume-regulated anion channels","date":"2024-11-26","source":"bioRxiv","url":"https://doi.org/10.1101/2024.11.24.625074","citation_count":0,"is_preprint":true}],"cost":{"stage1":{"model":"claude-sonnet-4-6","input_tokens":23235,"output_tokens":3952,"usd":0.064492,"stage1_stop_reason":"end_turn"},"stage2":{"model":"claude-opus-4-8","input_tokens":11637,"output_tokens":4348,"usd":0.083443,"stage2_stop_reason":"end_turn"},"total_usd":0.147935,"stage1_batch_id":"msgbatch_01BbTCMyrRPyhtEq6b1MdwCN","stage2_batch_id":"msgbatch_011dEed5bbxYvTAaQWKJevxu","note":"batch pricing = 50% of standard"},"stage1_raw":"```json\n{\n  \"discoveries\": [\n    {\n      \"year\": 2014,\n      \"finding\": \"LRRC8D is required for import of the antibiotic blasticidin S into mammalian cells, demonstrating a transport function. LRRC8D localizes to the plasma membrane and physically interacts with LRRC8A, LRRC8B, and LRRC8C.\",\n      \"method\": \"Genetic screen/loss-of-function, co-immunoprecipitation, localization and topology assays\",\n      \"journal\": \"The Journal of biological chemistry\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — functional KO phenotype with specific substrate readout plus co-IP for interactions, single lab but orthogonal methods\",\n      \"pmids\": [\"24782309\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2015,\n      \"finding\": \"LRRC8D is a subunit of heteromeric volume-regulated anion channels (VRACs). LRRC8A/LRRC8D-containing VRACs mediate ~50% of cellular cisplatin/carboplatin uptake under isotonic conditions, and cell swelling strongly enhances this uptake. Incorporation of LRRC8D increases VRAC permeability for cisplatin and taurine relative to channels lacking LRRC8D, indicating LRRC8D contributes to pore properties.\",\n      \"method\": \"CRISPR/siRNA knockout, isotopic drug uptake assays, electrophysiology, genetic rescue\",\n      \"journal\": \"The EMBO journal\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — multiple orthogonal methods (KO, uptake assays, electrophysiology), replicated by subsequent studies\",\n      \"pmids\": [\"26530471\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2017,\n      \"finding\": \"In primary rat astrocytes, LRRC8A/D-containing heteromeric VRACs preferentially mediate swelling-activated release of uncharged osmolytes (taurine, myo-inositol), whereas charged osmolyte (d-aspartate) release depends on LRRC8A/C/E-containing channels. LRRC8D knockdown strongly inhibits taurine and myo-inositol efflux without major effect on d-aspartate efflux, establishing subunit-specific substrate selectivity.\",\n      \"method\": \"RNAi knockdown, radiotracer flux assays with hypoosmotic challenge\",\n      \"journal\": \"The Journal of physiology\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — multiple substrates measured simultaneously with RNAi, single lab, replicated conceptually by other groups\",\n      \"pmids\": [\"28833202\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2017,\n      \"finding\": \"LRRC8A/D heteromeric channels are strongly inhibited by oxidation of intracellular cysteine residues (by chloramine-T or tert-butyl hydroperoxide), in contrast to LRRC8A/E heteromers which are potentiated. The extracellular loop domains (EL1, EL2) of LRRC8D contain external oxidation sites that confer susceptibility to oxidant-mediated inhibition.\",\n      \"method\": \"Electrophysiology of heterologously expressed fluorescently tagged LRRC8 heteromers, pharmacological oxidation, domain-swap chimeras\",\n      \"journal\": \"The Journal of physiology\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 1-2 / Moderate — direct electrophysiological measurement on defined heteromers plus chimera mutagenesis, single lab\",\n      \"pmids\": [\"28841766\", \"33932953\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2018,\n      \"finding\": \"Pancreatic islets prominently express LRRC8D alongside LRRC8A; LRRC8D-containing VRACs may mediate neurotransmitter-permeable channel activity in beta-cells, contributing to autocrine/paracrine signaling within islets. Loss of the essential subunit LRRC8A (which partners with LRRC8D) reduces first-phase glucose-induced insulin secretion and impairs glucose tolerance.\",\n      \"method\": \"Conditional knockout mice, patch-clamp electrophysiology, insulin secretion assays, glucose tolerance tests\",\n      \"journal\": \"Nature communications\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — conditional KO with defined cellular phenotype and multiple readouts; LRRC8D role inferred from expression and subunit composition, not direct KO of LRRC8D alone\",\n      \"pmids\": [\"29773801\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2018,\n      \"finding\": \"The first extracellular loop (EL1) of LRRC8D (connecting transmembrane domains 1 and 2) is essential for VRAC activity. Chimeric channels in which EL1 of LRRC8A is replaced with that of LRRC8D confer functional VRAC activity and volume-dependent regulation.\",\n      \"method\": \"Chimeric channel mutagenesis, whole-cell patch clamp, hypotonicity-induced current recording\",\n      \"journal\": \"The Journal of general physiology\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 1 / Moderate — mutagenesis with functional electrophysiological readout, single lab\",\n      \"pmids\": [\"29853476\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2020,\n      \"finding\": \"Cryo-EM structure of the human LRRC8D homo-hexamer reveals a two-fold symmetric arrangement. The extracellular pore constriction of LRRC8D is wider than in LRRC8A structures, explaining increased permeability for organic substrates. An N-terminal helix protrudes into the pore from the intracellular side and is implicated in gating. Structure-based electrophysiological mutagenesis confirmed these functional features.\",\n      \"method\": \"Cryo-EM structure determination, electrophysiology (structure-based mutagenesis)\",\n      \"journal\": \"Communications biology\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 / Moderate — high-resolution cryo-EM structure combined with electrophysiological validation of structural features, single lab but orthogonal methods\",\n      \"pmids\": [\"32415200\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2020,\n      \"finding\": \"LRRC8D inhibits cGAMP transport through VRAC. LRRC8A:C and LRRC8A:E heteromers transport cGAMP and other 2'3'-cyclic dinucleotides, whereas LRRC8D incorporation into the complex reduces this transport activity.\",\n      \"method\": \"CRISPR knockout screens, genetic rescue with individual subunits, cGAMP transport assays, STING pathway activation readout\",\n      \"journal\": \"Molecular cell\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — functional assay with multiple subunit knockouts and defined transport readout, single lab\",\n      \"pmids\": [\"33171122\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2021,\n      \"finding\": \"LRRC8D co-immunoprecipitates with NADPH oxidase 1 (Nox1) in vascular smooth muscle cells and co-localizes with Nox1 at the plasma membrane and in vesicles. LRRC8D knockdown potentiates NF-κB activation in response to TNFα, suggesting LRRC8D-containing VRACs negatively modulate this inflammatory signaling pathway.\",\n      \"method\": \"Co-immunoprecipitation, siRNA knockdown, NF-κB reporter assay, superoxide production assay\",\n      \"journal\": \"The Journal of physiology\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2-3 / Moderate — co-IP plus functional knockdown with multiple readouts, single lab\",\n      \"pmids\": [\"33932953\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2022,\n      \"finding\": \"LRRC8D localizes to basolateral membranes of proximal tubules in the kidney. Constitutive deletion of Lrrc8d in mice causes proximal tubular injury, increased diuresis, and mild Fanconi-like symptoms, establishing LRRC8A/D channels as required for basolateral exit of organic compounds including metabolites in proximal tubules.\",\n      \"method\": \"Epitope-tagged knock-in mice for localization, constitutive Lrrc8d knockout mice, histology, urine/serum metabolomics\",\n      \"journal\": \"Journal of the American Society of Nephrology : JASN\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — direct localization by knock-in tags plus KO with multiple functional readouts (histology, metabolomics, urine analysis), replicated across subunit deletions\",\n      \"pmids\": [\"35777784\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2022,\n      \"finding\": \"Loss of LRRC8D significantly reduces cis- and carboplatin uptake in BRCA1;p53-deficient mouse mammary tumor cells, resulting in reduced DNA damage and in vivo drug resistance. Lrrc8d deletion in mice does not affect viability or fertility, unlike Lrrc8a deletion.\",\n      \"method\": \"Conditional/constitutive gene knockout in mice, platinum drug uptake measurement, DNA damage assays, in vivo tumor treatment\",\n      \"journal\": \"Cancer research communications\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — in vivo KO with multiple mechanistic readouts, single lab, confirms prior in vitro findings\",\n      \"pmids\": [\"36467895\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2024,\n      \"finding\": \"In the cochlea, LRRC8D is expressed in sensory hair cells and in the stria vascularis (alongside LRRC8A and LRRC8E). Combined ablation of LRRC8D and LRRC8E results in cochlear degeneration and congenital deafness, with severe reduction in the endocochlear potential and loss of Kir4.1 (KCNJ10) expression, suggesting LRRC8D-containing VRACs transport metabolites (e.g., glutathione) needed to maintain inner ear redox potential.\",\n      \"method\": \"Conditional knockout mice, auditory brainstem response, endocochlear potential measurement, immunohistochemistry, Western blot\",\n      \"journal\": \"The Journal of biological chemistry\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — conditional double KO with defined electrophysiological and molecular phenotypes, single lab\",\n      \"pmids\": [\"38838775\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2025,\n      \"finding\": \"NAA60, an N-terminal acetyltransferase localized to the Golgi apparatus, acetylates the N-termini of LRRC8A and LRRC8D. Loss of NAA60 decreases cis- and carboplatin uptake; introduction of positively charged amino acids at the LRRC8A/D N-termini (mimicking absence of acetylation) similarly decreases platinum drug sensitivity, establishing N-terminal acetylation as a post-translational modification required for LRRC8A/D-mediated drug uptake.\",\n      \"method\": \"Co-immunoprecipitation/mass spectrometry, N-terminal acetylation assays, site-directed mutagenesis (charge substitution), drug uptake assays, in vivo tumor models\",\n      \"journal\": \"Communications biology\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 1-2 / Moderate — mutagenesis combined with functional uptake assay and in vivo validation, single lab, multiple orthogonal methods\",\n      \"pmids\": [\"41053424\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2024,\n      \"finding\": \"Cryo-EM structures of a native-like LRRC8A:D VRAC with 4:2 stoichiometry reveal that LRRC8D subunits increase hydrophobicity and widen the selectivity filter compared to LRRC8A homomers, explaining LRRC8D-unique substrate selectivity. Lipids occupy the pore in the closed state (as in LRRC8A:C VRACs). Incorporation of LRRC8D disrupts packing of cytoplasmic LRR domains, increases channel dynamics, and opens lateral fenestrations proposed to be necessary for pore lipid evacuation and channel activation. Electrophysiology confirmed lipid-gating is a general VRAC property.\",\n      \"method\": \"Cryo-EM structure determination (4:2 stoichiometry), electrophysiology\",\n      \"journal\": \"bioRxiv\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 1 / Weak — cryo-EM structure with electrophysiological validation but preprint, single lab\",\n      \"pmids\": [\"bio_10.1101_2024.11.24.625074\"],\n      \"is_preprint\": true\n    },\n    {\n      \"year\": 2025,\n      \"finding\": \"In lateral ventricular neural stem cells, LRRC8D (a core VRAC component) participates in GABA transport/signaling and provides negative feedback to ChAT+ neurons in the ACC-subependymal circuit, with evidence that LRRC8D-regulated chloride and GABA transport modulates neural stem cell proliferation.\",\n      \"method\": \"Immunohistochemistry, in vivo circuit activation, functional behavioral/proliferation readouts\",\n      \"journal\": \"Cells\",\n      \"confidence\": \"Low\",\n      \"confidence_rationale\": \"Tier 3 / Weak — single study, correlative localization and circuit manipulation without direct biochemical mechanistic dissection of LRRC8D's molecular role\",\n      \"pmids\": [\"40136675\"],\n      \"is_preprint\": false\n    }\n  ],\n  \"current_model\": \"LRRC8D is a facultative pore-forming subunit of heteromeric volume-regulated anion channels (VRACs), assembling with the obligatory LRRC8A subunit (and sometimes LRRC8B/C/E) as a hexamer with 4:2 stoichiometry; its incorporation widens the selectivity filter and increases pore hydrophobicity to confer elevated permeability for organic osmolytes (taurine, myo-inositol), platinum-based drugs (cisplatin, carboplatin), and blasticidin S, while inhibiting cGAMP transport; LRRC8D-containing channels are directly inhibited by oxidation of their extracellular loop cysteines, negatively modulate Nox1/NF-κB inflammatory signaling, and are functionally regulated by N-terminal acetylation by NAA60; in vivo, LRRC8A/D channels are required at the basolateral membrane of renal proximal tubules for metabolite export, and combined loss of LRRC8D/E causes cochlear degeneration and deafness.\"\n}\n```","stage2_raw":"{\n  \"mechanistic_narrative\": \"LRRC8D is a facultative pore-forming subunit of heteromeric volume-regulated anion channels (VRACs), which it forms by assembling at the plasma membrane with the obligatory LRRC8A subunit (and LRRC8B/C) [#0, #1]. Cryo-EM of a native-like LRRC8A:D channel with 4:2 stoichiometry shows that incorporation of LRRC8D widens the extracellular selectivity-filter constriction and increases pore hydrophobicity relative to LRRC8A homomers, the structural basis for its distinctive substrate profile [#6, #13]. Functionally, LRRC8D-containing channels confer elevated permeability to uncharged organic osmolytes such as taurine and myo-inositol and to platinum-based drugs (cisplatin, carboplatin), while charged osmolyte release depends on LRRC8A/C/E channels [#1, #2]. This drug-permeation activity is physiologically and clinically consequential: LRRC8D-mediated platinum uptake drives DNA damage in tumor cells, and its loss confers in vivo platinum resistance [#10], and the activity requires N-terminal acetylation of LRRC8A/D by the Golgi acetyltransferase NAA60 [#12]. LRRC8D incorporation conversely suppresses channel transport of the immune second messenger cGAMP [#7]. LRRC8D-containing channels are gated and modulated by their environment — directly inhibited by oxidation of extracellular-loop cysteines [#3], and lipid-occupied in the closed state with LRRC8D-induced lateral fenestrations proposed to permit pore lipid evacuation upon activation [#13]. In vivo, LRRC8A/D channels reside at the basolateral membrane of renal proximal tubules where they are required for export of organic metabolites [#9], and combined loss of LRRC8D and LRRC8E causes cochlear degeneration and congenital deafness with collapse of the endocochlear potential [#11].\",\n  \"teleology\": [\n    {\n      \"year\": 2014,\n      \"claim\": \"Established that LRRC8D has a transport function and physically assembles with other LRRC8 family members, answering whether LRRC8D is a plasma-membrane transport component.\",\n      \"evidence\": \"Genetic loss-of-function screen for blasticidin S import plus co-immunoprecipitation and topology assays in mammalian cells\",\n      \"pmids\": [\"24782309\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Did not define the channel identity or that VRAC is the relevant conductance\", \"Substrate scope limited to blasticidin S at this stage\"]\n    },\n    {\n      \"year\": 2015,\n      \"claim\": \"Identified LRRC8D as a VRAC subunit that shapes pore permeability, answering how LRRC8D contributes to anion-channel function and explaining its role in chemotherapy uptake.\",\n      \"evidence\": \"CRISPR/siRNA knockout, isotopic platinum-drug uptake assays, electrophysiology and genetic rescue\",\n      \"pmids\": [\"26530471\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Structural basis of altered permeability not yet resolved\", \"Stoichiometry of LRRC8D within the hexamer undefined\"]\n    },\n    {\n      \"year\": 2017,\n      \"claim\": \"Showed subunit-specific substrate selectivity, answering which osmolytes LRRC8D-containing channels preferentially conduct versus other heteromers.\",\n      \"evidence\": \"RNAi knockdown with radiotracer flux of taurine, myo-inositol and d-aspartate under hypoosmotic challenge in primary astrocytes\",\n      \"pmids\": [\"28833202\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Selectivity inferred from knockdown rather than reconstituted defined channels\", \"Single cell type\"]\n    },\n    {\n      \"year\": 2017,\n      \"claim\": \"Defined redox regulation of LRRC8D channels, answering how oxidation differentially modulates VRAC heteromers and localizing the oxidation-sensitive sites.\",\n      \"evidence\": \"Electrophysiology of defined fluorescently tagged heteromers with pharmacological oxidation and domain-swap chimeras\",\n      \"pmids\": [\"28841766\", \"33932953\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Specific cysteine residues conferring inhibition not fully enumerated\", \"Physiological oxidant relevant in vivo not established\"]\n    },\n    {\n      \"year\": 2018,\n      \"claim\": \"Mapped the first extracellular loop as essential for channel activity, answering which LRRC8D domain enables functional VRAC assembly and gating.\",\n      \"evidence\": \"Chimeric channel mutagenesis with whole-cell patch clamp under hypotonicity\",\n      \"pmids\": [\"29853476\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Mechanism by which EL1 confers volume sensing unresolved\", \"Single-subunit chimera context\"]\n    },\n    {\n      \"year\": 2018,\n      \"claim\": \"Implicated LRRC8D channels in pancreatic islet signaling, addressing a physiological context for LRRC8D-containing VRAC activity.\",\n      \"evidence\": \"Conditional LRRC8A knockout mice with patch-clamp, insulin secretion and glucose tolerance tests\",\n      \"pmids\": [\"29773801\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"LRRC8D role inferred from expression and partnership, not direct LRRC8D knockout\", \"Permeant signaling molecule not directly identified\"]\n    },\n    {\n      \"year\": 2020,\n      \"claim\": \"Provided a structural explanation for LRRC8D permeability, answering why LRRC8D channels conduct large organic substrates.\",\n      \"evidence\": \"Cryo-EM of the human LRRC8D homo-hexamer with structure-based electrophysiological mutagenesis\",\n      \"pmids\": [\"32415200\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Homo-hexamer is not the physiological heteromeric assembly\", \"Gating transitions not captured\"]\n    },\n    {\n      \"year\": 2020,\n      \"claim\": \"Showed LRRC8D inhibits cGAMP transport, answering how subunit composition selects against immune second-messenger permeation.\",\n      \"evidence\": \"CRISPR knockout screens, subunit rescue, cGAMP transport assays and STING activation readouts\",\n      \"pmids\": [\"33171122\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Structural basis for cGAMP exclusion by LRRC8D not defined\", \"Single-lab functional inference\"]\n    },\n    {\n      \"year\": 2021,\n      \"claim\": \"Linked LRRC8D to inflammatory signaling, answering whether LRRC8D-containing channels modulate Nox1/NF-\\u03baB pathways.\",\n      \"evidence\": \"Co-immunoprecipitation with Nox1, siRNA knockdown, NF-\\u03baB reporter and superoxide assays in vascular smooth muscle cells\",\n      \"pmids\": [\"33932953\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Co-IP without reciprocal validation\", \"Direct versus indirect modulation of NF-\\u03baB unresolved\"]\n    },\n    {\n      \"year\": 2022,\n      \"claim\": \"Established an in vivo physiological role in the kidney, answering whether LRRC8A/D channels export metabolites at the basolateral proximal tubule membrane.\",\n      \"evidence\": \"Epitope-tagged knock-in localization and constitutive Lrrc8d knockout mice with histology and urine/serum metabolomics\",\n      \"pmids\": [\"35777784\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Specific exported metabolites only partially defined\", \"Tissue-specific contribution versus systemic effects not fully separated\"]\n    },\n    {\n      \"year\": 2022,\n      \"claim\": \"Demonstrated in vivo chemotherapy relevance, answering whether LRRC8D loss confers platinum resistance and whether LRRC8D is dispensable for organismal viability.\",\n      \"evidence\": \"Conditional/constitutive knockout mice with platinum uptake, DNA damage assays and tumor treatment\",\n      \"pmids\": [\"36467895\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Mechanism of residual platinum uptake via other subunits not quantified\", \"Clinical translation untested\"]\n    },\n    {\n      \"year\": 2024,\n      \"claim\": \"Defined a cochlear requirement, answering the in vivo consequence of losing LRRC8D-containing channels in the inner ear.\",\n      \"evidence\": \"Conditional LRRC8D/E double knockout mice with auditory brainstem response, endocochlear potential measurement and immunohistochemistry\",\n      \"pmids\": [\"38838775\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Glutathione as the critical transported metabolite is inferential\", \"LRRC8D-specific contribution versus LRRC8E not separated\"]\n    },\n    {\n      \"year\": 2024,\n      \"claim\": \"Resolved the native-like heteromeric architecture and lipid-gating, answering how LRRC8D incorporation alters the pore and channel dynamics in the physiological 4:2 assembly.\",\n      \"evidence\": \"Cryo-EM of LRRC8A:D VRAC at 4:2 stoichiometry with electrophysiology (preprint)\",\n      \"pmids\": [\"bio_10.1101_2024.11.24.625074\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Preprint, not peer-reviewed\", \"Lipid-evacuation gating model awaits independent confirmation\"]\n    },\n    {\n      \"year\": 2025,\n      \"claim\": \"Identified N-terminal acetylation as a required post-translational modification, answering how LRRC8A/D channels are regulated for drug uptake.\",\n      \"evidence\": \"Co-IP/mass spectrometry, acetylation and charge-substitution mutagenesis, drug uptake assays and in vivo tumor models\",\n      \"pmids\": [\"41053424\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Mechanism by which N-terminal charge affects gating/permeation not structurally defined\", \"Single-lab finding\"]\n    },\n    {\n      \"year\": 2025,\n      \"claim\": \"Proposed a neural stem cell signaling role, addressing whether LRRC8D modulates GABA/chloride transport in a defined circuit.\",\n      \"evidence\": \"Immunohistochemistry, in vivo circuit activation and proliferation/behavioral readouts\",\n      \"pmids\": [\"40136675\"],\n      \"confidence\": \"Low\",\n      \"gaps\": [\"Correlative localization without direct biochemical dissection of LRRC8D's molecular role\", \"GABA permeation through LRRC8D channels not directly measured\"]\n    },\n    {\n      \"year\": null,\n      \"claim\": \"How LRRC8D subunit incorporation is regulated to tune channel composition and substrate selectivity across tissues, and the gating transitions linking volume sensing, lipid evacuation and pore opening, remain unresolved.\",\n      \"evidence\": \"\",\n      \"pmids\": [],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"No structure of an open/activated heteromeric channel\", \"Signals controlling tissue-specific subunit assembly unknown\", \"Full physiological substrate repertoire incomplete\"]\n    }\n  ],\n  \"mechanism_profile\": {\n    \"molecular_activity\": [\n      {\"term_id\": \"GO:0005215\", \"supporting_discovery_ids\": [0, 1, 2, 7, 9]},\n      {\"term_id\": \"GO:0005198\", \"supporting_discovery_ids\": [6, 13]}\n    ],\n    \"localization\": [\n      {\"term_id\": \"GO:0005886\", \"supporting_discovery_ids\": [0, 8, 9]},\n      {\"term_id\": \"GO:0031410\", \"supporting_discovery_ids\": [8]}\n    ],\n    \"pathway\": [\n      {\"term_id\": \"R-HSA-382551\", \"supporting_discovery_ids\": [1, 2, 9, 10]},\n      {\"term_id\": \"R-HSA-8953897\", \"supporting_discovery_ids\": [1, 2, 3]}\n    ],\n    \"complexes\": [\n      \"Volume-regulated anion channel (VRAC) LRRC8A:D heteromer\"\n    ],\n    \"partners\": [\n      \"LRRC8A\",\n      \"LRRC8B\",\n      \"LRRC8C\",\n      \"LRRC8E\",\n      \"NOX1\",\n      \"NAA60\"\n    ],\n    \"other_free_text\": []\n  }\n}","audit_flag":null,"evaluation":{"pairwise":"win","faith_supported":7,"faith_total":7,"faith_pct":100.0}}