{"gene":"KIRREL1","run_date":"2026-06-10T02:59:49","timeline":{"discoveries":[{"year":2001,"finding":"NEPH1 knockout mice develop proteinuria and podocyte foot process effacement, establishing that NEPH1 is required for maintaining the structural integrity of the glomerular filtration barrier in podocytes.","method":"Gene trap knockout mice; electron microscopy; proteinuria assay","journal":"Molecular and cellular biology","confidence":"High","confidence_rationale":"Tier 2 / Strong — clean KO with defined cellular phenotype (foot process effacement, proteinuria), replicated in multiple subsequent studies","pmids":["11416156"],"is_preprint":false},{"year":2002,"finding":"NEPH1 directly interacts with the C-terminal domain of podocin via a conserved binding motif; mutation of a centrally located tyrosine residue dramatically reduces NEPH1-podocin affinity. NEPH1 also triggers AP-1 activation requiring Tec family kinases.","method":"Co-immunoprecipitation; mutagenesis; transcriptional reporter assay","journal":"FASEB journal","confidence":"High","confidence_rationale":"Tier 2 / Moderate — reciprocal binding assays with mutagenesis identifying critical tyrosine, two orthogonal methods (pulldown + reporter assay)","pmids":["12424224"],"is_preprint":false},{"year":2003,"finding":"NEPH1 localizes to the glomerular slit diaphragm (confirmed by immunogold EM) and directly interacts with nephrin (via extracellular segments) and ZO-1 (via ZO-1 PDZ domains interacting with the cytoplasmic tail of Neph1). Neph1 forms dimers and multimers. Disrupting the Neph1-nephrin interaction in vivo by injecting subnephritogenic doses of both antibodies causes proteinuria and dramatically reduces ZO-1 protein levels without altering ZO-1 mRNA.","method":"Immunogold electron microscopy; co-immunoprecipitation; in vivo antibody injection; western blot","journal":"The Journal of clinical investigation","confidence":"High","confidence_rationale":"Tier 2 / Strong — subcellular localization by immunogold EM, multiple binding partners confirmed by Co-IP, in vivo functional perturbation, replicated by subsequent studies","pmids":["12865409"],"is_preprint":false},{"year":2003,"finding":"Neph1 localizes exclusively to the lateral margins of podocyte foot processes at the slit diaphragm insertion. Neph1 and Nephrin form cis heterodimers via their cytoplasmic domains, and interactions between extracellular domains of Nephrin with itself and with Neph1 were detected. Neph1 does not engage in homophilic interactions.","method":"Immunogold electron microscopy; co-immunoprecipitation; cell fractionation (detergent-resistant membrane)","journal":"The Journal of biological chemistry","confidence":"High","confidence_rationale":"Tier 2 / Strong — immunogold localization combined with biochemical Co-IP; negative result (no Neph1 homophilic interaction) explicitly tested; corroborated by multiple subsequent papers","pmids":["12646566"],"is_preprint":false},{"year":2003,"finding":"The extracellular Ig domains of NEPH1 mediate both homodimerization and heterodimerization with nephrin; as few as two Ig domains are sufficient for binding. These interactions are strictly dependent on post-translational glycosylation (bacterially expressed unglycosylated protein fails to bind).","method":"Co-immunoprecipitation; Ig-fusion pulldown; truncation analysis; glycosylation-deficient expression","journal":"Journal of the American Society of Nephrology","confidence":"High","confidence_rationale":"Tier 2 / Moderate — multiple truncation and glycosylation mutants tested in orthogonal pulldown/Co-IP assays in a single study","pmids":["12660326"],"is_preprint":false},{"year":2006,"finding":"Neph1 is expressed in the developing mammalian brain and localizes to dendritic shafts and synaptic sites. Neph1 interacts with the PDZ domain of the synaptic scaffolding protein CASK via its cytoplasmic tail, suggesting a role in synaptogenesis.","method":"In situ hybridization; immunohistochemistry; immunogold EM; co-immunoprecipitation","journal":"The Journal of comparative neurology","confidence":"Medium","confidence_rationale":"Tier 3 / Moderate — subcellular localization by immunogold EM with biochemical Co-IP for CASK interaction, single lab","pmids":["16874800"],"is_preprint":false},{"year":2007,"finding":"Upon engagement, Neph1 is phosphorylated on specific tyrosine residues by the Src family kinase Fyn, which recruits the adaptor Grb2. This Grb2 recruitment is necessary for Neph1-induced actin polymerization at the plasma membrane. Neph1 and Nephrin directly interact and cooperate by juxtaposing Grb2 and Nck1/2 at the membrane to augment actin polymerization efficiency.","method":"Phosphorylation assay; co-immunoprecipitation; dominant-negative and siRNA knockdown; actin polymerization assay; mutagenesis","journal":"Molecular and cellular biology","confidence":"High","confidence_rationale":"Tier 2 / Moderate — kinase-substrate relationship established in cells, multiple orthogonal methods (Co-IP, mutagenesis, functional actin assay), single lab","pmids":["17923684"],"is_preprint":false},{"year":2008,"finding":"Fyn phosphorylates the cytoplasmic domain of Neph1 at multiple tyrosines (including Y637 and Y638) both in vitro and in intact cells. Phosphorylated Neph1 specifically binds adaptor protein Grb2 and tyrosine kinase Csk in a phosphorylation-dependent manner (from glomerular lysates). Neph1 attenuates Fyn-elicited ERK activation, requiring intact Grb2-SH2 binding motif. Phosphorylation of Y637 is upregulated in vivo in podocyte injury models.","method":"In vitro kinase assay; peptide mass fingerprinting; site-directed mutagenesis; GST pulldown from rat glomerular lysates; co-immunoprecipitation","journal":"The Journal of biological chemistry","confidence":"High","confidence_rationale":"Tier 1 / Moderate — in vitro kinase assay with mutagenesis identifying specific phosphorylation sites, pulldown from native tissue, functional ERK signaling readout","pmids":["18258597"],"is_preprint":false},{"year":2008,"finding":"Renal ischemia induces rapid loss of interaction between slit diaphragm proteins Neph1 and ZO-1, with redistribution of both proteins from cell membrane to cytoplasm. Recovery from ATP-depletion injury restores Neph1 tyrosine phosphorylation (mediated by Fyn), which is required for re-establishing Neph1-ZO-1 binding and their co-localization at the membrane. Fyn-mediated Neph1 phosphorylation significantly increases Neph1-ZO-1 binding.","method":"In vivo rat ischemia model; co-immunoprecipitation; cell culture ATP depletion; immunofluorescence; Fyn kinase assay","journal":"The Journal of biological chemistry","confidence":"High","confidence_rationale":"Tier 2 / Moderate — in vivo and in vitro models, kinase identified (Fyn), interaction dynamics measured by Co-IP under injury and recovery conditions","pmids":["18922801"],"is_preprint":false},{"year":2009,"finding":"Neph1 interacts with large-conductance Ca2+-activated K+ (BK/Slo1) channels via all three extreme C-terminal Slo1 splice variants, as shown by reciprocal Co-IP from endogenous podocyte and neuron proteins and GST pulldown. Co-expression of Neph1 with Slo1 suppresses steady-state Slo1 surface expression in HEK293T cells. Conversely, siRNA knockdown of Neph1 in ciliary ganglion neurons increases surface Slo1 and BK current, while knockdown in podocytes decreases Slo1 surface expression and BK current.","method":"Reciprocal co-immunoprecipitation from endogenous proteins; GST pulldown; cell surface biotinylation; whole-cell electrophysiology; siRNA knockdown; confocal microscopy","journal":"American journal of physiology. Cell physiology","confidence":"High","confidence_rationale":"Tier 2 / Moderate — reciprocal Co-IP from native tissue plus functional electrophysiology and loss-of-function in two cell types; multiple orthogonal methods","pmids":["19794150"],"is_preprint":false},{"year":2011,"finding":"Neph1 and Nephrin can trans-interact across cells to induce cell adhesion (demonstrated in L fibroblasts lacking endogenous adhesion). Neph1 alone can induce cell adhesion. Trans-interaction of nephrin with Neph1 (or Neph3) is associated with down-regulation of nephrin tyrosine phosphorylation.","method":"L fibroblast cell adhesion assay; co-immunoprecipitation; phosphorylation analysis","journal":"The Biochemical journal","confidence":"Medium","confidence_rationale":"Tier 2 / Weak — clean cell adhesion assay with defined readout, but single lab with limited mechanistic follow-up on phosphorylation","pmids":["21306299"],"is_preprint":false},{"year":2011,"finding":"Motor protein Myo1c directly interacts with Neph1 in an actin-dependent manner and is required for targeting Neph1 to the podocyte cell membrane. Expression of dominant-negative Myo1c or siRNA depletion of Myo1c significantly reduces membrane localization of Neph1 and nephrin. Myo1c knockdown also impairs cell migration and tight junction formation.","method":"Co-immunoprecipitation in vivo and in vitro; dominant-negative expression; siRNA knockdown; cell surface fractionation; wound assay; transepithelial electric resistance","journal":"Molecular and cellular biology","confidence":"High","confidence_rationale":"Tier 2 / Moderate — direct interaction confirmed by Co-IP, loss-of-function via two approaches (DN and siRNA) with defined membrane localization phenotype, multiple functional readouts","pmids":["21402783"],"is_preprint":false},{"year":2012,"finding":"The solution structure of the Neph1 cytoplasmic domain (Neph1-CD) was determined by SWAXS (radius of gyration 21.3 Å, max dimension 70 Å). A structural model of the Neph1-CD·ZO-1-PDZ1 complex was constructed; beyond the C-terminal Thr-His-Val motif, residues Lys-761 and Tyr-762 in Neph1 are critical for ZO-1 binding (individual alanine mutations abolished binding).","method":"Small/wide angle X-ray scattering (SWAXS); circular dichroism; in vitro and in vivo pulldown with mutagenesis","journal":"The Journal of biological chemistry","confidence":"High","confidence_rationale":"Tier 1 / Moderate — structural determination combined with site-directed mutagenesis and functional binding validation","pmids":["22262837"],"is_preprint":false},{"year":2014,"finding":"Inhibiting Neph1 signaling by transducing its cytoplasmic domain (Neph1CD) into podocytes reduces puromycin aminonucleoside (PAN)-induced Neph1 phosphorylation, retains Neph1 in lipid raft fractions, maintains Neph1 at the membrane, and protects against cytoskeletal damage and albumin leakage. Maintaining high membrane levels of Neph1 via chimeric overexpression increases podocyte resistance to PAN injury.","method":"Protein transduction; lipid raft fractionation; immunofluorescence; transepithelial permeability assay; zebrafish in vivo injury model","journal":"The Journal of biological chemistry","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — multiple orthogonal methods in vitro plus in vivo zebrafish model; single lab","pmids":["24554715"],"is_preprint":false},{"year":2016,"finding":"The full-length Myo1c adopts an extended S-shaped conformation in solution; Neph1 binds to the C-terminal tail of Myo1c without inducing significant conformational change. A critical residue in Neph1 required for Myo1c binding was identified; point mutation abolished the interaction in vitro and in live cells. FRAP analysis confirmed Myo1c mediates intracellular vesicular movement and membrane turnover of Neph1.","method":"Small angle X-ray scattering (SAXS); mutagenesis; in vitro binding assay; live-cell imaging; fluorescence recovery after photobleaching (FRAP)","journal":"Molecular and cellular biology","confidence":"High","confidence_rationale":"Tier 1 / Moderate — structural SAXS data combined with mutagenesis and FRAP live-cell imaging in a single study","pmids":["27044863"],"is_preprint":false},{"year":2017,"finding":"The Neph1-ZO-1 protein-protein interaction interface contains a druggable pocket involving both proteins. The small molecule isodesmosine (ISD) enhances Neph1 cytoplasmic domain–ZO-1 binding in vitro and in vivo, and ISD-treated podocytes are resistant to injury-induced transepithelial permeability loss. Mouse and zebrafish experiments confirm ISD protects from injury-induced renal damage.","method":"Computational structural screening; biochemical binding assay; in vitro podocyte permeability assay; mouse and zebrafish injury models","journal":"Scientific reports","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — multiple in vitro and in vivo models but computational component and single lab","pmids":["28935902"],"is_preprint":false},{"year":2017,"finding":"CD80 (B7-1) interacts with Neph1 via their extracellular domains, as established by pulldown assay in HEK293 cells co-transfected with both proteins. CD80 overexpression in podocytes co-localizes with Neph1 and causes actin derangement.","method":"Co-immunoprecipitation/pulldown in HEK293 cells; immunofluorescence co-localization in podocytes","journal":"Clinical and experimental nephrology","confidence":"Low","confidence_rationale":"Tier 3 / Weak — single pulldown in overexpression system, limited mechanistic follow-up, single lab","pmids":["29022109"],"is_preprint":false},{"year":2019,"finding":"Homozygous mutations in KIRREL1 in patients with steroid-resistant nephrotic syndrome result in mutant KIRREL1 proteins that fail to localize to the podocyte cell membrane, indicating defective membrane trafficking.","method":"Patient genetics; immunofluorescence localization of mutant vs wild-type KIRREL1 in podocytes","journal":"Kidney international","confidence":"Medium","confidence_rationale":"Tier 2 / Weak — direct localization experiment with functional consequence (nephrotic syndrome), but single paper and limited mechanistic detail on trafficking mechanism","pmids":["31472902"],"is_preprint":false},{"year":2021,"finding":"NEPHRIN and NEPH1 are novel receptor proteins for hepatocyte growth factor (HGF), binding HGF directly through their extracellular domains (confirmed by surface plasmon resonance with purified recombinant proteins; NEPHRIN Kd ~20-fold higher affinity than NEPH1). HGF binding leads to phosphorylation of NEPHRIN and NEPH1 independently of the MET receptor. SHP-2 (PTPN11) mediates dephosphorylation of these proteins. HGF-induced phosphorylation promotes podocyte repair.","method":"Surface plasmon resonance with purified baculovirus-expressed recombinant proteins; molecular modeling; cell culture and Drosophila nephrocyte functional assays","journal":"The Journal of biological chemistry","confidence":"High","confidence_rationale":"Tier 1 / Moderate — in vitro binding reconstitution with purified proteins (SPR), site mapping, and functional in vitro and in vivo validation","pmids":["34391780"],"is_preprint":false},{"year":2022,"finding":"KIRREL1 physically interacts with SAV1 and recruits SAV1 to cell-cell contact sites, thereby activating the Hippo pathway. Knockout of KIRREL1 increases YAP activity in neighboring cells. During liver regeneration in mice, KIRREL1 knockout enhances hepatic YAP activity, hepatocyte reprogramming, and biliary epithelial cell proliferation.","method":"Co-immunoprecipitation; knockout mouse model; in vivo liver regeneration assay; YAP activity reporter","journal":"Nature communications","confidence":"High","confidence_rationale":"Tier 2 / Strong — direct physical interaction (Co-IP), in vivo KO phenotype with specific pathway readouts (YAP activity), replicated by independent lab in same year","pmids":["35177623"],"is_preprint":false},{"year":2022,"finding":"KIRREL1 interacts with both SAV1 and LATS1/2, promoting LATS1/2 activation by MST1/2 (Hippo kinases), thereby suppressing YAP/TAZ oncoproteins. YAP/TAZ in turn directly induce KIRREL1 expression in a TEAD1-4-dependent manner, constituting a negative feedback loop. Transgenic expression of KIRREL1 blocks tumorigenesis in a mouse intrahepatic cholangiocarcinoma model.","method":"Co-immunoprecipitation; LATS kinase activity assay; CRISPR screen; TEAD reporter assay; transgenic mouse tumor model","journal":"Cell reports","confidence":"High","confidence_rationale":"Tier 2 / Strong — kinase activity assay, multiple interactors confirmed by Co-IP, in vivo tumor model, replicated by independent lab in same year","pmids":["36044856"],"is_preprint":false},{"year":2022,"finding":"KIRREL binds directly to SAV1 to activate the Hippo tumor suppressor pathway. KIRREL loss markedly promotes tumor growth in vivo. This was discovered through an in vivo CRISPR screen of cell surface proteins and a separate CRISPR screen using a Hippo pathway reporter.","method":"In vivo CRISPR proliferation screen; Hippo pathway reporter CRISPR screen; direct binding assay","journal":"Proceedings of the National Academy of Sciences","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — CRISPR functional screens plus direct binding assay; partially overlapping findings with PMID 35177623 and 36044856","pmids":["35704761"],"is_preprint":false},{"year":2023,"finding":"KIRREL1 knockdown inhibits proliferation and angiogenesis of gastric cancer cells; overexpression promotes them. These effects are mediated through the PI3K/AKT/mTOR pathway: KIRREL1 silencing represses P-PI3K, P-AKT, P-mTOR, HIF-1α, and VEGF, while overexpression stimulates them. The AKT agonist IGF-1 and inhibitor LY294002 reverse these effects.","method":"Lentiviral knockdown/overexpression; western blot; pharmacological pathway inhibition/activation","journal":"Journal of cellular and molecular medicine","confidence":"Medium","confidence_rationale":"Tier 3 / Moderate — multiple cell-based assays with pharmacological validation of pathway; single lab, no direct binding to PI3K components demonstrated","pmids":["37909722"],"is_preprint":false},{"year":2024,"finding":"PRRXL1 homeodomain transcription factor directly regulates Neph1 transcription by binding to four intronic regions of the Neph1 gene (identified by ChIP), preventing premature Neph1 expression in dorsal horn superficial laminae at E14.5. Loss-of-function experiments show Neph1 is required for neurite branching, especially at distal neurites.","method":"Chromatin immunoprecipitation (ChIP); in situ hybridization; loss-of-function (Neph1 KO or knockdown); neurite morphometry","journal":"Neural development","confidence":"Medium","confidence_rationale":"Tier 2 / Weak — ChIP establishes direct transcriptional regulation, loss-of-function with morphological readout; single lab","pmids":["39049046"],"is_preprint":false},{"year":2025,"finding":"Cryo-electron tomography of human kidney tissue resolves the slit diaphragm as a fishnet-like lattice with crisscrossing strands at ~90°. An atomic model based on the Nephrin-Neph1 heterodimer fits the structure, with ~9 nm spacing in humans (vs 12.3 nm in mice and 15 nm in Drosophila), indicating the SD is a conserved heterodimeric fishnet assembly.","method":"Cryo-electron tomography of native human and mouse kidney tissue; atomic model fitting","journal":"bioRxiv","confidence":"Medium","confidence_rationale":"Tier 1 / Weak — high-resolution structural method on native tissue, preprint not yet peer-reviewed, single study","pmids":["bio_10.1101_2025.09.24.678239"],"is_preprint":true}],"current_model":"KIRREL1/NEPH1 is a transmembrane immunoglobulin superfamily protein that localizes to the glomerular slit diaphragm (forming part of a fishnet-like heterodimeric lattice with Nephrin) and to neuronal synapses, where it forms cis- and trans-heterodimers with Nephrin via both extracellular Ig domains and cytoplasmic interactions, is phosphorylated on specific tyrosines by the Src-family kinase Fyn, recruits Grb2 (via pY637/Y638) to drive actin polymerization through cooperation with Nck, binds ZO-1 via its cytoplasmic tail (stabilized by Fyn-mediated phosphorylation), is transported to the membrane by the motor protein Myo1c, acts as a receptor for HGF, and functions as an upstream positive regulator of the Hippo pathway by interacting with both SAV1 and LATS1/2 at cell-cell contacts to promote LATS1/2 activation and YAP/TAZ suppression."},"narrative":{"mechanistic_narrative":"KIRREL1/NEPH1 is a transmembrane immunoglobulin-superfamily adhesion receptor that is required to maintain the structural integrity of the glomerular filtration barrier in podocytes [PMID:11416156]. At the slit diaphragm it localizes to the lateral margins of podocyte foot processes [PMID:12865409, PMID:12646566] and assembles a heterodimeric, fishnet-like lattice with Nephrin: its extracellular Ig domains mediate glycosylation-dependent cis- and trans-heterodimerization with Nephrin (it does not engage in homophilic interactions in foot processes) [PMID:12646566, PMID:12660326, PMID:bio_10.1101_2025.09.24.678239], while its cytoplasmic tail couples to the slit-diaphragm scaffold through a C-terminal motif and residues Lys-761/Tyr-762 that bind the ZO-1 PDZ domain [PMID:12865409, PMID:22262837] and through podocin [PMID:12424224]. Upon engagement KIRREL1 is phosphorylated on cytoplasmic tyrosines (including Y637/Y638) by the Src-family kinase Fyn, which recruits Grb2 to drive actin polymerization in cooperation with Nephrin-bound Nck and also recruits Csk to attenuate ERK signaling [PMID:17923684, PMID:18258597]; Fyn-dependent phosphorylation likewise stabilizes the KIRREL1–ZO-1 interaction and membrane localization, a coupling that is acutely lost during ischemic and ATP-depletion injury and restored on recovery [PMID:18922801]. Membrane delivery and turnover of KIRREL1 depend on direct, actin-dependent binding to the motor protein Myo1c [PMID:21402783, PMID:27044863], and failure of mutant KIRREL1 to reach the podocyte membrane causes steroid-resistant nephrotic syndrome in patients with homozygous KIRREL1 mutations [PMID:31472902]. Beyond the kidney, KIRREL1 acts as a direct receptor for HGF, binding through its extracellular domain and undergoing MET-independent phosphorylation that promotes podocyte repair [PMID:34391780], and it functions as an upstream positive regulator of the Hippo pathway by recruiting SAV1 and LATS1/2 to cell–cell contacts to promote LATS1/2 activation and YAP/TAZ suppression within a TEAD-dependent negative feedback loop, restraining tumorigenesis in liver and biliary models [PMID:35177623, PMID:36044856, PMID:35704761]. KIRREL1/NEPH1 is also expressed at neuronal synapses and dendrites, where it binds CASK and is required for neurite branching [PMID:16874800, PMID:39049046].","teleology":[{"year":2001,"claim":"Established that NEPH1 is physiologically essential for the glomerular filtration barrier, framing it as a slit-diaphragm component rather than an incidental podocyte protein.","evidence":"Gene-trap knockout mice with EM and proteinuria readouts","pmids":["11416156"],"confidence":"High","gaps":["Did not define molecular partners or the biochemical basis of the phenotype","No mechanism for how loss leads to foot process effacement"]},{"year":2003,"claim":"Resolved where and with what NEPH1 acts at the slit diaphragm, showing it localizes to the slit-diaphragm insertion and forms a Nephrin-linked, ZO-1-coupled adhesion complex whose integrity sustains the barrier.","evidence":"Immunogold EM, Co-IP, in vivo antibody perturbation, cell fractionation","pmids":["12865409","12646566","12660326"],"confidence":"High","gaps":["Did not establish the intracellular signaling consequences of complex assembly","Trans- vs cis-orientation of Nephrin–NEPH1 not fully resolved at this stage"]},{"year":2002,"claim":"Identified podocin as a cytoplasmic NEPH1 partner and a tyrosine-dependent binding motif, linking NEPH1 to other slit-diaphragm scaffolding and to AP-1 transcriptional output.","evidence":"Co-IP, mutagenesis, transcriptional reporter assay","pmids":["12424224"],"confidence":"High","gaps":["Functional significance of AP-1 activation in podocytes unresolved","Tec-family kinase identity not pinned down"]},{"year":2008,"claim":"Defined the kinase-substrate logic of NEPH1 signaling, showing Fyn phosphorylates specific cytoplasmic tyrosines (Y637/Y638) to recruit Grb2 and Csk and drive actin polymerization, connecting adhesion to cytoskeletal remodeling.","evidence":"In vitro kinase assay, mass fingerprinting, mutagenesis, glomerular pulldown, actin polymerization assay","pmids":["17923684","18258597"],"confidence":"High","gaps":["Quantitative contribution of NEPH1 vs Nephrin to actin output not separated","In vivo relevance of ERK attenuation unclear"]},{"year":2008,"claim":"Showed the NEPH1–ZO-1 interaction is dynamically regulated by Fyn phosphorylation and disrupted by injury, providing a molecular mechanism for slit-diaphragm disassembly and recovery during ischemia.","evidence":"In vivo rat ischemia model, ATP depletion, Co-IP, immunofluorescence, Fyn kinase assay","pmids":["18922801"],"confidence":"High","gaps":["Upstream signal triggering Fyn reactivation during recovery not identified"]},{"year":2011,"claim":"Identified Myo1c as the motor delivering NEPH1 (and Nephrin) to the membrane, explaining how the slit-diaphragm complex is trafficked and assembled.","evidence":"Co-IP, dominant-negative and siRNA, surface fractionation, wound and TEER assays","pmids":["21402783"],"confidence":"High","gaps":["Regulation of Myo1c-dependent delivery not defined","Cargo selectivity mechanism unknown"]},{"year":2012,"claim":"Provided structural definition of the NEPH1 cytoplasmic domain and its ZO-1 interface, identifying Lys-761/Tyr-762 as critical contacts and rendering the interaction structurally tractable.","evidence":"SWAXS, circular dichroism, mutagenesis pulldowns","pmids":["22262837"],"confidence":"High","gaps":["No high-resolution atomic structure of the full complex","Phospho-regulation not captured structurally"]},{"year":2016,"claim":"Structurally characterized the NEPH1–Myo1c interaction and demonstrated by FRAP that Myo1c mediates vesicular movement and membrane turnover of NEPH1.","evidence":"SAXS, mutagenesis, in vitro binding, live-cell FRAP","pmids":["27044863"],"confidence":"High","gaps":["Coupling of motor activity to specific trafficking routes unresolved"]},{"year":2014,"claim":"Demonstrated that maintaining membrane NEPH1 and its signaling protects podocytes from injury, establishing NEPH1 stabilization as a candidate protective strategy.","evidence":"Protein transduction, lipid raft fractionation, permeability assays, zebrafish injury model","pmids":["24554715"],"confidence":"Medium","gaps":["Single lab","Mechanism by which Neph1CD retains the protein in rafts not fully defined"]},{"year":2017,"claim":"Showed the NEPH1–ZO-1 interface is druggable, with the small molecule isodesmosine enhancing binding and conferring injury resistance in vitro and in vivo.","evidence":"Computational screening, binding assay, podocyte permeability assay, mouse and zebrafish injury models","pmids":["28935902"],"confidence":"Medium","gaps":["Single lab","Direct target engagement specificity of ISD not orthogonally confirmed"]},{"year":2017,"claim":"Reported CD80 (B7-1) as an extracellular NEPH1 partner associated with actin derangement, hinting at an immune-injury link.","evidence":"Pulldown in HEK293 overexpression, immunofluorescence co-localization","pmids":["29022109"],"confidence":"Low","gaps":["Single pulldown in overexpression system without reciprocal validation","No endogenous interaction or in vivo evidence"]},{"year":2019,"claim":"Linked KIRREL1 directly to human disease, showing homozygous mutations cause steroid-resistant nephrotic syndrome through defective membrane trafficking of mutant protein.","evidence":"Patient genetics, immunofluorescence localization of mutant vs WT KIRREL1","pmids":["31472902"],"confidence":"Medium","gaps":["Single study","Trafficking step that fails not mechanistically defined"]},{"year":2009,"claim":"Extended NEPH1 function beyond scaffolding by showing it regulates BK/Slo1 channel surface expression and current in podocytes and neurons.","evidence":"Reciprocal endogenous Co-IP, GST pulldown, surface biotinylation, electrophysiology, siRNA","pmids":["19794150"],"confidence":"High","gaps":["Directionality of regulation differs between cell types and is unexplained","Structural basis of Slo1 binding not defined"]},{"year":2021,"claim":"Revealed NEPH1 (and Nephrin) as a direct HGF receptor undergoing MET-independent, SHP-2-reversible phosphorylation that promotes podocyte repair, recasting NEPH1 as a ligand-responsive signaling receptor.","evidence":"SPR with purified recombinant proteins, modeling, cell and Drosophila nephrocyte assays","pmids":["34391780"],"confidence":"High","gaps":["Downstream effectors of HGF-induced NEPH1 phosphorylation not fully mapped","Relative in vivo contribution vs MET signaling unclear"]},{"year":2022,"claim":"Defined a distinct role for KIRREL1 as an upstream positive regulator of the Hippo pathway, recruiting SAV1 and LATS1/2 to cell-cell contacts to activate LATS and suppress YAP/TAZ within a TEAD-dependent feedback loop, with tumor-suppressive consequences.","evidence":"Co-IP, KO and transgenic mouse models, CRISPR screens, LATS kinase and TEAD reporter assays, liver regeneration and cholangiocarcinoma models","pmids":["35177623","36044856","35704761"],"confidence":"High","gaps":["How adhesion-based KIRREL1 engagement is transduced to LATS activation mechanistically unresolved","Connection between slit-diaphragm and Hippo roles not reconciled"]},{"year":2023,"claim":"Reported a pro-tumorigenic, PI3K/AKT/mTOR-linked role for KIRREL1 in gastric cancer, contrasting with its Hippo tumor-suppressive function and indicating context-dependent outputs.","evidence":"Lentiviral knockdown/overexpression, western blot, pharmacological pathway modulation","pmids":["37909722"],"confidence":"Medium","gaps":["No direct binding to PI3K components shown","Single lab","Reconciliation with Hippo tumor-suppressor role absent"]},{"year":2024,"claim":"Established transcriptional control of Neph1 by the homeodomain factor PRRXL1 and a functional requirement for Neph1 in neurite branching, deepening its neuronal role.","evidence":"ChIP, in situ hybridization, loss-of-function, neurite morphometry","pmids":["39049046"],"confidence":"Medium","gaps":["Single lab","Molecular mechanism of Neph1 in neurite branching unresolved"]},{"year":2025,"claim":"Resolved the native slit diaphragm as a conserved Nephrin–NEPH1 heterodimeric fishnet lattice with species-specific spacing, providing the supramolecular architecture underlying the barrier.","evidence":"Cryo-electron tomography of native human and mouse kidney tissue with atomic model fitting (preprint)","pmids":["bio_10.1101_2025.09.24.678239"],"confidence":"Medium","gaps":["Preprint, not peer-reviewed","Single study","Dynamics of lattice remodeling not captured"]},{"year":null,"claim":"How KIRREL1's adhesion/scaffolding function at cell-cell contacts is mechanistically transduced into Hippo (LATS) activation, and how its opposing tumor-suppressive versus pro-proliferative outputs are determined by cellular context, remain unresolved.","evidence":"","pmids":[],"confidence":"Medium","gaps":["No mechanism linking extracellular engagement to LATS activation","Context-dependent tumor roles not reconciled","Integration of kidney, neuronal, and Hippo functions unknown"]}],"mechanism_profile":{"molecular_activity":[{"term_id":"GO:0098631","term_label":"cell adhesion mediator activity","supporting_discovery_ids":[3,4,10,24]},{"term_id":"GO:0001618","term_label":"virus receptor activity","supporting_discovery_ids":[18]},{"term_id":"GO:0060089","term_label":"molecular transducer activity","supporting_discovery_ids":[6,7,18]},{"term_id":"GO:0060090","term_label":"molecular adaptor activity","supporting_discovery_ids":[2,6,19,20]}],"localization":[{"term_id":"GO:0005886","term_label":"plasma membrane","supporting_discovery_ids":[2,3,11,17]},{"term_id":"GO:0005829","term_label":"cytosol","supporting_discovery_ids":[8]}],"pathway":[{"term_id":"R-HSA-162582","term_label":"Signal Transduction","supporting_discovery_ids":[6,7,19,20,18]},{"term_id":"R-HSA-1474244","term_label":"Extracellular matrix organization","supporting_discovery_ids":[2,3,24]},{"term_id":"R-HSA-1643685","term_label":"Disease","supporting_discovery_ids":[17,20]}],"complexes":["slit diaphragm (Nephrin-NEPH1 fishnet lattice)"],"partners":["NPHS1","NPHS2","TJP1","FYN","GRB2","MYO1C","SAV1","LATS1"],"other_free_text":[]}},"prefetch_data":{"uniprot":{"accession":"Q96J84","full_name":"Kin of IRRE-like protein 1","aliases":["Kin of irregular chiasm-like protein 1","Nephrin-like protein 1"],"length_aa":757,"mass_kda":83.5,"function":"Required for proper function of the glomerular filtration barrier. It is involved in the maintenance of a stable podocyte architecture with interdigitating foot processes connected by specialized cell-cell junctions, known as the slit diaphragm (PubMed:31472902). It is a signaling protein that needs the presence of TEC kinases to fully trans-activate the transcription factor AP-1 (By similarity)","subcellular_location":"Cell membrane","url":"https://www.uniprot.org/uniprotkb/Q96J84/entry"},"depmap":{"release":"DepMap","has_data":true,"is_common_essential":false,"resolved_as":"","url":"https://depmap.org/portal/gene/KIRREL1","classification":"Not Classified","n_dependent_lines":2,"n_total_lines":1208,"dependency_fraction":0.0016556291390728477},"opencell":{"profiled":false,"resolved_as":"","ensg_id":"","cell_line_id":"","localizations":[],"interactors":[],"url":"https://opencell.sf.czbiohub.org/search/KIRREL1","total_profiled":1310},"omim":[{"mim_id":"619201","title":"NEPHROTIC SYNDROME, TYPE 23; NPHS23","url":"https://www.omim.org/entry/619201"},{"mim_id":"607428","title":"KIRRE-LIKE NEPHRIN FAMILY ADHESION MOLECULE 1; KIRREL1","url":"https://www.omim.org/entry/607428"},{"mim_id":"256300","title":"NEPHROTIC SYNDROME, TYPE 1; NPHS1","url":"https://www.omim.org/entry/256300"}],"hpa":{"profiled":true,"resolved_as":"","reliability":"","locations":[],"tissue_specificity":"Low tissue specificity","tissue_distribution":"Detected in many","driving_tissues":[],"url":"https://www.proteinatlas.org/search/KIRREL1"},"hgnc":{"alias_symbol":["NEPH1"],"prev_symbol":["KIRREL"]},"alphafold":{"accession":"Q96J84","domains":[{"cath_id":"2.60.40.10","chopping":"18-118","consensus_level":"high","plddt":91.5239,"start":18,"end":118},{"cath_id":"2.60.40.10","chopping":"120-302","consensus_level":"medium","plddt":95.0719,"start":120,"end":302},{"cath_id":"2.60.40.10","chopping":"307-391","consensus_level":"high","plddt":94.8241,"start":307,"end":391},{"cath_id":"2.60.40.10","chopping":"395-490","consensus_level":"high","plddt":91.8434,"start":395,"end":490}],"viewer_url":"https://alphafold.ebi.ac.uk/entry/Q96J84","model_url":"https://alphafold.ebi.ac.uk/files/AF-Q96J84-F1-model_v6.cif","pae_url":"https://alphafold.ebi.ac.uk/files/AF-Q96J84-F1-predicted_aligned_error_v6.png","plddt_mean":75.88},"mouse_models":{"mgi_url":"https://www.informatics.jax.org/marker/summary?nomen=KIRREL1","jax_strain_url":"https://www.jax.org/strain/search?query=KIRREL1"},"sequence":{"accession":"Q96J84","fasta_url":"https://rest.uniprot.org/uniprotkb/Q96J84.fasta","uniprot_url":"https://www.uniprot.org/uniprotkb/Q96J84/entry","alphafold_viewer_url":"https://alphafold.ebi.ac.uk/entry/Q96J84"}},"corpus_meta":[{"pmid":"11416156","id":"PMC_11416156","title":"Proteinuria and perinatal lethality in mice lacking NEPH1, a novel protein with homology to NEPHRIN.","date":"2001","source":"Molecular and cellular biology","url":"https://pubmed.ncbi.nlm.nih.gov/11416156","citation_count":337,"is_preprint":false},{"pmid":"12424224","id":"PMC_12424224","title":"NEPH1 defines a novel family of podocin interacting proteins.","date":"2002","source":"FASEB journal : official publication of the Federation of American Societies for Experimental Biology","url":"https://pubmed.ncbi.nlm.nih.gov/12424224","citation_count":179,"is_preprint":false},{"pmid":"12865409","id":"PMC_12865409","title":"Neph1 and nephrin interaction in the slit diaphragm is an important determinant of glomerular permeability.","date":"2003","source":"The Journal of clinical investigation","url":"https://pubmed.ncbi.nlm.nih.gov/12865409","citation_count":172,"is_preprint":false},{"pmid":"12646566","id":"PMC_12646566","title":"Nephrin and Neph1 co-localize at the podocyte foot process intercellular 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NEPH1 also triggers AP-1 activation requiring Tec family kinases.\",\n      \"method\": \"Co-immunoprecipitation; mutagenesis; transcriptional reporter assay\",\n      \"journal\": \"FASEB journal\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — reciprocal binding assays with mutagenesis identifying critical tyrosine, two orthogonal methods (pulldown + reporter assay)\",\n      \"pmids\": [\"12424224\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2003,\n      \"finding\": \"NEPH1 localizes to the glomerular slit diaphragm (confirmed by immunogold EM) and directly interacts with nephrin (via extracellular segments) and ZO-1 (via ZO-1 PDZ domains interacting with the cytoplasmic tail of Neph1). Neph1 forms dimers and multimers. Disrupting the Neph1-nephrin interaction in vivo by injecting subnephritogenic doses of both antibodies causes proteinuria and dramatically reduces ZO-1 protein levels without altering ZO-1 mRNA.\",\n      \"method\": \"Immunogold electron microscopy; co-immunoprecipitation; in vivo antibody injection; western blot\",\n      \"journal\": \"The Journal of clinical investigation\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — subcellular localization by immunogold EM, multiple binding partners confirmed by Co-IP, in vivo functional perturbation, replicated by subsequent studies\",\n      \"pmids\": [\"12865409\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2003,\n      \"finding\": \"Neph1 localizes exclusively to the lateral margins of podocyte foot processes at the slit diaphragm insertion. Neph1 and Nephrin form cis heterodimers via their cytoplasmic domains, and interactions between extracellular domains of Nephrin with itself and with Neph1 were detected. Neph1 does not engage in homophilic interactions.\",\n      \"method\": \"Immunogold electron microscopy; co-immunoprecipitation; cell fractionation (detergent-resistant membrane)\",\n      \"journal\": \"The Journal of biological chemistry\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — immunogold localization combined with biochemical Co-IP; negative result (no Neph1 homophilic interaction) explicitly tested; corroborated by multiple subsequent papers\",\n      \"pmids\": [\"12646566\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2003,\n      \"finding\": \"The extracellular Ig domains of NEPH1 mediate both homodimerization and heterodimerization with nephrin; as few as two Ig domains are sufficient for binding. These interactions are strictly dependent on post-translational glycosylation (bacterially expressed unglycosylated protein fails to bind).\",\n      \"method\": \"Co-immunoprecipitation; Ig-fusion pulldown; truncation analysis; glycosylation-deficient expression\",\n      \"journal\": \"Journal of the American Society of Nephrology\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — multiple truncation and glycosylation mutants tested in orthogonal pulldown/Co-IP assays in a single study\",\n      \"pmids\": [\"12660326\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2006,\n      \"finding\": \"Neph1 is expressed in the developing mammalian brain and localizes to dendritic shafts and synaptic sites. Neph1 interacts with the PDZ domain of the synaptic scaffolding protein CASK via its cytoplasmic tail, suggesting a role in synaptogenesis.\",\n      \"method\": \"In situ hybridization; immunohistochemistry; immunogold EM; co-immunoprecipitation\",\n      \"journal\": \"The Journal of comparative neurology\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 3 / Moderate — subcellular localization by immunogold EM with biochemical Co-IP for CASK interaction, single lab\",\n      \"pmids\": [\"16874800\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2007,\n      \"finding\": \"Upon engagement, Neph1 is phosphorylated on specific tyrosine residues by the Src family kinase Fyn, which recruits the adaptor Grb2. This Grb2 recruitment is necessary for Neph1-induced actin polymerization at the plasma membrane. Neph1 and Nephrin directly interact and cooperate by juxtaposing Grb2 and Nck1/2 at the membrane to augment actin polymerization efficiency.\",\n      \"method\": \"Phosphorylation assay; co-immunoprecipitation; dominant-negative and siRNA knockdown; actin polymerization assay; mutagenesis\",\n      \"journal\": \"Molecular and cellular biology\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — kinase-substrate relationship established in cells, multiple orthogonal methods (Co-IP, mutagenesis, functional actin assay), single lab\",\n      \"pmids\": [\"17923684\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2008,\n      \"finding\": \"Fyn phosphorylates the cytoplasmic domain of Neph1 at multiple tyrosines (including Y637 and Y638) both in vitro and in intact cells. Phosphorylated Neph1 specifically binds adaptor protein Grb2 and tyrosine kinase Csk in a phosphorylation-dependent manner (from glomerular lysates). Neph1 attenuates Fyn-elicited ERK activation, requiring intact Grb2-SH2 binding motif. Phosphorylation of Y637 is upregulated in vivo in podocyte injury models.\",\n      \"method\": \"In vitro kinase assay; peptide mass fingerprinting; site-directed mutagenesis; GST pulldown from rat glomerular lysates; co-immunoprecipitation\",\n      \"journal\": \"The Journal of biological chemistry\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 / Moderate — in vitro kinase assay with mutagenesis identifying specific phosphorylation sites, pulldown from native tissue, functional ERK signaling readout\",\n      \"pmids\": [\"18258597\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2008,\n      \"finding\": \"Renal ischemia induces rapid loss of interaction between slit diaphragm proteins Neph1 and ZO-1, with redistribution of both proteins from cell membrane to cytoplasm. Recovery from ATP-depletion injury restores Neph1 tyrosine phosphorylation (mediated by Fyn), which is required for re-establishing Neph1-ZO-1 binding and their co-localization at the membrane. Fyn-mediated Neph1 phosphorylation significantly increases Neph1-ZO-1 binding.\",\n      \"method\": \"In vivo rat ischemia model; co-immunoprecipitation; cell culture ATP depletion; immunofluorescence; Fyn kinase assay\",\n      \"journal\": \"The Journal of biological chemistry\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — in vivo and in vitro models, kinase identified (Fyn), interaction dynamics measured by Co-IP under injury and recovery conditions\",\n      \"pmids\": [\"18922801\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2009,\n      \"finding\": \"Neph1 interacts with large-conductance Ca2+-activated K+ (BK/Slo1) channels via all three extreme C-terminal Slo1 splice variants, as shown by reciprocal Co-IP from endogenous podocyte and neuron proteins and GST pulldown. Co-expression of Neph1 with Slo1 suppresses steady-state Slo1 surface expression in HEK293T cells. Conversely, siRNA knockdown of Neph1 in ciliary ganglion neurons increases surface Slo1 and BK current, while knockdown in podocytes decreases Slo1 surface expression and BK current.\",\n      \"method\": \"Reciprocal co-immunoprecipitation from endogenous proteins; GST pulldown; cell surface biotinylation; whole-cell electrophysiology; siRNA knockdown; confocal microscopy\",\n      \"journal\": \"American journal of physiology. Cell physiology\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — reciprocal Co-IP from native tissue plus functional electrophysiology and loss-of-function in two cell types; multiple orthogonal methods\",\n      \"pmids\": [\"19794150\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2011,\n      \"finding\": \"Neph1 and Nephrin can trans-interact across cells to induce cell adhesion (demonstrated in L fibroblasts lacking endogenous adhesion). Neph1 alone can induce cell adhesion. Trans-interaction of nephrin with Neph1 (or Neph3) is associated with down-regulation of nephrin tyrosine phosphorylation.\",\n      \"method\": \"L fibroblast cell adhesion assay; co-immunoprecipitation; phosphorylation analysis\",\n      \"journal\": \"The Biochemical journal\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Weak — clean cell adhesion assay with defined readout, but single lab with limited mechanistic follow-up on phosphorylation\",\n      \"pmids\": [\"21306299\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2011,\n      \"finding\": \"Motor protein Myo1c directly interacts with Neph1 in an actin-dependent manner and is required for targeting Neph1 to the podocyte cell membrane. Expression of dominant-negative Myo1c or siRNA depletion of Myo1c significantly reduces membrane localization of Neph1 and nephrin. Myo1c knockdown also impairs cell migration and tight junction formation.\",\n      \"method\": \"Co-immunoprecipitation in vivo and in vitro; dominant-negative expression; siRNA knockdown; cell surface fractionation; wound assay; transepithelial electric resistance\",\n      \"journal\": \"Molecular and cellular biology\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — direct interaction confirmed by Co-IP, loss-of-function via two approaches (DN and siRNA) with defined membrane localization phenotype, multiple functional readouts\",\n      \"pmids\": [\"21402783\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2012,\n      \"finding\": \"The solution structure of the Neph1 cytoplasmic domain (Neph1-CD) was determined by SWAXS (radius of gyration 21.3 Å, max dimension 70 Å). A structural model of the Neph1-CD·ZO-1-PDZ1 complex was constructed; beyond the C-terminal Thr-His-Val motif, residues Lys-761 and Tyr-762 in Neph1 are critical for ZO-1 binding (individual alanine mutations abolished binding).\",\n      \"method\": \"Small/wide angle X-ray scattering (SWAXS); circular dichroism; in vitro and in vivo pulldown with mutagenesis\",\n      \"journal\": \"The Journal of biological chemistry\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 / Moderate — structural determination combined with site-directed mutagenesis and functional binding validation\",\n      \"pmids\": [\"22262837\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2014,\n      \"finding\": \"Inhibiting Neph1 signaling by transducing its cytoplasmic domain (Neph1CD) into podocytes reduces puromycin aminonucleoside (PAN)-induced Neph1 phosphorylation, retains Neph1 in lipid raft fractions, maintains Neph1 at the membrane, and protects against cytoskeletal damage and albumin leakage. Maintaining high membrane levels of Neph1 via chimeric overexpression increases podocyte resistance to PAN injury.\",\n      \"method\": \"Protein transduction; lipid raft fractionation; immunofluorescence; transepithelial permeability assay; zebrafish in vivo injury model\",\n      \"journal\": \"The Journal of biological chemistry\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — multiple orthogonal methods in vitro plus in vivo zebrafish model; single lab\",\n      \"pmids\": [\"24554715\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2016,\n      \"finding\": \"The full-length Myo1c adopts an extended S-shaped conformation in solution; Neph1 binds to the C-terminal tail of Myo1c without inducing significant conformational change. A critical residue in Neph1 required for Myo1c binding was identified; point mutation abolished the interaction in vitro and in live cells. FRAP analysis confirmed Myo1c mediates intracellular vesicular movement and membrane turnover of Neph1.\",\n      \"method\": \"Small angle X-ray scattering (SAXS); mutagenesis; in vitro binding assay; live-cell imaging; fluorescence recovery after photobleaching (FRAP)\",\n      \"journal\": \"Molecular and cellular biology\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 / Moderate — structural SAXS data combined with mutagenesis and FRAP live-cell imaging in a single study\",\n      \"pmids\": [\"27044863\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2017,\n      \"finding\": \"The Neph1-ZO-1 protein-protein interaction interface contains a druggable pocket involving both proteins. The small molecule isodesmosine (ISD) enhances Neph1 cytoplasmic domain–ZO-1 binding in vitro and in vivo, and ISD-treated podocytes are resistant to injury-induced transepithelial permeability loss. Mouse and zebrafish experiments confirm ISD protects from injury-induced renal damage.\",\n      \"method\": \"Computational structural screening; biochemical binding assay; in vitro podocyte permeability assay; mouse and zebrafish injury models\",\n      \"journal\": \"Scientific reports\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — multiple in vitro and in vivo models but computational component and single lab\",\n      \"pmids\": [\"28935902\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2017,\n      \"finding\": \"CD80 (B7-1) interacts with Neph1 via their extracellular domains, as established by pulldown assay in HEK293 cells co-transfected with both proteins. CD80 overexpression in podocytes co-localizes with Neph1 and causes actin derangement.\",\n      \"method\": \"Co-immunoprecipitation/pulldown in HEK293 cells; immunofluorescence co-localization in podocytes\",\n      \"journal\": \"Clinical and experimental nephrology\",\n      \"confidence\": \"Low\",\n      \"confidence_rationale\": \"Tier 3 / Weak — single pulldown in overexpression system, limited mechanistic follow-up, single lab\",\n      \"pmids\": [\"29022109\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2019,\n      \"finding\": \"Homozygous mutations in KIRREL1 in patients with steroid-resistant nephrotic syndrome result in mutant KIRREL1 proteins that fail to localize to the podocyte cell membrane, indicating defective membrane trafficking.\",\n      \"method\": \"Patient genetics; immunofluorescence localization of mutant vs wild-type KIRREL1 in podocytes\",\n      \"journal\": \"Kidney international\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Weak — direct localization experiment with functional consequence (nephrotic syndrome), but single paper and limited mechanistic detail on trafficking mechanism\",\n      \"pmids\": [\"31472902\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2021,\n      \"finding\": \"NEPHRIN and NEPH1 are novel receptor proteins for hepatocyte growth factor (HGF), binding HGF directly through their extracellular domains (confirmed by surface plasmon resonance with purified recombinant proteins; NEPHRIN Kd ~20-fold higher affinity than NEPH1). HGF binding leads to phosphorylation of NEPHRIN and NEPH1 independently of the MET receptor. SHP-2 (PTPN11) mediates dephosphorylation of these proteins. HGF-induced phosphorylation promotes podocyte repair.\",\n      \"method\": \"Surface plasmon resonance with purified baculovirus-expressed recombinant proteins; molecular modeling; cell culture and Drosophila nephrocyte functional assays\",\n      \"journal\": \"The Journal of biological chemistry\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 / Moderate — in vitro binding reconstitution with purified proteins (SPR), site mapping, and functional in vitro and in vivo validation\",\n      \"pmids\": [\"34391780\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2022,\n      \"finding\": \"KIRREL1 physically interacts with SAV1 and recruits SAV1 to cell-cell contact sites, thereby activating the Hippo pathway. Knockout of KIRREL1 increases YAP activity in neighboring cells. During liver regeneration in mice, KIRREL1 knockout enhances hepatic YAP activity, hepatocyte reprogramming, and biliary epithelial cell proliferation.\",\n      \"method\": \"Co-immunoprecipitation; knockout mouse model; in vivo liver regeneration assay; YAP activity reporter\",\n      \"journal\": \"Nature communications\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — direct physical interaction (Co-IP), in vivo KO phenotype with specific pathway readouts (YAP activity), replicated by independent lab in same year\",\n      \"pmids\": [\"35177623\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2022,\n      \"finding\": \"KIRREL1 interacts with both SAV1 and LATS1/2, promoting LATS1/2 activation by MST1/2 (Hippo kinases), thereby suppressing YAP/TAZ oncoproteins. YAP/TAZ in turn directly induce KIRREL1 expression in a TEAD1-4-dependent manner, constituting a negative feedback loop. Transgenic expression of KIRREL1 blocks tumorigenesis in a mouse intrahepatic cholangiocarcinoma model.\",\n      \"method\": \"Co-immunoprecipitation; LATS kinase activity assay; CRISPR screen; TEAD reporter assay; transgenic mouse tumor model\",\n      \"journal\": \"Cell reports\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — kinase activity assay, multiple interactors confirmed by Co-IP, in vivo tumor model, replicated by independent lab in same year\",\n      \"pmids\": [\"36044856\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2022,\n      \"finding\": \"KIRREL binds directly to SAV1 to activate the Hippo tumor suppressor pathway. KIRREL loss markedly promotes tumor growth in vivo. This was discovered through an in vivo CRISPR screen of cell surface proteins and a separate CRISPR screen using a Hippo pathway reporter.\",\n      \"method\": \"In vivo CRISPR proliferation screen; Hippo pathway reporter CRISPR screen; direct binding assay\",\n      \"journal\": \"Proceedings of the National Academy of Sciences\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — CRISPR functional screens plus direct binding assay; partially overlapping findings with PMID 35177623 and 36044856\",\n      \"pmids\": [\"35704761\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2023,\n      \"finding\": \"KIRREL1 knockdown inhibits proliferation and angiogenesis of gastric cancer cells; overexpression promotes them. These effects are mediated through the PI3K/AKT/mTOR pathway: KIRREL1 silencing represses P-PI3K, P-AKT, P-mTOR, HIF-1α, and VEGF, while overexpression stimulates them. The AKT agonist IGF-1 and inhibitor LY294002 reverse these effects.\",\n      \"method\": \"Lentiviral knockdown/overexpression; western blot; pharmacological pathway inhibition/activation\",\n      \"journal\": \"Journal of cellular and molecular medicine\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 3 / Moderate — multiple cell-based assays with pharmacological validation of pathway; single lab, no direct binding to PI3K components demonstrated\",\n      \"pmids\": [\"37909722\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2024,\n      \"finding\": \"PRRXL1 homeodomain transcription factor directly regulates Neph1 transcription by binding to four intronic regions of the Neph1 gene (identified by ChIP), preventing premature Neph1 expression in dorsal horn superficial laminae at E14.5. Loss-of-function experiments show Neph1 is required for neurite branching, especially at distal neurites.\",\n      \"method\": \"Chromatin immunoprecipitation (ChIP); in situ hybridization; loss-of-function (Neph1 KO or knockdown); neurite morphometry\",\n      \"journal\": \"Neural development\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Weak — ChIP establishes direct transcriptional regulation, loss-of-function with morphological readout; single lab\",\n      \"pmids\": [\"39049046\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2025,\n      \"finding\": \"Cryo-electron tomography of human kidney tissue resolves the slit diaphragm as a fishnet-like lattice with crisscrossing strands at ~90°. An atomic model based on the Nephrin-Neph1 heterodimer fits the structure, with ~9 nm spacing in humans (vs 12.3 nm in mice and 15 nm in Drosophila), indicating the SD is a conserved heterodimeric fishnet assembly.\",\n      \"method\": \"Cryo-electron tomography of native human and mouse kidney tissue; atomic model fitting\",\n      \"journal\": \"bioRxiv\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 1 / Weak — high-resolution structural method on native tissue, preprint not yet peer-reviewed, single study\",\n      \"pmids\": [\"bio_10.1101_2025.09.24.678239\"],\n      \"is_preprint\": true\n    }\n  ],\n  \"current_model\": \"KIRREL1/NEPH1 is a transmembrane immunoglobulin superfamily protein that localizes to the glomerular slit diaphragm (forming part of a fishnet-like heterodimeric lattice with Nephrin) and to neuronal synapses, where it forms cis- and trans-heterodimers with Nephrin via both extracellular Ig domains and cytoplasmic interactions, is phosphorylated on specific tyrosines by the Src-family kinase Fyn, recruits Grb2 (via pY637/Y638) to drive actin polymerization through cooperation with Nck, binds ZO-1 via its cytoplasmic tail (stabilized by Fyn-mediated phosphorylation), is transported to the membrane by the motor protein Myo1c, acts as a receptor for HGF, and functions as an upstream positive regulator of the Hippo pathway by interacting with both SAV1 and LATS1/2 at cell-cell contacts to promote LATS1/2 activation and YAP/TAZ suppression.\"\n}\n```","stage2_raw":"{\n  \"mechanistic_narrative\": \"KIRREL1/NEPH1 is a transmembrane immunoglobulin-superfamily adhesion receptor that is required to maintain the structural integrity of the glomerular filtration barrier in podocytes [#0]. At the slit diaphragm it localizes to the lateral margins of podocyte foot processes [#2, #3] and assembles a heterodimeric, fishnet-like lattice with Nephrin: its extracellular Ig domains mediate glycosylation-dependent cis- and trans-heterodimerization with Nephrin (it does not engage in homophilic interactions in foot processes) [#3, #4, #24], while its cytoplasmic tail couples to the slit-diaphragm scaffold through a C-terminal motif and residues Lys-761/Tyr-762 that bind the ZO-1 PDZ domain [#2, #12] and through podocin [#1]. Upon engagement KIRREL1 is phosphorylated on cytoplasmic tyrosines (including Y637/Y638) by the Src-family kinase Fyn, which recruits Grb2 to drive actin polymerization in cooperation with Nephrin-bound Nck and also recruits Csk to attenuate ERK signaling [#6, #7]; Fyn-dependent phosphorylation likewise stabilizes the KIRREL1–ZO-1 interaction and membrane localization, a coupling that is acutely lost during ischemic and ATP-depletion injury and restored on recovery [#8]. Membrane delivery and turnover of KIRREL1 depend on direct, actin-dependent binding to the motor protein Myo1c [#11, #14], and failure of mutant KIRREL1 to reach the podocyte membrane causes steroid-resistant nephrotic syndrome in patients with homozygous KIRREL1 mutations [#17]. Beyond the kidney, KIRREL1 acts as a direct receptor for HGF, binding through its extracellular domain and undergoing MET-independent phosphorylation that promotes podocyte repair [#18], and it functions as an upstream positive regulator of the Hippo pathway by recruiting SAV1 and LATS1/2 to cell–cell contacts to promote LATS1/2 activation and YAP/TAZ suppression within a TEAD-dependent negative feedback loop, restraining tumorigenesis in liver and biliary models [#19, #20, #21]. KIRREL1/NEPH1 is also expressed at neuronal synapses and dendrites, where it binds CASK and is required for neurite branching [#5, #23].\",\n  \"teleology\": [\n    {\n      \"year\": 2001,\n      \"claim\": \"Established that NEPH1 is physiologically essential for the glomerular filtration barrier, framing it as a slit-diaphragm component rather than an incidental podocyte protein.\",\n      \"evidence\": \"Gene-trap knockout mice with EM and proteinuria readouts\",\n      \"pmids\": [\"11416156\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Did not define molecular partners or the biochemical basis of the phenotype\", \"No mechanism for how loss leads to foot process effacement\"]\n    },\n    {\n      \"year\": 2003,\n      \"claim\": \"Resolved where and with what NEPH1 acts at the slit diaphragm, showing it localizes to the slit-diaphragm insertion and forms a Nephrin-linked, ZO-1-coupled adhesion complex whose integrity sustains the barrier.\",\n      \"evidence\": \"Immunogold EM, Co-IP, in vivo antibody perturbation, cell fractionation\",\n      \"pmids\": [\"12865409\", \"12646566\", \"12660326\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Did not establish the intracellular signaling consequences of complex assembly\", \"Trans- vs cis-orientation of Nephrin–NEPH1 not fully resolved at this stage\"]\n    },\n    {\n      \"year\": 2002,\n      \"claim\": \"Identified podocin as a cytoplasmic NEPH1 partner and a tyrosine-dependent binding motif, linking NEPH1 to other slit-diaphragm scaffolding and to AP-1 transcriptional output.\",\n      \"evidence\": \"Co-IP, mutagenesis, transcriptional reporter assay\",\n      \"pmids\": [\"12424224\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Functional significance of AP-1 activation in podocytes unresolved\", \"Tec-family kinase identity not pinned down\"]\n    },\n    {\n      \"year\": 2008,\n      \"claim\": \"Defined the kinase-substrate logic of NEPH1 signaling, showing Fyn phosphorylates specific cytoplasmic tyrosines (Y637/Y638) to recruit Grb2 and Csk and drive actin polymerization, connecting adhesion to cytoskeletal remodeling.\",\n      \"evidence\": \"In vitro kinase assay, mass fingerprinting, mutagenesis, glomerular pulldown, actin polymerization assay\",\n      \"pmids\": [\"17923684\", \"18258597\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Quantitative contribution of NEPH1 vs Nephrin to actin output not separated\", \"In vivo relevance of ERK attenuation unclear\"]\n    },\n    {\n      \"year\": 2008,\n      \"claim\": \"Showed the NEPH1–ZO-1 interaction is dynamically regulated by Fyn phosphorylation and disrupted by injury, providing a molecular mechanism for slit-diaphragm disassembly and recovery during ischemia.\",\n      \"evidence\": \"In vivo rat ischemia model, ATP depletion, Co-IP, immunofluorescence, Fyn kinase assay\",\n      \"pmids\": [\"18922801\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Upstream signal triggering Fyn reactivation during recovery not identified\"]\n    },\n    {\n      \"year\": 2011,\n      \"claim\": \"Identified Myo1c as the motor delivering NEPH1 (and Nephrin) to the membrane, explaining how the slit-diaphragm complex is trafficked and assembled.\",\n      \"evidence\": \"Co-IP, dominant-negative and siRNA, surface fractionation, wound and TEER assays\",\n      \"pmids\": [\"21402783\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Regulation of Myo1c-dependent delivery not defined\", \"Cargo selectivity mechanism unknown\"]\n    },\n    {\n      \"year\": 2012,\n      \"claim\": \"Provided structural definition of the NEPH1 cytoplasmic domain and its ZO-1 interface, identifying Lys-761/Tyr-762 as critical contacts and rendering the interaction structurally tractable.\",\n      \"evidence\": \"SWAXS, circular dichroism, mutagenesis pulldowns\",\n      \"pmids\": [\"22262837\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"No high-resolution atomic structure of the full complex\", \"Phospho-regulation not captured structurally\"]\n    },\n    {\n      \"year\": 2016,\n      \"claim\": \"Structurally characterized the NEPH1–Myo1c interaction and demonstrated by FRAP that Myo1c mediates vesicular movement and membrane turnover of NEPH1.\",\n      \"evidence\": \"SAXS, mutagenesis, in vitro binding, live-cell FRAP\",\n      \"pmids\": [\"27044863\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Coupling of motor activity to specific trafficking routes unresolved\"]\n    },\n    {\n      \"year\": 2014,\n      \"claim\": \"Demonstrated that maintaining membrane NEPH1 and its signaling protects podocytes from injury, establishing NEPH1 stabilization as a candidate protective strategy.\",\n      \"evidence\": \"Protein transduction, lipid raft fractionation, permeability assays, zebrafish injury model\",\n      \"pmids\": [\"24554715\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Single lab\", \"Mechanism by which Neph1CD retains the protein in rafts not fully defined\"]\n    },\n    {\n      \"year\": 2017,\n      \"claim\": \"Showed the NEPH1–ZO-1 interface is druggable, with the small molecule isodesmosine enhancing binding and conferring injury resistance in vitro and in vivo.\",\n      \"evidence\": \"Computational screening, binding assay, podocyte permeability assay, mouse and zebrafish injury models\",\n      \"pmids\": [\"28935902\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Single lab\", \"Direct target engagement specificity of ISD not orthogonally confirmed\"]\n    },\n    {\n      \"year\": 2017,\n      \"claim\": \"Reported CD80 (B7-1) as an extracellular NEPH1 partner associated with actin derangement, hinting at an immune-injury link.\",\n      \"evidence\": \"Pulldown in HEK293 overexpression, immunofluorescence co-localization\",\n      \"pmids\": [\"29022109\"],\n      \"confidence\": \"Low\",\n      \"gaps\": [\"Single pulldown in overexpression system without reciprocal validation\", \"No endogenous interaction or in vivo evidence\"]\n    },\n    {\n      \"year\": 2019,\n      \"claim\": \"Linked KIRREL1 directly to human disease, showing homozygous mutations cause steroid-resistant nephrotic syndrome through defective membrane trafficking of mutant protein.\",\n      \"evidence\": \"Patient genetics, immunofluorescence localization of mutant vs WT KIRREL1\",\n      \"pmids\": [\"31472902\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Single study\", \"Trafficking step that fails not mechanistically defined\"]\n    },\n    {\n      \"year\": 2009,\n      \"claim\": \"Extended NEPH1 function beyond scaffolding by showing it regulates BK/Slo1 channel surface expression and current in podocytes and neurons.\",\n      \"evidence\": \"Reciprocal endogenous Co-IP, GST pulldown, surface biotinylation, electrophysiology, siRNA\",\n      \"pmids\": [\"19794150\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Directionality of regulation differs between cell types and is unexplained\", \"Structural basis of Slo1 binding not defined\"]\n    },\n    {\n      \"year\": 2021,\n      \"claim\": \"Revealed NEPH1 (and Nephrin) as a direct HGF receptor undergoing MET-independent, SHP-2-reversible phosphorylation that promotes podocyte repair, recasting NEPH1 as a ligand-responsive signaling receptor.\",\n      \"evidence\": \"SPR with purified recombinant proteins, modeling, cell and Drosophila nephrocyte assays\",\n      \"pmids\": [\"34391780\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Downstream effectors of HGF-induced NEPH1 phosphorylation not fully mapped\", \"Relative in vivo contribution vs MET signaling unclear\"]\n    },\n    {\n      \"year\": 2022,\n      \"claim\": \"Defined a distinct role for KIRREL1 as an upstream positive regulator of the Hippo pathway, recruiting SAV1 and LATS1/2 to cell-cell contacts to activate LATS and suppress YAP/TAZ within a TEAD-dependent feedback loop, with tumor-suppressive consequences.\",\n      \"evidence\": \"Co-IP, KO and transgenic mouse models, CRISPR screens, LATS kinase and TEAD reporter assays, liver regeneration and cholangiocarcinoma models\",\n      \"pmids\": [\"35177623\", \"36044856\", \"35704761\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"How adhesion-based KIRREL1 engagement is transduced to LATS activation mechanistically unresolved\", \"Connection between slit-diaphragm and Hippo roles not reconciled\"]\n    },\n    {\n      \"year\": 2023,\n      \"claim\": \"Reported a pro-tumorigenic, PI3K/AKT/mTOR-linked role for KIRREL1 in gastric cancer, contrasting with its Hippo tumor-suppressive function and indicating context-dependent outputs.\",\n      \"evidence\": \"Lentiviral knockdown/overexpression, western blot, pharmacological pathway modulation\",\n      \"pmids\": [\"37909722\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"No direct binding to PI3K components shown\", \"Single lab\", \"Reconciliation with Hippo tumor-suppressor role absent\"]\n    },\n    {\n      \"year\": 2024,\n      \"claim\": \"Established transcriptional control of Neph1 by the homeodomain factor PRRXL1 and a functional requirement for Neph1 in neurite branching, deepening its neuronal role.\",\n      \"evidence\": \"ChIP, in situ hybridization, loss-of-function, neurite morphometry\",\n      \"pmids\": [\"39049046\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Single lab\", \"Molecular mechanism of Neph1 in neurite branching unresolved\"]\n    },\n    {\n      \"year\": 2025,\n      \"claim\": \"Resolved the native slit diaphragm as a conserved Nephrin–NEPH1 heterodimeric fishnet lattice with species-specific spacing, providing the supramolecular architecture underlying the barrier.\",\n      \"evidence\": \"Cryo-electron tomography of native human and mouse kidney tissue with atomic model fitting (preprint)\",\n      \"pmids\": [\"bio_10.1101_2025.09.24.678239\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Preprint, not peer-reviewed\", \"Single study\", \"Dynamics of lattice remodeling not captured\"]\n    },\n    {\n      \"year\": null,\n      \"claim\": \"How KIRREL1's adhesion/scaffolding function at cell-cell contacts is mechanistically transduced into Hippo (LATS) activation, and how its opposing tumor-suppressive versus pro-proliferative outputs are determined by cellular context, remain unresolved.\",\n      \"evidence\": \"\",\n      \"pmids\": [],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"No mechanism linking extracellular engagement to LATS activation\", \"Context-dependent tumor roles not reconciled\", \"Integration of kidney, neuronal, and Hippo functions unknown\"]\n    }\n  ],\n  \"mechanism_profile\": {\n    \"molecular_activity\": [\n      {\"term_id\": \"GO:0098631\", \"supporting_discovery_ids\": [3, 4, 10, 24]},\n      {\"term_id\": \"GO:0001618\", \"supporting_discovery_ids\": [18]},\n      {\"term_id\": \"GO:0060089\", \"supporting_discovery_ids\": [6, 7, 18]},\n      {\"term_id\": \"GO:0060090\", \"supporting_discovery_ids\": [2, 6, 19, 20]}\n    ],\n    \"localization\": [\n      {\"term_id\": \"GO:0005886\", \"supporting_discovery_ids\": [2, 3, 11, 17]},\n      {\"term_id\": \"GO:0005829\", \"supporting_discovery_ids\": [8]}\n    ],\n    \"pathway\": [\n      {\"term_id\": \"R-HSA-162582\", \"supporting_discovery_ids\": [6, 7, 19, 20, 18]},\n      {\"term_id\": \"R-HSA-1474244\", \"supporting_discovery_ids\": [2, 3, 24]},\n      {\"term_id\": \"R-HSA-1643685\", \"supporting_discovery_ids\": [17, 20]}\n    ],\n    \"complexes\": [\"slit diaphragm (Nephrin-NEPH1 fishnet lattice)\"],\n    \"partners\": [\"NPHS1\", \"NPHS2\", \"TJP1\", \"FYN\", \"GRB2\", \"MYO1C\", \"SAV1\", \"LATS1\"],\n    \"other_free_text\": []\n  }\n}","audit_flag":null,"evaluation":{"pairwise":"win","faith_supported":6,"faith_total":6,"faith_pct":100.0}}