{"gene":"SLC10A1","run_date":"2026-06-10T07:46:32","timeline":{"discoveries":[{"year":2022,"finding":"Cryo-EM structures of human, bovine, and rat NTCPs in the apo state reveal a tunnel across the membrane and a possible transport route for the bile acid substrate. The structure of human NTCP in complex with the myristoylated preS1 domain of the HBV large surface protein, together with mutation and transport assays, shows that preS1 and the bile acid substrate compete for the extracellular opening of the tunnel, establishing the binding mode for viral entry.","method":"Cryo-electron microscopy, mutation assays, transport assays","journal":"Nature","confidence":"High","confidence_rationale":"Tier 1 / Strong — cryo-EM structure with mutagenesis and functional transport assays in a single rigorous study; independently reported by two concurrent Nature papers","pmids":["35580629"],"is_preprint":false},{"year":2022,"finding":"Cryo-EM structure of NTCP bound to an antibody shows the transporter lacks the first transmembrane helix found in other SLC10 proteins and has an N-terminus exposed on the extracellular face. Comparison with related proteins indicates a common mechanism of bile acid transport, and an additional pocket formed by residues known to interact with preS1 was identified.","method":"Cryo-electron microscopy, structural comparison","journal":"Nature","confidence":"High","confidence_rationale":"Tier 1 / Strong — cryo-EM structure; independently replicated by concurrent parallel paper (PMID:35580629)","pmids":["35580630"],"is_preprint":false},{"year":2024,"finding":"Cryo-EM structure of bulevirtide (BLV/Myrcludex B)-bound human NTCP reveals BLV forms two domains: a plug lodged in the bile salt transport tunnel of NTCP and a string covering the extracellular surface, with the N-terminal myristoyl group interacting with the lipid-exposed surface of NTCP. This structure explains how BLV blocks bile salt transport and provides a structural basis for HBV/HDV host specificity.","method":"Cryo-electron microscopy","journal":"Nature communications","confidence":"High","confidence_rationale":"Tier 1 / Strong — atomic-resolution cryo-EM structure with mechanistic interpretation; single rigorous study with multiple orthogonal validations implied by structural detail","pmids":["38509088"],"is_preprint":false},{"year":2013,"finding":"NTCP (SLC10A1) was confirmed as the functional HBV entry receptor: HepG2 cells engineered to overexpress human NTCP became susceptible to HBV infection, and knockdown of NTCP blocked infection. Compounds that inhibit NTCP transporter activity (including cyclosporin A and oxysterols) also blocked HBV infection, linking bile acid transport function to viral entry.","method":"NTCP overexpression in HepG2 cells, siRNA knockdown, HBV infection assay, inhibitor studies","journal":"Biochemical and biophysical research communications","confidence":"High","confidence_rationale":"Tier 2 / Strong — gain-of-function and loss-of-function with defined phenotypic readout; replicated across multiple labs","pmids":["24342612"],"is_preprint":false},{"year":2013,"finding":"Cyclosporin A inhibits HBV and HDV entry by directly interfering with NTCP receptor function in a cyclophilin-independent manner. Binding of the HBVpreS1 domain to NTCP was blocked by cyclosporin A, and an NTCP variant deficient in HBVpreS1 binding but competent for bile salt transport was resistant to cyclosporin A, mapping the interaction site to the preS1-binding domain on NTCP.","method":"HepaRG cells and NTCP-expressing hepatoma cell lines, taurocholate uptake assay, HBVpreS1 binding assay, cyclophilin siRNA, NTCP variant studies","journal":"Journal of hepatology","confidence":"High","confidence_rationale":"Tier 2 / Strong — multiple orthogonal methods (transport assay, binding assay, RNAi, variant rescue), replicated across multiple cell models","pmids":["24295872"],"is_preprint":false},{"year":2014,"finding":"The myristoylated preS1 domain of HBV inhibits bile acid transport by NTCP, and inversely, bile acid conjugates (taurine/glycine conjugates of cholic acid and ursodeoxycholic acid) inhibit HBV infection in a concentration-dependent manner. NTCP expression, transport function, preS1 peptide binding, and HBV infection follow comparable kinetics, establishing direct functional overlap between the bile acid transport site and the HBV receptor site.","method":"Bile acid transport assays, HBV infection assays, myr-preS1 peptide binding in primary hepatocytes and NTCP-transfected cells","journal":"Journal of hepatology","confidence":"High","confidence_rationale":"Tier 2 / Strong — reciprocal inhibition demonstrated in primary human hepatocytes and multiple engineered cell lines with quantitative kinetics","pmids":["24845614"],"is_preprint":false},{"year":2005,"finding":"Dephosphorylation of Ser-226 in the third cytoplasmic loop of NTCP facilitates plasma membrane retention. Mutation of Ser-226 to Ala decreased NTCP phosphorylation by 30% and increased taurocholate uptake and plasma membrane retention 2.5–3.2-fold; cAMP failed to further increase translocation of S226A-NTCP, establishing Ser-226 as the cAMP-regulated phosphorylation site controlling NTCP plasma membrane localization.","method":"Metabolic [32P] labeling, clostripain digestion, site-directed mutagenesis, taurocholate uptake assay, cell surface expression in transfected HuH-7 cells","journal":"The Journal of biological chemistry","confidence":"High","confidence_rationale":"Tier 1 / Strong — site-directed mutagenesis combined with phosphorylation mapping and functional transport assays in the same study","pmids":["16027164"],"is_preprint":false},{"year":2002,"finding":"PP2B (calcineurin), a Ca2+/calmodulin-dependent phosphatase, mediates cAMP-induced dephosphorylation and translocation of NTCP to the plasma membrane. The PP2B inhibitor cypermethrin reversed cAMP-mediated NTCP dephosphorylation and translocation, and PP2B directly dephosphorylated immunoprecipitated NTCP from control but not cAMP-treated hepatocytes.","method":"PP2B inhibitors (cypermethrin, FK-506), taurocholate uptake assay, NTCP immunoprecipitation, PP2B activity assay in isolated rat hepatocytes","journal":"American journal of physiology. Gastrointestinal and liver physiology","confidence":"High","confidence_rationale":"Tier 1 / Strong — in vitro dephosphorylation of immunoprecipitated NTCP by PP2B, pharmacological inhibition with multiple compounds, and functional readout","pmids":["12065290"],"is_preprint":false},{"year":2002,"finding":"Protein kinase B (PKB/Akt) mediates cAMP- and cell swelling-stimulated Na+/taurocholate cotransport and NTCP translocation to the plasma membrane via the PI3K/PKB signaling pathway. Dominant-negative PKB blocked cAMP- and swelling-induced increases in TC uptake and NTCP translocation, while constitutively active PKB increased both.","method":"Dominant-negative and constitutively active PKB transfection, taurocholate uptake assay, NTCP translocation assay in HuH-Ntcp cells","journal":"The Journal of biological chemistry","confidence":"High","confidence_rationale":"Tier 2 / Strong — genetic dominant-negative and constitutively active constructs with functional readout; multiple stimuli tested","pmids":["12034724"],"is_preprint":false},{"year":2006,"finding":"NTCP-containing vesicles are present on intracellular recycling endosomes and move bidirectionally on microtubules using kinesin-1 and dynein motors. PKCζ is specifically required for microtubule-based motility of NTCP vesicles: PI(3,4,5)P3 activates PKCζ and enhances motility, while specific inhibition of PKCζ blocks motility of NTCP-containing vesicles but not late vesicles.","method":"In vitro and whole-cell immunofluorescence microscopy, live-cell imaging of NTCP-GFP, PKCζ inhibitor, colocalization with endosomal markers","journal":"Traffic (Copenhagen, Denmark)","confidence":"High","confidence_rationale":"Tier 2 / Strong — direct live-imaging and in vitro motility assays with pharmacological and genetic inhibition; functional consequence demonstrated","pmids":["16734659"],"is_preprint":false},{"year":2005,"finding":"Rat Ntcp/human NTCP is degraded by the ubiquitin-proteasome system via ER-associated degradation (ERAD). Proteasome inhibition led to accumulation of core-glycosylated (ER-resident) intracellular NTCP that co-localized with ubiquitin at the microtubule organization center as polyubiquitinated aggresomes.","method":"Proteasome inhibitors (MG-132, lactacystin), co-localization studies, polyubiquitination assay, glycosylation analysis in stably transfected HepG2 cells","journal":"Biological chemistry","confidence":"High","confidence_rationale":"Tier 2 / Strong — multiple pharmacological inhibitors, biochemical characterization of ubiquitinated species, and glycosylation state analysis","pmids":["16218878"],"is_preprint":false},{"year":2002,"finding":"Interleukin-1β suppresses Ntcp gene expression via a JNK-dependent mechanism: JNK phosphorylates RXR, reducing nuclear RXR:RAR binding activity and thereby decreasing RXR:RAR-mediated transactivation of the Ntcp promoter. JNK inhibition by curcumin or dominant-negative JNK completely blocked IL-1β-mediated suppression, while ERK and p38 inhibitors had no effect.","method":"Ntcp promoter luciferase assays, JNK expression plasmids (dominant-negative and wild-type), pharmacological inhibitors, EMSA, primary rat hepatocytes","journal":"The Journal of biological chemistry","confidence":"High","confidence_rationale":"Tier 2 / Strong — multiple orthogonal methods (promoter assay, dominant-negative JNK, pharmacological inhibitors, EMSA); confirmed in primary hepatocytes","pmids":["12105223"],"is_preprint":false},{"year":2003,"finding":"HNF1α, HNF4α, and RXRα/RARα activate the rat Ntcp promoter but not the human or mouse NTCP/Ntcp promoters, whereas C/EBPβ activation is specific to human and mouse. HNF3β is the only transcription factor binding and active at a conserved motif in all three species, where it inhibits NTCP/Ntcp promoter activity. Small heterodimer partner (SHP) does not affect NTCP/Ntcp promoter activity from the conserved region.","method":"Transfection-based luciferase reporter assays, electrophoretic mobility shift assay (EMSA), cotransfection in Huh7 cells","journal":"American journal of physiology. Gastrointestinal and liver physiology","confidence":"High","confidence_rationale":"Tier 2 / Strong — EMSA and promoter luciferase assays across three species with multiple transcription factors","pmids":["14701722"],"is_preprint":false},{"year":2002,"finding":"Short heterodimer partner 1 (SHP-1) induction by retained bile acids temporally precedes and likely mediates downregulation of Ntcp in obstructive cholestasis. In bile duct-ligated mice, SHP-1 mRNA peaked at 6 h after ligation, followed 6 h later by the nadir of Ntcp mRNA and protein.","method":"Northern and Western blotting of SHP-1 and Ntcp in CBDL mice, immunofluorescence","journal":"American journal of physiology. Gastrointestinal and liver physiology","confidence":"Medium","confidence_rationale":"Tier 3 / Moderate — temporal correlation in a genetic knockout model without direct mechanistic intervention; confirmed in two genotypes (FXR+/+ and FXR-/-) in follow-up paper","pmids":["11751172"],"is_preprint":false},{"year":2005,"finding":"In obstructive cholestasis (CBDL), Ntcp downregulation is bile acid–mediated and FXR-dependent, not cytokine-mediated. FXR knockout mice fail to repress Ntcp after bile acid feeding or CBDL; LPS-induced Ntcp repression is FXR-independent. The mechanism involves reduced nuclear levels and DNA binding of HNF-1α, HNF-4α, RXRα, and RARα.","method":"FXR knockout mice, bile duct ligation, cholic acid feeding, LPS treatment, EMSA, Western blotting, cytokine inactivation","journal":"American journal of physiology. Gastrointestinal and liver physiology","confidence":"High","confidence_rationale":"Tier 2 / Strong — genetic epistasis with FXR-/- mice using multiple interventions and multiple readouts","pmids":["16002565"],"is_preprint":false},{"year":2005,"finding":"Kupffer cell depletion with liposomal clodronate prevents endotoxin-induced suppression of Ntcp expression. In endotoxin-exposed rats, KC depletion preserved Ntcp RNA and protein expression and maintained RXR:RAR and HNF1α transcription factor binding activity.","method":"Liposomal clodronate KC depletion, LPS exposure, Northern/Western blotting, EMSA in rats","journal":"Journal of hepatology","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — depletion experiment with defined molecular readout (transcription factor binding), single lab","pmids":["15629514"],"is_preprint":false},{"year":2005,"finding":"In obstructive cholestasis in rodents, accumulating bile acids—independent of cytokines—downregulate Ntcp through repression of HNF-1α and HNF-4α. Both TNF-α/IL-1β neutralization and Kupffer cell depletion failed to restore Ntcp expression, while SHP mRNA increased 3–5-fold.","method":"Cytokine neutralization (etanercept, anakinra), KC depletion by liposome clodronate, Western/Northern blotting, EMSA in CBDL rats and mice","journal":"Hepatology (Baltimore, Md.)","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — pharmacological and cellular depletion with multiple molecular readouts; single lab","pmids":["15723437"],"is_preprint":false},{"year":2003,"finding":"NTCP (Ntcp) facilitates hepatocellular uptake of the lethal mushroom toxin α-amanitin. HepG2 cells stably transfected with rat Ntcp showed markedly increased sensitivity to α-amanitin cytotoxicity compared to non-transfected cells, as measured by suppression of cytokine-induced IL-1Ra mRNA synthesis.","method":"Stable transfection of rat Ntcp in HepG2 cells, taurocholate uptake kinetics, functional toxicity assay (IL-1Ra mRNA induction)","journal":"Archives of toxicology","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — gain-of-function with a defined functional assay; single lab","pmids":["14598021"],"is_preprint":false},{"year":2006,"finding":"Human NTCP and BSEP reconstituted together in LLC-PK1 polarized cells mediate vectorial transcellular transport of bile acids from the basolateral to apical side, with substrate specificities showing taurine conjugates > glycine conjugates > unconjugated bile salts for NTCP. Single-transporter cells showed no vectorial transport.","method":"Double transfection of LLC-PK1 cells with NTCP and BSEP, transcellular bile acid flux assays, kinetic analysis","journal":"American journal of physiology. Gastrointestinal and liver physiology","confidence":"High","confidence_rationale":"Tier 1 / Strong — reconstitution of vectorial transport in an epithelial cell system with kinetic characterization","pmids":["16474011"],"is_preprint":false},{"year":2017,"finding":"N-linked glycosylation of NTCP at residues N5 and N11 is required for efficient trafficking to the plasma membrane and for HBV infection. NTCP lacking both glycosylation sites (N5,11Q) failed to support HBV infection, showed minimal cellular expression, and was degraded in lysosomes, while single-glycan variants retained normal bile acid transport function and HBV infectability.","method":"Site-directed mutagenesis of glycosylation sites, cell surface biotinylation, lysosomal degradation assay, HBV infection assay, bile acid uptake assay in HepG2 cells","journal":"PloS one","confidence":"High","confidence_rationale":"Tier 1-2 / Strong — mutagenesis combined with multiple functional assays and trafficking analysis","pmids":["28125599"],"is_preprint":false},{"year":2003,"finding":"The C800T (Ser267Phe) polymorphism in NTCP causes near-complete loss of bile acid uptake function while preserving transport of the non-bile acid substrate estrone sulfate, identifying position 267 as part of a region critical and specific for bile acid substrate recognition. Cell surface biotinylation showed normal plasma membrane expression of this variant.","method":"Cell surface biotinylation, taurocholate and estrone sulfate transport assays, immunofluorescence confocal microscopy in HepG2 and transfected cells","journal":"The Journal of biological chemistry","confidence":"High","confidence_rationale":"Tier 2 / Strong — substrate-selective transport with normal membrane expression; multiple substrates and multiple cell models tested","pmids":["14660639"],"is_preprint":false},{"year":2013,"finding":"A dileucine motif in the third intracellular loop of Ntcp is essential for both endocytosis and plasma membrane targeting, indicating a dual trafficking function. PKC-mediated endocytosis of Ntcp from the plasma membrane is clathrin-dependent and is followed by lysosomal degradation. Mutation of Thr225 and Ser226 inhibited PKC-mediated endocytosis.","method":"Flow cytometry, immunofluorescence, Western blotting, site-directed mutagenesis of dileucine motif and phosphorylation sites in HepG2 cells","journal":"American journal of physiology. Gastrointestinal and liver physiology","confidence":"High","confidence_rationale":"Tier 2 / Strong — mutagenesis of multiple residues with parallel functional and localization readouts","pmids":["24008362"],"is_preprint":false},{"year":2010,"finding":"Nitric oxide (NO) inhibits taurocholate uptake by NTCP via S-nitrosylation of cysteine residues on NTCP, causing non-competitive inhibition (decreased Vmax, unchanged Km) and reduction of NTCP at the plasma membrane. Dithiothreitol reversed both S-nitrosylation and transport inhibition.","method":"Biotin switch assay for S-nitrosylation, taurocholate kinetic uptake assay, plasma membrane biotinylation, NO donor treatment in HuH-NTCP cells","journal":"American journal of physiology. Gastrointestinal and liver physiology","confidence":"High","confidence_rationale":"Tier 1-2 / Strong — S-nitrosylation established by biotin switch with mechanistic rescue by dithiothreitol; kinetic analysis performed","pmids":["21109590"],"is_preprint":false},{"year":2012,"finding":"Plasma membrane localization of PKCδ, rather than its kinase activity, is necessary for cAMP-induced NTCP translocation and Rab4 activation. Kinase-dead dominant-negative PKCδ and PKCδ knockdown both still increased plasma membrane NTCP and Rab4 activity when PKCδ was at the membrane. In contrast, kinase activity of PKCδ is required for cAMP-induced MRP2 translocation.","method":"Wild-type, kinase-dead DN-PKCδ transfection, siRNA knockdown, Rab4 activity assay, rottlerin and LY294002 inhibitors in HuH-NTCP cells","journal":"American journal of physiology. Gastrointestinal and liver physiology","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — genetic and pharmacological dissection with multiple constructs; single lab","pmids":["22744337"],"is_preprint":false},{"year":2015,"finding":"IL-6 inhibits HBV entry into hepatocytes by downregulating NTCP: IL-6 pretreatment reduced NTCP mRNA by 98%, reduced NTCP-mediated taurocholate uptake by 80%, and inhibited HBV entry by up to 90%. Restoration of NTCP expression suppressed the inhibitory effect of IL-6, confirming that NTCP downregulation mediates the effect.","method":"HBV infection assay (cccDNA and HBsAg readouts), taurocholate uptake assay, NTCP mRNA quantification, NTCP restoration experiment","journal":"Virology","confidence":"High","confidence_rationale":"Tier 2 / Strong — loss-of-function with rescue experiment and multiple readouts; dose-dependent kinetics demonstrated","pmids":["25765005"],"is_preprint":false},{"year":2014,"finding":"Retinoic acid receptor (RAR) regulates human NTCP promoter activity via a binding site at nucleotides −112 to −96 of the NTCP gene. Pharmacological antagonism of RAR (Ro41-5253, CD2665) reduced NTCP mRNA and protein expression, blocked viral entry, and inhibited HBV spread across multiple genotypes.","method":"NTCP promoter luciferase assays, RAR ChIP, pharmacological RAR antagonists, HBV infection assay, NTCP mRNA/protein quantification in HepG2-hNTCP cells","journal":"The Journal of biological chemistry","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — promoter mapping with ChIP and functional readout; single lab","pmids":["25550158"],"is_preprint":false},{"year":2016,"finding":"NTCP augments HCV infection by mediating bile-acid-dependent repression of interferon-stimulated genes (ISGs), including IFITM3. Gain- and loss-of-function studies in hepatocytes showed NTCP regulates innate antiviral immunity, with NTCP-mediated bile acid transport suppressing ISG expression and thereby increasing permissiveness to HCV infection.","method":"NTCP overexpression and knockdown in hepatoma cells, ISG expression profiling, HCV infection assay, bile acid treatment","journal":"Cell reports","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — gain- and loss-of-function with defined molecular mechanism (ISG repression); single lab","pmids":["27783949"],"is_preprint":false},{"year":2021,"finding":"NTCP oligomerization occurs downstream of the NTCP-EGFR interaction and is required for HBV internalization. Troglitazone directly binds NTCP allosterically (non-competitive inhibition of bile acid uptake) and blocks NTCP oligomerization, inhibiting HBV internalization without affecting HBV attachment or the NTCP-EGFR interaction. The F274A mutation in NTCP disrupts oligomerization and HBV internalization without affecting viral surface binding.","method":"Surface plasmon resonance, transporter kinetics, alanine scanning mutagenesis, coimmunoprecipitation, NTCP oligomerization assay in transfected cells","journal":"Journal of virology","confidence":"High","confidence_rationale":"Tier 2 / Strong — multiple orthogonal methods (SPR, mutagenesis, CoIP, kinetics) dissecting an ordered entry mechanism","pmids":["34613794"],"is_preprint":false},{"year":2022,"finding":"IFITM3 is a protein-protein interaction partner of NTCP identified by membrane yeast-two-hybrid and confirmed by co-immunoprecipitation. IFITM3 knockdown significantly reduced HBV and HDV infection of NTCP-expressing HuH7 cells and primary human hepatocytes, despite intact myr-preS1 peptide binding to NTCP, indicating IFITM3 facilitates a post-attachment step of HBV/HDV entry.","method":"Membrane yeast-two-hybrid, co-immunoprecipitation, IFITM3 siRNA knockdown, HBV/HDV infection assays, myr-preS1 binding assay","journal":"Viruses","confidence":"Medium","confidence_rationale":"Tier 2-3 / Moderate — membrane Y2H confirmed by CoIP with defined functional consequence in primary hepatocytes; single lab","pmids":["35458456"],"is_preprint":false},{"year":2019,"finding":"OATP1B3 forms hetero-oligomers with NTCP in both HEK293 cells and frozen human liver sections. Coexpression of NTCP with OATP1B3 in HEK293 cells increased OATP1B3 plasma membrane expression but decreased its apparent turnover rate for the substrate cholecystokinin-8.","method":"Co-immunoprecipitation, proximity ligation assay, plasma membrane expression quantification, transport assay in HEK293T cells and human liver sections","journal":"Drug metabolism and disposition","confidence":"Medium","confidence_rationale":"Tier 2-3 / Moderate — CoIP and proximity ligation in native tissue and cell system with functional readout; single lab","pmids":["32482756"],"is_preprint":false},{"year":2014,"finding":"SLC10A1 deficiency (NTCP deficiency) was identified as an inborn error of metabolism in a patient with a homozygous R252H mutation. Functional studies showed markedly reduced taurocholate uptake activity, and immunofluorescence/surface biotinylation showed the mutant protein is virtually absent from the plasma membrane, confirming NTCP is the main hepatic import system for conjugated bile salts.","method":"SLC10A1 sequencing, taurocholic acid uptake assay, immunofluorescence, surface biotinylation","journal":"Hepatology (Baltimore, Md.)","confidence":"High","confidence_rationale":"Tier 2 / Strong — human genetic disease with functional and localization validation; multiple orthogonal methods","pmids":["24867799"],"is_preprint":false},{"year":2019,"finding":"Cyclin D1 transcriptionally suppresses NTCP expression by inhibiting NTCP promoter activity during cell cycle progression. Ectopic expression of NTCP in HepG2 and Huh-7 cells suppressed hepatocyte growth by arresting cells in G0/G1 phase, identifying a functional link between NTCP expression and cell cycle regulation.","method":"NTCP promoter luciferase assay, cyclin D1 overexpression, cell cycle analysis (flow cytometry), ectopic NTCP expression in HCC cell lines","journal":"Oncotarget","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — promoter assay and gain-of-function with cell cycle readout; single lab","pmids":["28915572"],"is_preprint":false},{"year":2020,"finding":"HBV uptake into HepG2-NTCP cells is dependent on the actin cytoskeleton and occurs via clathrin-mediated endocytosis. HBV internalisation was inhibited by pitstop-2 and siRNA silencing of clathrin heavy chain, adaptor protein AP-2, and dynamin-2. Entry via caveolae or macropinocytosis was excluded. HBV entry in clathrin-coated pits was visualised by electron microscopy and cryo-EM with immunogold labelling.","method":"siRNA knockdown of clathrin, AP-2, dynamin-2; pharmacological inhibitors; electron microscopy; cryo-EM immunogold labelling in HepG2-NTCP cells","journal":"Cellular microbiology","confidence":"High","confidence_rationale":"Tier 2 / Strong — RNAi and pharmacological inhibitors with ultrastructural visualization; multiple orthogonal approaches","pmids":["32216005"],"is_preprint":false},{"year":2019,"finding":"NTCP genetic deletion in mice reduces diet-induced obesity, attenuates hepatic steatosis, and lowers plasma cholesterol by prolonging postprandial plasma bile acid elevations. The effect on obesity protection is TGR5-independent (NTCP/TGR5 double knockout mice showed equal protection). Mechanistically, NTCP knockout was associated with decreased intestinal fat absorption and increased uncoupled respiration in brown adipose tissue.","method":"NTCP knockout mice, NTCP/TGR5 double knockout mice, metabolic phenotyping, intestinal fat absorption assay, brown adipose tissue respiration measurement","journal":"JCI insight","confidence":"High","confidence_rationale":"Tier 2 / Strong — genetic knockout with defined metabolic phenotype; double KO epistasis demonstrates TGR5 independence","pmids":["31237863"],"is_preprint":false},{"year":2019,"finding":"Myrcludex B (bulevirtide) binds NTCP with very high affinity, and the NTCP:Myrcludex B interaction is extremely long-lived. Pre-bound Myrcludex B can transfer from one NTCP molecule to newly synthesized NTCP, partly escaping co-degradation. The normalization of plasma bile salt levels correlates with NTCP protein turnover rate.","method":"FITC-labeled Myrcludex B tracking, biotin-labeled NTCP turnover assay, FRET by fluorescence lifetime imaging microscopy in U2OS cells","journal":"JHEP reports","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — FRET-FLIM and pulse-chase protein turnover; single lab, direct binding dynamics","pmids":["32039379"],"is_preprint":false},{"year":2011,"finding":"The A64T polymorphism of NTCP (SLC10A1) significantly decreased transport of both taurocholate and rosuvastatin compared to wild-type. The S267F polymorphism showed decreased taurocholate uptake but increased rosuvastatin uptake (substrate-dependent functional change), demonstrating position 267 as a substrate selectivity determinant.","method":"Stable transfection of NTCP variants, taurocholate and rosuvastatin uptake assays in transfected cells","journal":"Xenobiotica","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — functional characterization of naturally occurring variants with multiple substrates; single lab","pmids":["21341987"],"is_preprint":false},{"year":1999,"finding":"Two alternatively spliced isoforms of mouse Ntcp (Ntcp1 encoding 362 aa and Ntcp2 encoding 317 aa with a shorter C-terminus from intron retention) both mediate saturable Na+-dependent transport of taurocholate when expressed in Xenopus oocytes.","method":"cDNA library screening, Xenopus oocyte expression, taurocholate uptake assay","journal":"Biochimica et biophysica acta","confidence":"High","confidence_rationale":"Tier 1 / Strong — functional reconstitution in Xenopus oocytes with saturable kinetics for both isoforms","pmids":["10209268"],"is_preprint":false},{"year":2020,"finding":"Position S267 in NTCP functions as an evolutionary 'rheostat': all 20 amino acid substitutions at this position produce progressive, varied effects on transport kinetics (Km and Vmax) and substrate specificity for taurocholic acid, estrone-3-sulfate, and rosuvastatin, rather than simple on/off effects. Stability modeling showed substitution tolerance correlates with surface expression but not transport activity.","method":"Systematic mutagenesis of all 20 amino acids at position 267, transport assays with three substrates, surface expression analysis, Rosetta stability modeling","journal":"The Journal of biological chemistry","confidence":"High","confidence_rationale":"Tier 1-2 / Strong — comprehensive systematic mutagenesis with multiple substrates and stability modeling; mechanistically informative","pmids":["33168628"],"is_preprint":false}],"current_model":"SLC10A1/NTCP is an electrogenic, sodium-dependent bile acid cotransporter localized to the basolateral membrane of hepatocytes, where it functions as the primary importer of conjugated bile acids from portal blood into hepatocytes; its cryo-EM structure reveals a transmembrane tunnel through which bile acids and the HBV preS1 domain compete for access, NTCP undergoes regulated trafficking to and from the plasma membrane controlled by PI3K/PKB/Akt signaling, PP2B-mediated dephosphorylation of Ser-226, PKCζ-dependent microtubule-based vesicle transport, clathrin-dependent endocytosis triggered by a dileucine motif, and ubiquitin-proteasome-mediated ERAD; its expression is transcriptionally regulated by RXR:RAR, HNF1α, HNF4α, RAR, and cyclin D1, and post-translationally regulated by N-linked glycosylation (required for plasma membrane targeting) and S-nitrosylation (inhibitory); NTCP also serves as the high-affinity receptor for HBV and HDV entry via the myristoylated preS1 domain, which blocks the extracellular entrance to the bile acid transport tunnel, with viral internalization requiring NTCP oligomerization downstream of EGFR co-receptor interaction and proceeding through clathrin-mediated endocytosis."},"narrative":{"mechanistic_narrative":"SLC10A1 (NTCP) is the principal sodium-dependent transporter that imports conjugated bile acids into hepatocytes, functioning at the rate-limiting basolateral uptake step of the enterohepatic bile salt cycle [PMID:16474011, PMID:24867799]. It transports taurine and glycine conjugates in preference to unconjugated bile salts, and when reconstituted with the apical exporter BSEP it reconstitutes vectorial basolateral-to-apical bile acid flux across polarized epithelium [PMID:16474011]. Cryo-EM structures show a transmembrane tunnel forming the substrate translocation route; residue 267 lies within a region critical for bile acid recognition, where substitutions act as a graded rheostat over transport kinetics and substrate selectivity while sparing the non-bile-acid substrate estrone sulfate [PMID:35580629, PMID:14660639, PMID:33168628]. NTCP activity at the plasma membrane is set by regulated trafficking: PI3K/PKB(Akt) signaling and PP2B(calcineurin)-mediated dephosphorylation of Ser-226 promote membrane insertion, PKCζ drives microtubule-based motility of NTCP recycling vesicles, a dileucine motif and PKC-triggered clathrin-dependent endocytosis remove it, and misfolded NTCP is cleared by ubiquitin-proteasome ERAD [PMID:16027164, PMID:12065290, PMID:12034724, PMID:16734659, PMID:24008362, PMID:16218878]. N-linked glycosylation is required for plasma membrane targeting, and S-nitrosylation of cysteines non-competitively inhibits transport [PMID:28125599, PMID:21109590]. Transcription of the gene is controlled by HNF1α, HNF4α, RXR:RAR and RAR, and is repressed in cholestasis through bile-acid/FXR-dependent loss of these activators and by inflammatory JNK signaling [PMID:14701722, PMID:25550158, PMID:16002565, PMID:12105223]. Independently of its metabolic role, NTCP is the high-affinity hepatocyte entry receptor for HBV and HDV: the myristoylated preS1 domain of the HBV large surface protein binds the extracellular mouth of the transport tunnel in direct competition with bile acid substrate, and viral internalization requires EGFR-dependent NTCP oligomerization followed by clathrin-mediated endocytosis [PMID:35580629, PMID:24342612, PMID:24845614, PMID:34613794, PMID:32216005]. Homozygous loss-of-function mutation (R252H) causes NTCP deficiency, an inborn error of bile salt metabolism with markedly reduced bile acid uptake [PMID:24867799].","teleology":[{"year":1999,"claim":"Establishing that NTCP intrinsically carries out sodium-dependent bile acid transport defined its core molecular activity independent of cellular context.","evidence":"Functional expression of two mouse Ntcp splice isoforms in Xenopus oocytes with saturable taurocholate uptake","pmids":["10209268"],"confidence":"High","gaps":["Oocyte system does not address polarized hepatocyte localization","Did not define substrate-recognition residues"]},{"year":2002,"claim":"Identifying PP2B-mediated dephosphorylation and PI3K/PKB signaling as drivers of NTCP membrane translocation showed that transporter activity is acutely controlled by regulated trafficking rather than expression alone.","evidence":"PP2B inhibition and immunoprecipitated-NTCP dephosphorylation in rat hepatocytes; dominant-negative/constitutively active PKB with translocation and uptake readouts","pmids":["12065290","12034724"],"confidence":"High","gaps":["Did not identify the target phosphosite","Upstream stimulus integration unresolved at this stage"]},{"year":2002,"claim":"Showing that inflammatory signaling represses Ntcp transcription via JNK-phosphorylated RXR linked NTCP downregulation to the hepatic response to cytokines and cholestasis.","evidence":"Ntcp promoter luciferase, dominant-negative JNK, EMSA in primary rat hepatocytes; SHP-1 induction timing in bile-duct-ligated mice","pmids":["12105223","11751172"],"confidence":"High","gaps":["SHP-1 link was temporal correlation only","Did not separate cytokine vs bile-acid contributions"]},{"year":2003,"claim":"Mapping species-specific and conserved transcription factor inputs (HNF1α, HNF4α, RXR/RAR, HNF3β, C/EBPβ) defined the regulatory architecture of the NTCP promoter.","evidence":"Cross-species luciferase reporter and EMSA assays in Huh7 cells","pmids":["14701722"],"confidence":"High","gaps":["Species divergence complicates extrapolation to human regulation","In vivo occupancy not established here"]},{"year":2003,"claim":"Demonstrating that the S267F polymorphism abolishes bile acid uptake while sparing estrone sulfate and membrane expression localized substrate recognition to a discrete region.","evidence":"Variant transport assays with multiple substrates plus cell-surface biotinylation in HepG2/transfected cells","pmids":["14660639"],"confidence":"High","gaps":["Atomic basis of substrate selectivity not yet resolved","Physiological consequence in carriers not addressed"]},{"year":2005,"claim":"Pinpointing Ser-226 as the cAMP-regulated phosphosite governing membrane retention connected the kinase/phosphatase circuitry to a defined molecular switch.","evidence":"32P labeling, clostripain mapping, S226A mutagenesis with uptake and surface expression in HuH-7 cells","pmids":["16027164"],"confidence":"High","gaps":["Kinase phosphorylating Ser-226 not directly identified","Interplay with the dileucine endocytosis motif not yet defined"]},{"year":2005,"claim":"Establishing FXR-dependent, bile-acid-mediated (not cytokine-mediated) repression of Ntcp in obstructive cholestasis resolved the dominant in vivo control mechanism.","evidence":"FXR knockout mice, bile duct ligation, cholic acid feeding, LPS, EMSA; plus Kupffer-cell depletion and cytokine neutralization controls","pmids":["16002565","15629514","15723437"],"confidence":"High","gaps":["FXR-to-HNF axis intermediate steps incomplete","LPS/inflammatory repression operates by a distinct, less-defined pathway"]},{"year":2005,"claim":"Identifying ERAD/ubiquitin-proteasome clearance of misfolded NTCP defined the protein's quality-control degradation route.","evidence":"Proteasome inhibitors, polyubiquitination assay, glycosylation analysis and MTOC aggresome colocalization in HepG2 cells","pmids":["16218878"],"confidence":"High","gaps":["Specific E3 ligase not identified","Relationship to surface-pool turnover unresolved"]},{"year":2006,"claim":"Reconstituting vectorial bile acid transport with NTCP and BSEP, and defining PKCζ-dependent vesicular motility, established NTCP's role in transcellular flux and its trafficking machinery.","evidence":"Double-transfected polarized LLC-PK1 transcellular flux; live imaging of NTCP-GFP vesicles with PKCζ inhibition and motor/endosome markers","pmids":["16474011","16734659"],"confidence":"High","gaps":["Coupling of motility to specific signaling stimuli incomplete","Kinesin/dynein switching control not defined"]},{"year":2010,"claim":"Showing S-nitrosylation of NTCP cysteines causes reversible non-competitive transport inhibition added a redox-based post-translational control layer.","evidence":"Biotin switch assay, kinetic uptake, surface biotinylation, DTT rescue in HuH-NTCP cells","pmids":["21109590"],"confidence":"High","gaps":["Modified cysteine residues not individually mapped","Physiological NO source not defined"]},{"year":2013,"claim":"Defining a dileucine motif with dual endocytosis/membrane-targeting roles and clathrin-dependent PKC-triggered internalization completed the retrieval arm of NTCP trafficking.","evidence":"Mutagenesis of dileucine motif and Thr225/Ser226, flow cytometry, immunofluorescence in HepG2 cells","pmids":["24008362"],"confidence":"High","gaps":["Adaptor recognizing the motif not identified","Reconciliation with Ser-226 dephosphorylation switch incomplete"]},{"year":2013,"claim":"Confirming NTCP as the functional HBV entry receptor and mapping cyclosporin A action to the preS1-binding site established the receptor function and its overlap with transport.","evidence":"NTCP overexpression/knockdown HBV infection assays; cyclosporin A inhibition, preS1 binding, and transport-competent/binding-deficient variants","pmids":["24342612","24295872"],"confidence":"High","gaps":["Post-attachment internalization steps not yet defined","Co-receptor requirements unresolved at this stage"]},{"year":2014,"claim":"Demonstrating reciprocal competition between preS1 and bile acids, and that R252H loss-of-function causes human NTCP deficiency, tied the viral and metabolic functions to a shared site and proved NTCP's role as the main hepatic bile salt importer.","evidence":"Reciprocal inhibition kinetics in primary hepatocytes and cell lines; patient sequencing with uptake assay and surface localization","pmids":["24845614","24867799"],"confidence":"High","gaps":["Spectrum of clinical phenotypes in deficiency not detailed","Quantitative overlap of viral vs substrate sites not structurally resolved here"]},{"year":2014,"claim":"Identifying RAR control of the human NTCP promoter linked nuclear receptor signaling to both transporter expression and HBV permissiveness.","evidence":"Promoter luciferase, RAR ChIP, RAR antagonists with HBV infection in HepG2-hNTCP","pmids":["25550158"],"confidence":"Medium","gaps":["Single lab; in vivo relevance not established","Endogenous RAR ligand context unclear"]},{"year":2017,"claim":"Establishing that N-linked glycosylation at N5/N11 is required for membrane trafficking and HBV infection clarified a biosynthetic prerequisite for both functions.","evidence":"Glycosylation-site mutagenesis, surface biotinylation, lysosomal degradation and HBV/uptake assays in HepG2 cells","pmids":["28125599"],"confidence":"High","gaps":["Glycan-dependent chaperone interactions not identified","Effect on oligomerization not tested"]},{"year":2020,"claim":"Systematic saturation mutagenesis at position 267 reframed substrate recognition as a tunable rheostat coupling stability/expression and substrate-specific kinetics.","evidence":"All-20-amino-acid substitution with three substrates, surface expression, and Rosetta stability modeling; prior A64T/S267F variant kinetics","pmids":["33168628","21341987"],"confidence":"High","gaps":["Structural rationalization of rheostat behavior incomplete","Population-level pharmacogenetic impact not assessed"]},{"year":2021,"claim":"Resolving that NTCP oligomerization downstream of EGFR is required for HBV internalization, and defining clathrin-dependent uptake, established the ordered post-attachment entry mechanism.","evidence":"SPR, F274A mutagenesis, CoIP, oligomerization assays with troglitazone; clathrin/AP-2/dynamin-2 knockdown and EM in HepG2-NTCP","pmids":["34613794","32216005"],"confidence":"High","gaps":["Stoichiometry of the oligomer not defined","How EGFR engagement triggers oligomerization unresolved"]},{"year":2022,"claim":"Cryo-EM structures of NTCP, alone and with preS1, provided the atomic framework for the bile acid transport tunnel and the structural basis of viral receptor engagement.","evidence":"Cryo-EM of human/bovine/rat NTCP and antibody/preS1 complexes with mutation and transport assays","pmids":["35580629","35580630"],"confidence":"High","gaps":["Substrate-bound and intermediate conformational states not captured","Sodium coupling mechanism not fully detailed"]},{"year":2022,"claim":"Identifying IFITM3 as an NTCP interaction partner facilitating a post-attachment HBV/HDV entry step expanded the receptor complex beyond NTCP itself.","evidence":"Membrane yeast-two-hybrid, CoIP, IFITM3 knockdown with HBV/HDV infection and preS1 binding in HuH7 and primary hepatocytes","pmids":["35458456"],"confidence":"Medium","gaps":["Single lab; mechanism of IFITM3 action at the entry step undefined","Direct vs indirect interaction not fully resolved"]},{"year":2024,"claim":"The bulevirtide-bound NTCP structure explained how the therapeutic peptide plugs the transport tunnel and underlies HBV/HDV host specificity.","evidence":"Cryo-EM of BLV-bound human NTCP; prior high-affinity, long-lived binding/turnover measurements","pmids":["38509088","32039379"],"confidence":"High","gaps":["In vivo pharmacodynamic linkage to NTCP turnover only correlative","Resistance mechanisms not addressed"]},{"year":null,"claim":"How NTCP's bile-acid transport function mechanistically intersects with broader physiology—innate antiviral immunity, cell-cycle control, and systemic metabolism—remains incompletely defined.","evidence":"Open question integrating NTCP/ISG repression, cyclin D1 promoter suppression, and knockout metabolic phenotypes","pmids":[],"confidence":"Medium","gaps":["Causal chain from bile-acid transport to ISG/IFITM3 repression not fully mapped","Mechanism coupling NTCP expression to G0/G1 arrest unresolved","Tissue-level metabolic effects of NTCP loss in humans not characterized"]}],"mechanism_profile":{"molecular_activity":[{"term_id":"GO:0005215","term_label":"transporter activity","supporting_discovery_ids":[18,30,36,20,37]},{"term_id":"GO:0140104","term_label":"molecular carrier activity","supporting_discovery_ids":[18,36]},{"term_id":"GO:0001618","term_label":"virus receptor activity","supporting_discovery_ids":[3,4,5,0]},{"term_id":"GO:0038024","term_label":"cargo receptor activity","supporting_discovery_ids":[17]}],"localization":[{"term_id":"GO:0005886","term_label":"plasma membrane","supporting_discovery_ids":[6,8,18,19,30]},{"term_id":"GO:0005768","term_label":"endosome","supporting_discovery_ids":[9,21]},{"term_id":"GO:0005783","term_label":"endoplasmic reticulum","supporting_discovery_ids":[10,19]},{"term_id":"GO:0005815","term_label":"microtubule organizing center","supporting_discovery_ids":[10]}],"pathway":[{"term_id":"R-HSA-382551","term_label":"Transport of small molecules","supporting_discovery_ids":[18,30,36]},{"term_id":"R-HSA-1430728","term_label":"Metabolism","supporting_discovery_ids":[18,33]},{"term_id":"R-HSA-1643685","term_label":"Disease","supporting_discovery_ids":[3,30,5]},{"term_id":"R-HSA-168256","term_label":"Immune System","supporting_discovery_ids":[24,26,28]},{"term_id":"R-HSA-74160","term_label":"Gene expression (Transcription)","supporting_discovery_ids":[11,12,25,14]}],"complexes":[],"partners":["BSEP","IFITM3","OATP1B3","EGFR"],"other_free_text":[]}},"prefetch_data":{"uniprot":{"accession":"Q14973","full_name":"Hepatic sodium/bile acid cotransporter","aliases":["Cell growth-inhibiting gene 29 protein","Na(+)/bile acid cotransporter","Na(+)/taurocholate transport protein","Sodium/taurocholate cotransporting polypeptide","NTCP","Solute carrier family 10 member 1","SLC10A1"],"length_aa":349,"mass_kda":38.1,"function":"As a major transporter of conjugated bile salts from plasma into the hepatocyte, it plays a key role in the enterohepatic circulation of bile salts necessary for the solubilization and absorption of dietary fat and fat-soluble vitamins (PubMed:14660639, PubMed:24867799, PubMed:34060352, PubMed:8132774). It is strictly dependent on the extracellular presence of sodium (PubMed:14660639, PubMed:24867799, PubMed:34060352, PubMed:8132774). It exhibits broad substrate specificity and transports various bile acids, such as taurocholate, cholate, as well as non-bile acid organic compounds, such as estrone sulfate (PubMed:14660639, PubMed:34060352). Works collaboratively with the ileal transporter (NTCP2), the organic solute transporter (OST), and the bile salt export pump (BSEP), to ensure efficacious biological recycling of bile acids during enterohepatic circulation (PubMed:33222321) (Microbial infection) Acts as an entry receptor for hepatitis B virus (HBV) (PubMed:23150796). The recognition for human SLC10A1/NTCP is highly specific (PubMed:35545671, PubMed:35580629, PubMed:35580630, PubMed:38233573)","subcellular_location":"Cell membrane","url":"https://www.uniprot.org/uniprotkb/Q14973/entry"},"depmap":{"release":"DepMap","has_data":true,"is_common_essential":false,"resolved_as":"","url":"https://depmap.org/portal/gene/SLC10A1","classification":"Not Classified","n_dependent_lines":1,"n_total_lines":1208,"dependency_fraction":0.0008278145695364238},"opencell":{"profiled":false,"resolved_as":"","ensg_id":"","cell_line_id":"","localizations":[],"interactors":[],"url":"https://opencell.sf.czbiohub.org/search/SLC10A1","total_profiled":1310},"omim":[{"mim_id":"619874","title":"CHOLESTASIS, PROGRESSIVE FAMILIAL INTRAHEPATIC, 11; PFIC11","url":"https://www.omim.org/entry/619874"},{"mim_id":"619256","title":"HYPERCHOLANEMIA, FAMILIAL, 2; FHCA2","url":"https://www.omim.org/entry/619256"},{"mim_id":"610424","title":"HEPATITIS B VIRUS, SUSCEPTIBILITY TO","url":"https://www.omim.org/entry/610424"},{"mim_id":"607961","title":"SEMAPHORIN 7A; SEMA7A","url":"https://www.omim.org/entry/607961"},{"mim_id":"607748","title":"HYPERCHOLANEMIA, FAMILIAL 1; FHCA1","url":"https://www.omim.org/entry/607748"}],"hpa":{"profiled":true,"resolved_as":"","reliability":"","locations":[],"tissue_specificity":"Tissue enriched","tissue_distribution":"Detected in single","driving_tissues":[{"tissue":"liver","ntpm":399.1}],"url":"https://www.proteinatlas.org/search/SLC10A1"},"hgnc":{"alias_symbol":["NTCP","NTCP1"],"prev_symbol":[]},"alphafold":{"accession":"Q14973","domains":[{"cath_id":"1.20.1530.20","chopping":"23-309","consensus_level":"high","plddt":90.7295,"start":23,"end":309}],"viewer_url":"https://alphafold.ebi.ac.uk/entry/Q14973","model_url":"https://alphafold.ebi.ac.uk/files/AF-Q14973-F1-model_v6.cif","pae_url":"https://alphafold.ebi.ac.uk/files/AF-Q14973-F1-predicted_aligned_error_v6.png","plddt_mean":83.5},"mouse_models":{"mgi_url":"https://www.informatics.jax.org/marker/summary?nomen=SLC10A1","jax_strain_url":"https://www.jax.org/strain/search?query=SLC10A1"},"sequence":{"accession":"Q14973","fasta_url":"https://rest.uniprot.org/uniprotkb/Q14973.fasta","uniprot_url":"https://www.uniprot.org/uniprotkb/Q14973/entry","alphafold_viewer_url":"https://alphafold.ebi.ac.uk/entry/Q14973"}},"corpus_meta":[{"pmid":"24342612","id":"PMC_24342612","title":"Evaluation 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The structure of human NTCP in complex with the myristoylated preS1 domain of the HBV large surface protein, together with mutation and transport assays, shows that preS1 and the bile acid substrate compete for the extracellular opening of the tunnel, establishing the binding mode for viral entry.\",\n      \"method\": \"Cryo-electron microscopy, mutation assays, transport assays\",\n      \"journal\": \"Nature\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 / Strong — cryo-EM structure with mutagenesis and functional transport assays in a single rigorous study; independently reported by two concurrent Nature papers\",\n      \"pmids\": [\"35580629\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2022,\n      \"finding\": \"Cryo-EM structure of NTCP bound to an antibody shows the transporter lacks the first transmembrane helix found in other SLC10 proteins and has an N-terminus exposed on the extracellular face. Comparison with related proteins indicates a common mechanism of bile acid transport, and an additional pocket formed by residues known to interact with preS1 was identified.\",\n      \"method\": \"Cryo-electron microscopy, structural comparison\",\n      \"journal\": \"Nature\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 / Strong — cryo-EM structure; independently replicated by concurrent parallel paper (PMID:35580629)\",\n      \"pmids\": [\"35580630\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2024,\n      \"finding\": \"Cryo-EM structure of bulevirtide (BLV/Myrcludex B)-bound human NTCP reveals BLV forms two domains: a plug lodged in the bile salt transport tunnel of NTCP and a string covering the extracellular surface, with the N-terminal myristoyl group interacting with the lipid-exposed surface of NTCP. This structure explains how BLV blocks bile salt transport and provides a structural basis for HBV/HDV host specificity.\",\n      \"method\": \"Cryo-electron microscopy\",\n      \"journal\": \"Nature communications\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 / Strong — atomic-resolution cryo-EM structure with mechanistic interpretation; single rigorous study with multiple orthogonal validations implied by structural detail\",\n      \"pmids\": [\"38509088\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2013,\n      \"finding\": \"NTCP (SLC10A1) was confirmed as the functional HBV entry receptor: HepG2 cells engineered to overexpress human NTCP became susceptible to HBV infection, and knockdown of NTCP blocked infection. Compounds that inhibit NTCP transporter activity (including cyclosporin A and oxysterols) also blocked HBV infection, linking bile acid transport function to viral entry.\",\n      \"method\": \"NTCP overexpression in HepG2 cells, siRNA knockdown, HBV infection assay, inhibitor studies\",\n      \"journal\": \"Biochemical and biophysical research communications\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — gain-of-function and loss-of-function with defined phenotypic readout; replicated across multiple labs\",\n      \"pmids\": [\"24342612\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2013,\n      \"finding\": \"Cyclosporin A inhibits HBV and HDV entry by directly interfering with NTCP receptor function in a cyclophilin-independent manner. Binding of the HBVpreS1 domain to NTCP was blocked by cyclosporin A, and an NTCP variant deficient in HBVpreS1 binding but competent for bile salt transport was resistant to cyclosporin A, mapping the interaction site to the preS1-binding domain on NTCP.\",\n      \"method\": \"HepaRG cells and NTCP-expressing hepatoma cell lines, taurocholate uptake assay, HBVpreS1 binding assay, cyclophilin siRNA, NTCP variant studies\",\n      \"journal\": \"Journal of hepatology\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — multiple orthogonal methods (transport assay, binding assay, RNAi, variant rescue), replicated across multiple cell models\",\n      \"pmids\": [\"24295872\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2014,\n      \"finding\": \"The myristoylated preS1 domain of HBV inhibits bile acid transport by NTCP, and inversely, bile acid conjugates (taurine/glycine conjugates of cholic acid and ursodeoxycholic acid) inhibit HBV infection in a concentration-dependent manner. NTCP expression, transport function, preS1 peptide binding, and HBV infection follow comparable kinetics, establishing direct functional overlap between the bile acid transport site and the HBV receptor site.\",\n      \"method\": \"Bile acid transport assays, HBV infection assays, myr-preS1 peptide binding in primary hepatocytes and NTCP-transfected cells\",\n      \"journal\": \"Journal of hepatology\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — reciprocal inhibition demonstrated in primary human hepatocytes and multiple engineered cell lines with quantitative kinetics\",\n      \"pmids\": [\"24845614\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2005,\n      \"finding\": \"Dephosphorylation of Ser-226 in the third cytoplasmic loop of NTCP facilitates plasma membrane retention. Mutation of Ser-226 to Ala decreased NTCP phosphorylation by 30% and increased taurocholate uptake and plasma membrane retention 2.5–3.2-fold; cAMP failed to further increase translocation of S226A-NTCP, establishing Ser-226 as the cAMP-regulated phosphorylation site controlling NTCP plasma membrane localization.\",\n      \"method\": \"Metabolic [32P] labeling, clostripain digestion, site-directed mutagenesis, taurocholate uptake assay, cell surface expression in transfected HuH-7 cells\",\n      \"journal\": \"The Journal of biological chemistry\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 / Strong — site-directed mutagenesis combined with phosphorylation mapping and functional transport assays in the same study\",\n      \"pmids\": [\"16027164\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2002,\n      \"finding\": \"PP2B (calcineurin), a Ca2+/calmodulin-dependent phosphatase, mediates cAMP-induced dephosphorylation and translocation of NTCP to the plasma membrane. The PP2B inhibitor cypermethrin reversed cAMP-mediated NTCP dephosphorylation and translocation, and PP2B directly dephosphorylated immunoprecipitated NTCP from control but not cAMP-treated hepatocytes.\",\n      \"method\": \"PP2B inhibitors (cypermethrin, FK-506), taurocholate uptake assay, NTCP immunoprecipitation, PP2B activity assay in isolated rat hepatocytes\",\n      \"journal\": \"American journal of physiology. Gastrointestinal and liver physiology\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 / Strong — in vitro dephosphorylation of immunoprecipitated NTCP by PP2B, pharmacological inhibition with multiple compounds, and functional readout\",\n      \"pmids\": [\"12065290\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2002,\n      \"finding\": \"Protein kinase B (PKB/Akt) mediates cAMP- and cell swelling-stimulated Na+/taurocholate cotransport and NTCP translocation to the plasma membrane via the PI3K/PKB signaling pathway. Dominant-negative PKB blocked cAMP- and swelling-induced increases in TC uptake and NTCP translocation, while constitutively active PKB increased both.\",\n      \"method\": \"Dominant-negative and constitutively active PKB transfection, taurocholate uptake assay, NTCP translocation assay in HuH-Ntcp cells\",\n      \"journal\": \"The Journal of biological chemistry\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — genetic dominant-negative and constitutively active constructs with functional readout; multiple stimuli tested\",\n      \"pmids\": [\"12034724\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2006,\n      \"finding\": \"NTCP-containing vesicles are present on intracellular recycling endosomes and move bidirectionally on microtubules using kinesin-1 and dynein motors. PKCζ is specifically required for microtubule-based motility of NTCP vesicles: PI(3,4,5)P3 activates PKCζ and enhances motility, while specific inhibition of PKCζ blocks motility of NTCP-containing vesicles but not late vesicles.\",\n      \"method\": \"In vitro and whole-cell immunofluorescence microscopy, live-cell imaging of NTCP-GFP, PKCζ inhibitor, colocalization with endosomal markers\",\n      \"journal\": \"Traffic (Copenhagen, Denmark)\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — direct live-imaging and in vitro motility assays with pharmacological and genetic inhibition; functional consequence demonstrated\",\n      \"pmids\": [\"16734659\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2005,\n      \"finding\": \"Rat Ntcp/human NTCP is degraded by the ubiquitin-proteasome system via ER-associated degradation (ERAD). Proteasome inhibition led to accumulation of core-glycosylated (ER-resident) intracellular NTCP that co-localized with ubiquitin at the microtubule organization center as polyubiquitinated aggresomes.\",\n      \"method\": \"Proteasome inhibitors (MG-132, lactacystin), co-localization studies, polyubiquitination assay, glycosylation analysis in stably transfected HepG2 cells\",\n      \"journal\": \"Biological chemistry\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — multiple pharmacological inhibitors, biochemical characterization of ubiquitinated species, and glycosylation state analysis\",\n      \"pmids\": [\"16218878\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2002,\n      \"finding\": \"Interleukin-1β suppresses Ntcp gene expression via a JNK-dependent mechanism: JNK phosphorylates RXR, reducing nuclear RXR:RAR binding activity and thereby decreasing RXR:RAR-mediated transactivation of the Ntcp promoter. JNK inhibition by curcumin or dominant-negative JNK completely blocked IL-1β-mediated suppression, while ERK and p38 inhibitors had no effect.\",\n      \"method\": \"Ntcp promoter luciferase assays, JNK expression plasmids (dominant-negative and wild-type), pharmacological inhibitors, EMSA, primary rat hepatocytes\",\n      \"journal\": \"The Journal of biological chemistry\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — multiple orthogonal methods (promoter assay, dominant-negative JNK, pharmacological inhibitors, EMSA); confirmed in primary hepatocytes\",\n      \"pmids\": [\"12105223\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2003,\n      \"finding\": \"HNF1α, HNF4α, and RXRα/RARα activate the rat Ntcp promoter but not the human or mouse NTCP/Ntcp promoters, whereas C/EBPβ activation is specific to human and mouse. HNF3β is the only transcription factor binding and active at a conserved motif in all three species, where it inhibits NTCP/Ntcp promoter activity. Small heterodimer partner (SHP) does not affect NTCP/Ntcp promoter activity from the conserved region.\",\n      \"method\": \"Transfection-based luciferase reporter assays, electrophoretic mobility shift assay (EMSA), cotransfection in Huh7 cells\",\n      \"journal\": \"American journal of physiology. Gastrointestinal and liver physiology\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — EMSA and promoter luciferase assays across three species with multiple transcription factors\",\n      \"pmids\": [\"14701722\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2002,\n      \"finding\": \"Short heterodimer partner 1 (SHP-1) induction by retained bile acids temporally precedes and likely mediates downregulation of Ntcp in obstructive cholestasis. In bile duct-ligated mice, SHP-1 mRNA peaked at 6 h after ligation, followed 6 h later by the nadir of Ntcp mRNA and protein.\",\n      \"method\": \"Northern and Western blotting of SHP-1 and Ntcp in CBDL mice, immunofluorescence\",\n      \"journal\": \"American journal of physiology. Gastrointestinal and liver physiology\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 3 / Moderate — temporal correlation in a genetic knockout model without direct mechanistic intervention; confirmed in two genotypes (FXR+/+ and FXR-/-) in follow-up paper\",\n      \"pmids\": [\"11751172\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2005,\n      \"finding\": \"In obstructive cholestasis (CBDL), Ntcp downregulation is bile acid–mediated and FXR-dependent, not cytokine-mediated. FXR knockout mice fail to repress Ntcp after bile acid feeding or CBDL; LPS-induced Ntcp repression is FXR-independent. The mechanism involves reduced nuclear levels and DNA binding of HNF-1α, HNF-4α, RXRα, and RARα.\",\n      \"method\": \"FXR knockout mice, bile duct ligation, cholic acid feeding, LPS treatment, EMSA, Western blotting, cytokine inactivation\",\n      \"journal\": \"American journal of physiology. Gastrointestinal and liver physiology\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — genetic epistasis with FXR-/- mice using multiple interventions and multiple readouts\",\n      \"pmids\": [\"16002565\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2005,\n      \"finding\": \"Kupffer cell depletion with liposomal clodronate prevents endotoxin-induced suppression of Ntcp expression. In endotoxin-exposed rats, KC depletion preserved Ntcp RNA and protein expression and maintained RXR:RAR and HNF1α transcription factor binding activity.\",\n      \"method\": \"Liposomal clodronate KC depletion, LPS exposure, Northern/Western blotting, EMSA in rats\",\n      \"journal\": \"Journal of hepatology\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — depletion experiment with defined molecular readout (transcription factor binding), single lab\",\n      \"pmids\": [\"15629514\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2005,\n      \"finding\": \"In obstructive cholestasis in rodents, accumulating bile acids—independent of cytokines—downregulate Ntcp through repression of HNF-1α and HNF-4α. Both TNF-α/IL-1β neutralization and Kupffer cell depletion failed to restore Ntcp expression, while SHP mRNA increased 3–5-fold.\",\n      \"method\": \"Cytokine neutralization (etanercept, anakinra), KC depletion by liposome clodronate, Western/Northern blotting, EMSA in CBDL rats and mice\",\n      \"journal\": \"Hepatology (Baltimore, Md.)\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — pharmacological and cellular depletion with multiple molecular readouts; single lab\",\n      \"pmids\": [\"15723437\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2003,\n      \"finding\": \"NTCP (Ntcp) facilitates hepatocellular uptake of the lethal mushroom toxin α-amanitin. HepG2 cells stably transfected with rat Ntcp showed markedly increased sensitivity to α-amanitin cytotoxicity compared to non-transfected cells, as measured by suppression of cytokine-induced IL-1Ra mRNA synthesis.\",\n      \"method\": \"Stable transfection of rat Ntcp in HepG2 cells, taurocholate uptake kinetics, functional toxicity assay (IL-1Ra mRNA induction)\",\n      \"journal\": \"Archives of toxicology\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — gain-of-function with a defined functional assay; single lab\",\n      \"pmids\": [\"14598021\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2006,\n      \"finding\": \"Human NTCP and BSEP reconstituted together in LLC-PK1 polarized cells mediate vectorial transcellular transport of bile acids from the basolateral to apical side, with substrate specificities showing taurine conjugates > glycine conjugates > unconjugated bile salts for NTCP. Single-transporter cells showed no vectorial transport.\",\n      \"method\": \"Double transfection of LLC-PK1 cells with NTCP and BSEP, transcellular bile acid flux assays, kinetic analysis\",\n      \"journal\": \"American journal of physiology. Gastrointestinal and liver physiology\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 / Strong — reconstitution of vectorial transport in an epithelial cell system with kinetic characterization\",\n      \"pmids\": [\"16474011\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2017,\n      \"finding\": \"N-linked glycosylation of NTCP at residues N5 and N11 is required for efficient trafficking to the plasma membrane and for HBV infection. NTCP lacking both glycosylation sites (N5,11Q) failed to support HBV infection, showed minimal cellular expression, and was degraded in lysosomes, while single-glycan variants retained normal bile acid transport function and HBV infectability.\",\n      \"method\": \"Site-directed mutagenesis of glycosylation sites, cell surface biotinylation, lysosomal degradation assay, HBV infection assay, bile acid uptake assay in HepG2 cells\",\n      \"journal\": \"PloS one\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1-2 / Strong — mutagenesis combined with multiple functional assays and trafficking analysis\",\n      \"pmids\": [\"28125599\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2003,\n      \"finding\": \"The C800T (Ser267Phe) polymorphism in NTCP causes near-complete loss of bile acid uptake function while preserving transport of the non-bile acid substrate estrone sulfate, identifying position 267 as part of a region critical and specific for bile acid substrate recognition. Cell surface biotinylation showed normal plasma membrane expression of this variant.\",\n      \"method\": \"Cell surface biotinylation, taurocholate and estrone sulfate transport assays, immunofluorescence confocal microscopy in HepG2 and transfected cells\",\n      \"journal\": \"The Journal of biological chemistry\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — substrate-selective transport with normal membrane expression; multiple substrates and multiple cell models tested\",\n      \"pmids\": [\"14660639\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2013,\n      \"finding\": \"A dileucine motif in the third intracellular loop of Ntcp is essential for both endocytosis and plasma membrane targeting, indicating a dual trafficking function. PKC-mediated endocytosis of Ntcp from the plasma membrane is clathrin-dependent and is followed by lysosomal degradation. Mutation of Thr225 and Ser226 inhibited PKC-mediated endocytosis.\",\n      \"method\": \"Flow cytometry, immunofluorescence, Western blotting, site-directed mutagenesis of dileucine motif and phosphorylation sites in HepG2 cells\",\n      \"journal\": \"American journal of physiology. Gastrointestinal and liver physiology\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — mutagenesis of multiple residues with parallel functional and localization readouts\",\n      \"pmids\": [\"24008362\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2010,\n      \"finding\": \"Nitric oxide (NO) inhibits taurocholate uptake by NTCP via S-nitrosylation of cysteine residues on NTCP, causing non-competitive inhibition (decreased Vmax, unchanged Km) and reduction of NTCP at the plasma membrane. Dithiothreitol reversed both S-nitrosylation and transport inhibition.\",\n      \"method\": \"Biotin switch assay for S-nitrosylation, taurocholate kinetic uptake assay, plasma membrane biotinylation, NO donor treatment in HuH-NTCP cells\",\n      \"journal\": \"American journal of physiology. Gastrointestinal and liver physiology\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1-2 / Strong — S-nitrosylation established by biotin switch with mechanistic rescue by dithiothreitol; kinetic analysis performed\",\n      \"pmids\": [\"21109590\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2012,\n      \"finding\": \"Plasma membrane localization of PKCδ, rather than its kinase activity, is necessary for cAMP-induced NTCP translocation and Rab4 activation. Kinase-dead dominant-negative PKCδ and PKCδ knockdown both still increased plasma membrane NTCP and Rab4 activity when PKCδ was at the membrane. In contrast, kinase activity of PKCδ is required for cAMP-induced MRP2 translocation.\",\n      \"method\": \"Wild-type, kinase-dead DN-PKCδ transfection, siRNA knockdown, Rab4 activity assay, rottlerin and LY294002 inhibitors in HuH-NTCP cells\",\n      \"journal\": \"American journal of physiology. Gastrointestinal and liver physiology\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — genetic and pharmacological dissection with multiple constructs; single lab\",\n      \"pmids\": [\"22744337\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2015,\n      \"finding\": \"IL-6 inhibits HBV entry into hepatocytes by downregulating NTCP: IL-6 pretreatment reduced NTCP mRNA by 98%, reduced NTCP-mediated taurocholate uptake by 80%, and inhibited HBV entry by up to 90%. Restoration of NTCP expression suppressed the inhibitory effect of IL-6, confirming that NTCP downregulation mediates the effect.\",\n      \"method\": \"HBV infection assay (cccDNA and HBsAg readouts), taurocholate uptake assay, NTCP mRNA quantification, NTCP restoration experiment\",\n      \"journal\": \"Virology\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — loss-of-function with rescue experiment and multiple readouts; dose-dependent kinetics demonstrated\",\n      \"pmids\": [\"25765005\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2014,\n      \"finding\": \"Retinoic acid receptor (RAR) regulates human NTCP promoter activity via a binding site at nucleotides −112 to −96 of the NTCP gene. Pharmacological antagonism of RAR (Ro41-5253, CD2665) reduced NTCP mRNA and protein expression, blocked viral entry, and inhibited HBV spread across multiple genotypes.\",\n      \"method\": \"NTCP promoter luciferase assays, RAR ChIP, pharmacological RAR antagonists, HBV infection assay, NTCP mRNA/protein quantification in HepG2-hNTCP cells\",\n      \"journal\": \"The Journal of biological chemistry\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — promoter mapping with ChIP and functional readout; single lab\",\n      \"pmids\": [\"25550158\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2016,\n      \"finding\": \"NTCP augments HCV infection by mediating bile-acid-dependent repression of interferon-stimulated genes (ISGs), including IFITM3. Gain- and loss-of-function studies in hepatocytes showed NTCP regulates innate antiviral immunity, with NTCP-mediated bile acid transport suppressing ISG expression and thereby increasing permissiveness to HCV infection.\",\n      \"method\": \"NTCP overexpression and knockdown in hepatoma cells, ISG expression profiling, HCV infection assay, bile acid treatment\",\n      \"journal\": \"Cell reports\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — gain- and loss-of-function with defined molecular mechanism (ISG repression); single lab\",\n      \"pmids\": [\"27783949\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2021,\n      \"finding\": \"NTCP oligomerization occurs downstream of the NTCP-EGFR interaction and is required for HBV internalization. Troglitazone directly binds NTCP allosterically (non-competitive inhibition of bile acid uptake) and blocks NTCP oligomerization, inhibiting HBV internalization without affecting HBV attachment or the NTCP-EGFR interaction. The F274A mutation in NTCP disrupts oligomerization and HBV internalization without affecting viral surface binding.\",\n      \"method\": \"Surface plasmon resonance, transporter kinetics, alanine scanning mutagenesis, coimmunoprecipitation, NTCP oligomerization assay in transfected cells\",\n      \"journal\": \"Journal of virology\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — multiple orthogonal methods (SPR, mutagenesis, CoIP, kinetics) dissecting an ordered entry mechanism\",\n      \"pmids\": [\"34613794\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2022,\n      \"finding\": \"IFITM3 is a protein-protein interaction partner of NTCP identified by membrane yeast-two-hybrid and confirmed by co-immunoprecipitation. IFITM3 knockdown significantly reduced HBV and HDV infection of NTCP-expressing HuH7 cells and primary human hepatocytes, despite intact myr-preS1 peptide binding to NTCP, indicating IFITM3 facilitates a post-attachment step of HBV/HDV entry.\",\n      \"method\": \"Membrane yeast-two-hybrid, co-immunoprecipitation, IFITM3 siRNA knockdown, HBV/HDV infection assays, myr-preS1 binding assay\",\n      \"journal\": \"Viruses\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2-3 / Moderate — membrane Y2H confirmed by CoIP with defined functional consequence in primary hepatocytes; single lab\",\n      \"pmids\": [\"35458456\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2019,\n      \"finding\": \"OATP1B3 forms hetero-oligomers with NTCP in both HEK293 cells and frozen human liver sections. Coexpression of NTCP with OATP1B3 in HEK293 cells increased OATP1B3 plasma membrane expression but decreased its apparent turnover rate for the substrate cholecystokinin-8.\",\n      \"method\": \"Co-immunoprecipitation, proximity ligation assay, plasma membrane expression quantification, transport assay in HEK293T cells and human liver sections\",\n      \"journal\": \"Drug metabolism and disposition\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2-3 / Moderate — CoIP and proximity ligation in native tissue and cell system with functional readout; single lab\",\n      \"pmids\": [\"32482756\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2014,\n      \"finding\": \"SLC10A1 deficiency (NTCP deficiency) was identified as an inborn error of metabolism in a patient with a homozygous R252H mutation. Functional studies showed markedly reduced taurocholate uptake activity, and immunofluorescence/surface biotinylation showed the mutant protein is virtually absent from the plasma membrane, confirming NTCP is the main hepatic import system for conjugated bile salts.\",\n      \"method\": \"SLC10A1 sequencing, taurocholic acid uptake assay, immunofluorescence, surface biotinylation\",\n      \"journal\": \"Hepatology (Baltimore, Md.)\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — human genetic disease with functional and localization validation; multiple orthogonal methods\",\n      \"pmids\": [\"24867799\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2019,\n      \"finding\": \"Cyclin D1 transcriptionally suppresses NTCP expression by inhibiting NTCP promoter activity during cell cycle progression. Ectopic expression of NTCP in HepG2 and Huh-7 cells suppressed hepatocyte growth by arresting cells in G0/G1 phase, identifying a functional link between NTCP expression and cell cycle regulation.\",\n      \"method\": \"NTCP promoter luciferase assay, cyclin D1 overexpression, cell cycle analysis (flow cytometry), ectopic NTCP expression in HCC cell lines\",\n      \"journal\": \"Oncotarget\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — promoter assay and gain-of-function with cell cycle readout; single lab\",\n      \"pmids\": [\"28915572\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2020,\n      \"finding\": \"HBV uptake into HepG2-NTCP cells is dependent on the actin cytoskeleton and occurs via clathrin-mediated endocytosis. HBV internalisation was inhibited by pitstop-2 and siRNA silencing of clathrin heavy chain, adaptor protein AP-2, and dynamin-2. Entry via caveolae or macropinocytosis was excluded. HBV entry in clathrin-coated pits was visualised by electron microscopy and cryo-EM with immunogold labelling.\",\n      \"method\": \"siRNA knockdown of clathrin, AP-2, dynamin-2; pharmacological inhibitors; electron microscopy; cryo-EM immunogold labelling in HepG2-NTCP cells\",\n      \"journal\": \"Cellular microbiology\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — RNAi and pharmacological inhibitors with ultrastructural visualization; multiple orthogonal approaches\",\n      \"pmids\": [\"32216005\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2019,\n      \"finding\": \"NTCP genetic deletion in mice reduces diet-induced obesity, attenuates hepatic steatosis, and lowers plasma cholesterol by prolonging postprandial plasma bile acid elevations. The effect on obesity protection is TGR5-independent (NTCP/TGR5 double knockout mice showed equal protection). Mechanistically, NTCP knockout was associated with decreased intestinal fat absorption and increased uncoupled respiration in brown adipose tissue.\",\n      \"method\": \"NTCP knockout mice, NTCP/TGR5 double knockout mice, metabolic phenotyping, intestinal fat absorption assay, brown adipose tissue respiration measurement\",\n      \"journal\": \"JCI insight\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — genetic knockout with defined metabolic phenotype; double KO epistasis demonstrates TGR5 independence\",\n      \"pmids\": [\"31237863\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2019,\n      \"finding\": \"Myrcludex B (bulevirtide) binds NTCP with very high affinity, and the NTCP:Myrcludex B interaction is extremely long-lived. Pre-bound Myrcludex B can transfer from one NTCP molecule to newly synthesized NTCP, partly escaping co-degradation. The normalization of plasma bile salt levels correlates with NTCP protein turnover rate.\",\n      \"method\": \"FITC-labeled Myrcludex B tracking, biotin-labeled NTCP turnover assay, FRET by fluorescence lifetime imaging microscopy in U2OS cells\",\n      \"journal\": \"JHEP reports\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — FRET-FLIM and pulse-chase protein turnover; single lab, direct binding dynamics\",\n      \"pmids\": [\"32039379\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2011,\n      \"finding\": \"The A64T polymorphism of NTCP (SLC10A1) significantly decreased transport of both taurocholate and rosuvastatin compared to wild-type. The S267F polymorphism showed decreased taurocholate uptake but increased rosuvastatin uptake (substrate-dependent functional change), demonstrating position 267 as a substrate selectivity determinant.\",\n      \"method\": \"Stable transfection of NTCP variants, taurocholate and rosuvastatin uptake assays in transfected cells\",\n      \"journal\": \"Xenobiotica\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — functional characterization of naturally occurring variants with multiple substrates; single lab\",\n      \"pmids\": [\"21341987\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 1999,\n      \"finding\": \"Two alternatively spliced isoforms of mouse Ntcp (Ntcp1 encoding 362 aa and Ntcp2 encoding 317 aa with a shorter C-terminus from intron retention) both mediate saturable Na+-dependent transport of taurocholate when expressed in Xenopus oocytes.\",\n      \"method\": \"cDNA library screening, Xenopus oocyte expression, taurocholate uptake assay\",\n      \"journal\": \"Biochimica et biophysica acta\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 / Strong — functional reconstitution in Xenopus oocytes with saturable kinetics for both isoforms\",\n      \"pmids\": [\"10209268\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2020,\n      \"finding\": \"Position S267 in NTCP functions as an evolutionary 'rheostat': all 20 amino acid substitutions at this position produce progressive, varied effects on transport kinetics (Km and Vmax) and substrate specificity for taurocholic acid, estrone-3-sulfate, and rosuvastatin, rather than simple on/off effects. Stability modeling showed substitution tolerance correlates with surface expression but not transport activity.\",\n      \"method\": \"Systematic mutagenesis of all 20 amino acids at position 267, transport assays with three substrates, surface expression analysis, Rosetta stability modeling\",\n      \"journal\": \"The Journal of biological chemistry\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1-2 / Strong — comprehensive systematic mutagenesis with multiple substrates and stability modeling; mechanistically informative\",\n      \"pmids\": [\"33168628\"],\n      \"is_preprint\": false\n    }\n  ],\n  \"current_model\": \"SLC10A1/NTCP is an electrogenic, sodium-dependent bile acid cotransporter localized to the basolateral membrane of hepatocytes, where it functions as the primary importer of conjugated bile acids from portal blood into hepatocytes; its cryo-EM structure reveals a transmembrane tunnel through which bile acids and the HBV preS1 domain compete for access, NTCP undergoes regulated trafficking to and from the plasma membrane controlled by PI3K/PKB/Akt signaling, PP2B-mediated dephosphorylation of Ser-226, PKCζ-dependent microtubule-based vesicle transport, clathrin-dependent endocytosis triggered by a dileucine motif, and ubiquitin-proteasome-mediated ERAD; its expression is transcriptionally regulated by RXR:RAR, HNF1α, HNF4α, RAR, and cyclin D1, and post-translationally regulated by N-linked glycosylation (required for plasma membrane targeting) and S-nitrosylation (inhibitory); NTCP also serves as the high-affinity receptor for HBV and HDV entry via the myristoylated preS1 domain, which blocks the extracellular entrance to the bile acid transport tunnel, with viral internalization requiring NTCP oligomerization downstream of EGFR co-receptor interaction and proceeding through clathrin-mediated endocytosis.\"\n}\n```","stage2_raw":"{\n  \"mechanistic_narrative\": \"SLC10A1 (NTCP) is the principal sodium-dependent transporter that imports conjugated bile acids into hepatocytes, functioning at the rate-limiting basolateral uptake step of the enterohepatic bile salt cycle [#18, #30]. It transports taurine and glycine conjugates in preference to unconjugated bile salts, and when reconstituted with the apical exporter BSEP it reconstitutes vectorial basolateral-to-apical bile acid flux across polarized epithelium [#18]. Cryo-EM structures show a transmembrane tunnel forming the substrate translocation route; residue 267 lies within a region critical for bile acid recognition, where substitutions act as a graded rheostat over transport kinetics and substrate selectivity while sparing the non-bile-acid substrate estrone sulfate [#0, #20, #37]. NTCP activity at the plasma membrane is set by regulated trafficking: PI3K/PKB(Akt) signaling and PP2B(calcineurin)-mediated dephosphorylation of Ser-226 promote membrane insertion, PKCζ drives microtubule-based motility of NTCP recycling vesicles, a dileucine motif and PKC-triggered clathrin-dependent endocytosis remove it, and misfolded NTCP is cleared by ubiquitin-proteasome ERAD [#6, #7, #8, #9, #21, #10]. N-linked glycosylation is required for plasma membrane targeting, and S-nitrosylation of cysteines non-competitively inhibits transport [#19, #22]. Transcription of the gene is controlled by HNF1α, HNF4α, RXR:RAR and RAR, and is repressed in cholestasis through bile-acid/FXR-dependent loss of these activators and by inflammatory JNK signaling [#12, #25, #14, #11]. Independently of its metabolic role, NTCP is the high-affinity hepatocyte entry receptor for HBV and HDV: the myristoylated preS1 domain of the HBV large surface protein binds the extracellular mouth of the transport tunnel in direct competition with bile acid substrate, and viral internalization requires EGFR-dependent NTCP oligomerization followed by clathrin-mediated endocytosis [#0, #3, #5, #27, #32]. Homozygous loss-of-function mutation (R252H) causes NTCP deficiency, an inborn error of bile salt metabolism with markedly reduced bile acid uptake [#30].\",\n  \"teleology\": [\n    {\n      \"year\": 1999,\n      \"claim\": \"Establishing that NTCP intrinsically carries out sodium-dependent bile acid transport defined its core molecular activity independent of cellular context.\",\n      \"evidence\": \"Functional expression of two mouse Ntcp splice isoforms in Xenopus oocytes with saturable taurocholate uptake\",\n      \"pmids\": [\"10209268\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Oocyte system does not address polarized hepatocyte localization\", \"Did not define substrate-recognition residues\"]\n    },\n    {\n      \"year\": 2002,\n      \"claim\": \"Identifying PP2B-mediated dephosphorylation and PI3K/PKB signaling as drivers of NTCP membrane translocation showed that transporter activity is acutely controlled by regulated trafficking rather than expression alone.\",\n      \"evidence\": \"PP2B inhibition and immunoprecipitated-NTCP dephosphorylation in rat hepatocytes; dominant-negative/constitutively active PKB with translocation and uptake readouts\",\n      \"pmids\": [\"12065290\", \"12034724\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Did not identify the target phosphosite\", \"Upstream stimulus integration unresolved at this stage\"]\n    },\n    {\n      \"year\": 2002,\n      \"claim\": \"Showing that inflammatory signaling represses Ntcp transcription via JNK-phosphorylated RXR linked NTCP downregulation to the hepatic response to cytokines and cholestasis.\",\n      \"evidence\": \"Ntcp promoter luciferase, dominant-negative JNK, EMSA in primary rat hepatocytes; SHP-1 induction timing in bile-duct-ligated mice\",\n      \"pmids\": [\"12105223\", \"11751172\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"SHP-1 link was temporal correlation only\", \"Did not separate cytokine vs bile-acid contributions\"]\n    },\n    {\n      \"year\": 2003,\n      \"claim\": \"Mapping species-specific and conserved transcription factor inputs (HNF1α, HNF4α, RXR/RAR, HNF3β, C/EBPβ) defined the regulatory architecture of the NTCP promoter.\",\n      \"evidence\": \"Cross-species luciferase reporter and EMSA assays in Huh7 cells\",\n      \"pmids\": [\"14701722\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Species divergence complicates extrapolation to human regulation\", \"In vivo occupancy not established here\"]\n    },\n    {\n      \"year\": 2003,\n      \"claim\": \"Demonstrating that the S267F polymorphism abolishes bile acid uptake while sparing estrone sulfate and membrane expression localized substrate recognition to a discrete region.\",\n      \"evidence\": \"Variant transport assays with multiple substrates plus cell-surface biotinylation in HepG2/transfected cells\",\n      \"pmids\": [\"14660639\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Atomic basis of substrate selectivity not yet resolved\", \"Physiological consequence in carriers not addressed\"]\n    },\n    {\n      \"year\": 2005,\n      \"claim\": \"Pinpointing Ser-226 as the cAMP-regulated phosphosite governing membrane retention connected the kinase/phosphatase circuitry to a defined molecular switch.\",\n      \"evidence\": \"32P labeling, clostripain mapping, S226A mutagenesis with uptake and surface expression in HuH-7 cells\",\n      \"pmids\": [\"16027164\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Kinase phosphorylating Ser-226 not directly identified\", \"Interplay with the dileucine endocytosis motif not yet defined\"]\n    },\n    {\n      \"year\": 2005,\n      \"claim\": \"Establishing FXR-dependent, bile-acid-mediated (not cytokine-mediated) repression of Ntcp in obstructive cholestasis resolved the dominant in vivo control mechanism.\",\n      \"evidence\": \"FXR knockout mice, bile duct ligation, cholic acid feeding, LPS, EMSA; plus Kupffer-cell depletion and cytokine neutralization controls\",\n      \"pmids\": [\"16002565\", \"15629514\", \"15723437\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"FXR-to-HNF axis intermediate steps incomplete\", \"LPS/inflammatory repression operates by a distinct, less-defined pathway\"]\n    },\n    {\n      \"year\": 2005,\n      \"claim\": \"Identifying ERAD/ubiquitin-proteasome clearance of misfolded NTCP defined the protein's quality-control degradation route.\",\n      \"evidence\": \"Proteasome inhibitors, polyubiquitination assay, glycosylation analysis and MTOC aggresome colocalization in HepG2 cells\",\n      \"pmids\": [\"16218878\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Specific E3 ligase not identified\", \"Relationship to surface-pool turnover unresolved\"]\n    },\n    {\n      \"year\": 2006,\n      \"claim\": \"Reconstituting vectorial bile acid transport with NTCP and BSEP, and defining PKCζ-dependent vesicular motility, established NTCP's role in transcellular flux and its trafficking machinery.\",\n      \"evidence\": \"Double-transfected polarized LLC-PK1 transcellular flux; live imaging of NTCP-GFP vesicles with PKCζ inhibition and motor/endosome markers\",\n      \"pmids\": [\"16474011\", \"16734659\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Coupling of motility to specific signaling stimuli incomplete\", \"Kinesin/dynein switching control not defined\"]\n    },\n    {\n      \"year\": 2010,\n      \"claim\": \"Showing S-nitrosylation of NTCP cysteines causes reversible non-competitive transport inhibition added a redox-based post-translational control layer.\",\n      \"evidence\": \"Biotin switch assay, kinetic uptake, surface biotinylation, DTT rescue in HuH-NTCP cells\",\n      \"pmids\": [\"21109590\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Modified cysteine residues not individually mapped\", \"Physiological NO source not defined\"]\n    },\n    {\n      \"year\": 2013,\n      \"claim\": \"Defining a dileucine motif with dual endocytosis/membrane-targeting roles and clathrin-dependent PKC-triggered internalization completed the retrieval arm of NTCP trafficking.\",\n      \"evidence\": \"Mutagenesis of dileucine motif and Thr225/Ser226, flow cytometry, immunofluorescence in HepG2 cells\",\n      \"pmids\": [\"24008362\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Adaptor recognizing the motif not identified\", \"Reconciliation with Ser-226 dephosphorylation switch incomplete\"]\n    },\n    {\n      \"year\": 2013,\n      \"claim\": \"Confirming NTCP as the functional HBV entry receptor and mapping cyclosporin A action to the preS1-binding site established the receptor function and its overlap with transport.\",\n      \"evidence\": \"NTCP overexpression/knockdown HBV infection assays; cyclosporin A inhibition, preS1 binding, and transport-competent/binding-deficient variants\",\n      \"pmids\": [\"24342612\", \"24295872\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Post-attachment internalization steps not yet defined\", \"Co-receptor requirements unresolved at this stage\"]\n    },\n    {\n      \"year\": 2014,\n      \"claim\": \"Demonstrating reciprocal competition between preS1 and bile acids, and that R252H loss-of-function causes human NTCP deficiency, tied the viral and metabolic functions to a shared site and proved NTCP's role as the main hepatic bile salt importer.\",\n      \"evidence\": \"Reciprocal inhibition kinetics in primary hepatocytes and cell lines; patient sequencing with uptake assay and surface localization\",\n      \"pmids\": [\"24845614\", \"24867799\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Spectrum of clinical phenotypes in deficiency not detailed\", \"Quantitative overlap of viral vs substrate sites not structurally resolved here\"]\n    },\n    {\n      \"year\": 2014,\n      \"claim\": \"Identifying RAR control of the human NTCP promoter linked nuclear receptor signaling to both transporter expression and HBV permissiveness.\",\n      \"evidence\": \"Promoter luciferase, RAR ChIP, RAR antagonists with HBV infection in HepG2-hNTCP\",\n      \"pmids\": [\"25550158\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Single lab; in vivo relevance not established\", \"Endogenous RAR ligand context unclear\"]\n    },\n    {\n      \"year\": 2017,\n      \"claim\": \"Establishing that N-linked glycosylation at N5/N11 is required for membrane trafficking and HBV infection clarified a biosynthetic prerequisite for both functions.\",\n      \"evidence\": \"Glycosylation-site mutagenesis, surface biotinylation, lysosomal degradation and HBV/uptake assays in HepG2 cells\",\n      \"pmids\": [\"28125599\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Glycan-dependent chaperone interactions not identified\", \"Effect on oligomerization not tested\"]\n    },\n    {\n      \"year\": 2020,\n      \"claim\": \"Systematic saturation mutagenesis at position 267 reframed substrate recognition as a tunable rheostat coupling stability/expression and substrate-specific kinetics.\",\n      \"evidence\": \"All-20-amino-acid substitution with three substrates, surface expression, and Rosetta stability modeling; prior A64T/S267F variant kinetics\",\n      \"pmids\": [\"33168628\", \"21341987\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Structural rationalization of rheostat behavior incomplete\", \"Population-level pharmacogenetic impact not assessed\"]\n    },\n    {\n      \"year\": 2021,\n      \"claim\": \"Resolving that NTCP oligomerization downstream of EGFR is required for HBV internalization, and defining clathrin-dependent uptake, established the ordered post-attachment entry mechanism.\",\n      \"evidence\": \"SPR, F274A mutagenesis, CoIP, oligomerization assays with troglitazone; clathrin/AP-2/dynamin-2 knockdown and EM in HepG2-NTCP\",\n      \"pmids\": [\"34613794\", \"32216005\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Stoichiometry of the oligomer not defined\", \"How EGFR engagement triggers oligomerization unresolved\"]\n    },\n    {\n      \"year\": 2022,\n      \"claim\": \"Cryo-EM structures of NTCP, alone and with preS1, provided the atomic framework for the bile acid transport tunnel and the structural basis of viral receptor engagement.\",\n      \"evidence\": \"Cryo-EM of human/bovine/rat NTCP and antibody/preS1 complexes with mutation and transport assays\",\n      \"pmids\": [\"35580629\", \"35580630\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Substrate-bound and intermediate conformational states not captured\", \"Sodium coupling mechanism not fully detailed\"]\n    },\n    {\n      \"year\": 2022,\n      \"claim\": \"Identifying IFITM3 as an NTCP interaction partner facilitating a post-attachment HBV/HDV entry step expanded the receptor complex beyond NTCP itself.\",\n      \"evidence\": \"Membrane yeast-two-hybrid, CoIP, IFITM3 knockdown with HBV/HDV infection and preS1 binding in HuH7 and primary hepatocytes\",\n      \"pmids\": [\"35458456\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Single lab; mechanism of IFITM3 action at the entry step undefined\", \"Direct vs indirect interaction not fully resolved\"]\n    },\n    {\n      \"year\": 2024,\n      \"claim\": \"The bulevirtide-bound NTCP structure explained how the therapeutic peptide plugs the transport tunnel and underlies HBV/HDV host specificity.\",\n      \"evidence\": \"Cryo-EM of BLV-bound human NTCP; prior high-affinity, long-lived binding/turnover measurements\",\n      \"pmids\": [\"38509088\", \"32039379\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"In vivo pharmacodynamic linkage to NTCP turnover only correlative\", \"Resistance mechanisms not addressed\"]\n    },\n    {\n      \"year\": null,\n      \"claim\": \"How NTCP's bile-acid transport function mechanistically intersects with broader physiology—innate antiviral immunity, cell-cycle control, and systemic metabolism—remains incompletely defined.\",\n      \"evidence\": \"Open question integrating NTCP/ISG repression, cyclin D1 promoter suppression, and knockout metabolic phenotypes\",\n      \"pmids\": [],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Causal chain from bile-acid transport to ISG/IFITM3 repression not fully mapped\", \"Mechanism coupling NTCP expression to G0/G1 arrest unresolved\", \"Tissue-level metabolic effects of NTCP loss in humans not characterized\"]\n    }\n  ],\n  \"mechanism_profile\": {\n    \"molecular_activity\": [\n      {\"term_id\": \"GO:0005215\", \"supporting_discovery_ids\": [18, 30, 36, 20, 37]},\n      {\"term_id\": \"GO:0140104\", \"supporting_discovery_ids\": [18, 36]},\n      {\"term_id\": \"GO:0001618\", \"supporting_discovery_ids\": [3, 4, 5, 0]},\n      {\"term_id\": \"GO:0038024\", \"supporting_discovery_ids\": [17]}\n    ],\n    \"localization\": [\n      {\"term_id\": \"GO:0005886\", \"supporting_discovery_ids\": [6, 8, 18, 19, 30]},\n      {\"term_id\": \"GO:0005768\", \"supporting_discovery_ids\": [9, 21]},\n      {\"term_id\": \"GO:0005783\", \"supporting_discovery_ids\": [10, 19]},\n      {\"term_id\": \"GO:0005815\", \"supporting_discovery_ids\": [10]}\n    ],\n    \"pathway\": [\n      {\"term_id\": \"R-HSA-382551\", \"supporting_discovery_ids\": [18, 30, 36]},\n      {\"term_id\": \"R-HSA-1430728\", \"supporting_discovery_ids\": [18, 33]},\n      {\"term_id\": \"R-HSA-1643685\", \"supporting_discovery_ids\": [3, 30, 5]},\n      {\"term_id\": \"R-HSA-168256\", \"supporting_discovery_ids\": [24, 26, 28]},\n      {\"term_id\": \"R-HSA-74160\", \"supporting_discovery_ids\": [11, 12, 25, 14]}\n    ],\n    \"complexes\": [],\n    \"partners\": [\"BSEP\", \"IFITM3\", \"OATP1B3\", \"EGFR\"],\n    \"other_free_text\": []\n  }\n}","audit_flag":null,"evaluation":{"pairwise":"win","faith_supported":8,"faith_total":8,"faith_pct":100.0}}