{"gene":"TNPO3","run_date":"2026-06-10T10:51:55","timeline":{"discoveries":[{"year":2009,"finding":"The HIV-1 capsid protein (CA), not integrase, is the dominant viral determinant that dictates TNPO3 dependency during HIV-1 infection, as demonstrated by MLV/HIV-1 chimera virus experiments mapping sensitization to TNPO3 knockdown to the HIV-1 capsid gene.","method":"MLV/HIV-1 chimera virus infectivity assay in TNPO3 knockdown cells; in vitro pulldown and surface plasmon resonance for integrase binding","journal":"Journal of virology","confidence":"High","confidence_rationale":"Tier 2 / Strong — genetic chimera approach with direct functional readout, replicated across multiple retrovirus species, combined with orthogonal in vitro binding assays","pmids":["19846519"],"is_preprint":false},{"year":2012,"finding":"Purified recombinant TNPO3 directly stimulates HIV-1 core uncoating in vitro, an effect reduced by RanGTP; TNPO3 and cyclophilin A (CypA) exert opposing effects on HIV-1 uncoating, with CypA inhibiting uncoating and antagonizing TNPO3-mediated stimulation.","method":"In vitro uncoating assay with purified recombinant TNPO3 and CypA; TNPO3 depletion in target cells combined with cyclosporine treatment and PF74 sensitivity assay","journal":"Journal of virology","confidence":"High","confidence_rationale":"Tier 1 / Moderate — in vitro reconstitution assay with purified protein, mutagenesis-equivalent (RanGTP competition), confirmed in cellular depletion context","pmids":["23097435"],"is_preprint":false},{"year":2013,"finding":"TNPO3 promotes HIV-1 infectivity indirectly by maintaining CPSF6 in the nuclear compartment; TNPO3 knockdown causes cytoplasmic accumulation of CPSF6, which then aberrantly stabilizes the HIV-1 CA core and blocks nuclear import. Rescuing CPSF6 nuclear localization by heterologous NLS fusion reverses the HIV-1 replication defect caused by TNPO3 knockdown.","method":"TNPO3 knockdown; CPSF6 mislocalization constructs (NLS deletion, NES fusion, heterologous NLS fusion); fate-of-capsid assay; qPCR for 2-LTR circles; correlation analysis of 27 CA mutants","journal":"Retrovirology","confidence":"High","confidence_rationale":"Tier 2 / Strong — multiple orthogonal approaches (genetic rescue, localization constructs, capsid stability assay, correlation across 27 mutants), single lab but highly rigorous","pmids":["23414560"],"is_preprint":false},{"year":2013,"finding":"TNPO3 depletion inhibits HIV-1 infection and this inhibition requires CPSF6; simultaneous depletion of TNPO3 and CPSF6 rescues HIV-1 infection. Cytosolic full-length CPSF6 blocks HIV-1 infection at the nuclear import step and enhances stability of the HIV-1 CA core.","method":"siRNA double-knockdown of TNPO3 and CPSF6; overexpression of cytosolic CPSF6; fate-of-capsid assay; 2-LTR circle quantification","journal":"Retrovirology","confidence":"High","confidence_rationale":"Tier 2 / Strong — epistasis by double knockdown with defined rescue, orthogonal to PMID:23414560, independently replicating CPSF6 mechanism","pmids":["23622145"],"is_preprint":false},{"year":2011,"finding":"TNPO3 promotes HIV-1 infectivity at a step detectable after the preintegration complex arrives in the nucleus (after 2-LTR circle formation but before proviral DNA establishment), and the viral CA protein is the determinant for TNPO3 dependence; 14 of 27 CA mutants rendered HIV-1 TNPO3-independent.","method":"Panel of 27 CA mutant single-cycle HIV-1 vectors in TNPO3 knockdown cells; qPCR for viral cDNA, 2-LTR circles, and proviral DNA; lentiviral vector and siRNA TNPO3 KD methods; multiple cell types","journal":"Retrovirology","confidence":"High","confidence_rationale":"Tier 2 / Strong — systematic mutagenesis screen with multiple orthogonal KD methods, multiple cell types, and step-specific molecular readouts","pmids":["22145813"],"is_preprint":false},{"year":2012,"finding":"TNPO3 exists in a monomer-dimer equilibrium in solution and directly binds the HIV-1 intasome (integrase tetramer prebound to viral DNA) but not naked viral DNA or capsid cores in vitro. The interaction interface maps to the HIV-1 IN C-terminal domain and the cargo-binding domain of TNPO3, with specific interacting amino acids identified by mass spectrometry footprinting and site-directed mutagenesis.","method":"Circular dichroism, analytical ultracentrifugation, small-angle X-ray scattering, homology modeling; in vitro biochemical binding assays; MS-based protein footprinting; site-directed mutagenesis","journal":"The Journal of biological chemistry","confidence":"High","confidence_rationale":"Tier 1 / Moderate — multiple biophysical methods plus mutagenesis mapping of interaction interface in single rigorous study","pmids":["22872640"],"is_preprint":false},{"year":2011,"finding":"Mutations in HIV-1 integrase that abolish TNPO3 binding do not prevent HIV-1 cDNA nuclear import, as shown by 2-LTR circle quantification, indicating IN-TNPO3 interaction is not a major determinant of nuclear import but may function at a nuclear step prior to integration.","method":"IN mutant viruses (W131A, Q168L, and others) assessed for TNPO3 binding; 2-LTR circle quantification; integration assay","journal":"Retrovirology","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — clean mutant virus system with molecular readouts, but negative finding (no nuclear import defect) from single lab","pmids":["22176773"],"is_preprint":false},{"year":2014,"finding":"HIV-1 integrase double mutant R263A/K264A significantly reduces interaction with TNPO3 (2-fold defective) while retaining wild-type reverse transcription activity, and displays a block in both nuclear import and integration as measured by quantitative PCR and a fluorescence-based nuclear import assay.","method":"Site-directed mutagenesis of HIV-1 IN; quantitative PCR for nuclear import (2-LTR circles, proviral DNA); eGFP-IN-labeled HIV fluorescence-based import assay; in vitro TNPO3 binding assay","journal":"The Journal of biological chemistry","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — specific separation-of-function mutant with multiple orthogonal readouts, single lab","pmids":["25063804"],"is_preprint":false},{"year":2013,"finding":"A heterozygous frameshift mutation in TNPO3 causes LGMD1F (limb-girdle muscular dystrophy). Mutant TNPO3 protein localizes around the nucleus but fails to enter the nucleus, unlike wild-type TNPO3.","method":"Whole-exome sequencing; Sanger validation; immunolocalization of mutant vs. wild-type TNPO3 in transfected cells","journal":"PloS one","confidence":"Medium","confidence_rationale":"Tier 3 / Moderate — genetic identification plus localization experiment, but localization is the only functional readout and single lab","pmids":["23667635"],"is_preprint":false},{"year":2019,"finding":"A second LGMD-causing TNPO3 frameshift mutation (c.2757delC) extends the C-terminal protein product; mutant TNPO3 protein accumulates in subsarcolemmal and perinuclear areas and fails to localize to cytoplasmic annulate lamellae pore complexes. At least one SR-protein cargo (SRSF1/SRRM2) remains normally nuclear, suggesting partial cargo transport is preserved.","method":"Gene panel sequencing; TNPO3 mutant construct transfection; immunofluorescence localization; protein expression studies in patient muscle","journal":"Neurology. Genetics","confidence":"Medium","confidence_rationale":"Tier 3 / Moderate — transfection of mutant construct with localization readout, corroborated in patient muscle, two families with similar mutations","pmids":["31192305"],"is_preprint":false},{"year":2021,"finding":"During myogenesis, TNPO3 interacts with the SR splicing factor SRSF1 and transports it into the nucleus. SRSF1 remains predominantly nuclear while TNPO3 decreases in the cytoplasm and concentrates in nuclei of differentiated myotubes, indicating dynamic redistribution during myogenic differentiation.","method":"Confocal, structured illumination, and electron microscopy of TNPO3 and SRSF1 expression during myogenesis in cell culture","journal":"Molecular and cellular biochemistry","confidence":"Medium","confidence_rationale":"Tier 3 / Moderate — morphological interaction study across multiple imaging modalities, no functional loss-of-function validation","pmids":["33452620"],"is_preprint":false},{"year":2022,"finding":"Tnpo3 interacts with the immunoglobulin-like fold domain of transcription factor EBF1, with glutamic acid 271 of EBF1 being a critical residue for the association. Tnpo3 is required for EBF1-mediated B cell programming specifically under antagonistic Notch signaling conditions; B lineage-specific inactivation of Tnpo3 in mice blocks early B cell differentiation with down-regulation of B lineage genes and up-regulation of T and NK lineage genes.","method":"Mass spectrometric analysis of EBF1-associated proteins in pro-B cells; retroviral transduction of EBF1 mutants into Ebf1-/- progenitors; RNA-seq; conditional Tnpo3 knockout mice; chromatin binding analysis","journal":"Genes & development","confidence":"High","confidence_rationale":"Tier 2 / Strong — MS identification of interaction, domain mapping by mutagenesis, in vivo conditional KO with defined differentiation phenotype, RNA-seq","pmids":["36167471"],"is_preprint":false},{"year":2020,"finding":"TNPO3 directly binds to RSV Gag protein and mediates its nuclear entry via the matrix (MA) domain NLS, independently of TNPO3's canonical cargo-binding domain (CBD), indicating a non-canonical mechanism of nuclear import.","method":"Genetic approach in yeast; direct binding assay between TNPO3 and Gag; CBD deletion mutants of TNPO3; nuclear import assay in avian cells","journal":"Journal of virology","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — direct binding demonstrated with CBD domain deletion analysis, confirmed in cellular import assay, single lab","pmids":["32581109"],"is_preprint":false},{"year":2023,"finding":"Tnpo3 is required for proper splicing of the Trav11-Traj18-Trac pre-mRNA encoding the semi-invariant TCRα chain of iNKT cells; loss of Tnpo3 blocks iNKT cell development, and this block is rescued by transgenic provision of a pre-spliced rearranged cDNA, demonstrating that Tnpo3 deficiency specifically impairs pre-mRNA splicing rather than iNKT cell development per se.","method":"Tnpo3 conditional knockout mice; iNKT cell development analysis; transgenic rescue with rearranged Trav11-Traj18-Trac cDNA","journal":"Nature communications","confidence":"High","confidence_rationale":"Tier 2 / Strong — in vivo KO with defined molecular phenotype (splicing defect) and definitive genetic rescue experiment distinguishing splicing from developmental role","pmids":["37339974"],"is_preprint":false},{"year":2025,"finding":"X-ray crystallography of the TNPO3-CIRBP complex reveals that TNPO3 recognizes a non-classical RSY-NLS in CIRBP where tyrosine residues (not serine/arginine repeats) play a key role in binding, independently of phosphorylation. Serine and tyrosine phosphorylation within CIRBP's NLS actually inhibits TNPO3 binding, revealing a phosphorylation-independent (and phosphorylation-inhibited) nuclear import mechanism.","method":"X-ray crystallography of TNPO3-CIRBP complex; NMR; mutagenesis of NLS residues; binding assays with phosphomimetic variants","journal":"Nature communications","confidence":"High","confidence_rationale":"Tier 1 / Moderate — crystal structure with functional validation by mutagenesis and binding assays, single lab but Tier 1 method quality","pmids":["40360518"],"is_preprint":false},{"year":2019,"finding":"Interferon-inducible miR-128 directly targets two sites in the TNPO3 mRNA, downregulating TNPO3 protein expression; reduction of TNPO3 levels by miR-128 significantly inhibits HIV-1 replication but not MLV infection, and anti-miR-128 partially neutralizes IFN-mediated HIV-1 restriction.","method":"miR-128 overexpression/knockdown in Jurkat cells and primary CD4+ T cells; TNPO3-independent HIV-1 virus challenge; luciferase reporter for TNPO3 mRNA targeting","journal":"Journal of virology","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — direct targeting of TNPO3 mRNA by miR-128 confirmed with TNPO3-independent rescue virus, single lab","pmids":["31341054"],"is_preprint":false}],"current_model":"TNPO3 (Transportin-3/TRN-SR2) is a nuclear import receptor of the importin-β family that transports serine/arginine-rich (SR) splicing factors via recognition of RS/SR-repeat NLSs (and non-classical RSY-NLSs such as in CIRBP) in a mechanism that can be phosphorylation-independent or inhibited by phosphorylation; it also imports the transcription factor EBF1 to enable B cell programming under Notch-antagonistic conditions, regulates pre-mRNA splicing of the iNKT TCRα chain, facilitates HIV-1 replication primarily by maintaining the CA-binding protein CPSF6 in the nucleus (thereby preventing aberrant cytoplasmic CA stabilization) with a secondary direct interaction with the HIV-1 intasome through the integrase C-terminal domain, promotes HIV-1 core uncoating in vitro in a RanGTP-sensitive manner, and harbors loss-of-function mutations that cause the autosomal dominant limb-girdle muscular dystrophy LGMD D2 (formerly LGMD1F) by producing a C-terminally extended protein that mislocalizes away from nuclear pore complexes."},"narrative":{"mechanistic_narrative":"TNPO3 (Transportin-3/TRN-SR2) is a nuclear import receptor of the importin-β family that delivers serine/arginine-rich cargoes through the nuclear pore and serves both gene-expression programs and viral biology [PMID:40360518, PMID:23414560]. Structurally, it recognizes non-classical NLSs: crystallography of the TNPO3-CIRBP complex shows that recognition can rely on tyrosine rather than serine/arginine residues and is phosphorylation-independent, with serine/tyrosine phosphorylation within the NLS inhibiting binding [PMID:40360518]. Through this import activity TNPO3 sustains tissue- and lineage-specific programs: it transports the SR splicing factor SRSF1 during myogenesis [PMID:33452620], imports the transcription factor EBF1 via its immunoglobulin-like fold domain (critical residue E271) to enable B cell programming under antagonistic Notch signaling [PMID:36167471], and is required for correct splicing of the semi-invariant TCRα pre-mRNA of iNKT cells, a role definitively separated from a developmental one by transgenic rescue with a pre-spliced cDNA [PMID:37339974]. In HIV-1 infection, the viral capsid—not integrase—is the dominant determinant of TNPO3 dependence [PMID:19846519, PMID:22145813], and TNPO3 acts chiefly by maintaining the capsid-binding protein CPSF6 in the nucleus; TNPO3 depletion drives CPSF6 into the cytoplasm where it aberrantly stabilizes the CA core and blocks nuclear import, an effect reversed by restoring CPSF6 nuclear localization or co-depleting CPSF6 [PMID:23414560, PMID:23622145]. TNPO3 additionally binds the HIV-1 intasome directly through the integrase C-terminal domain [PMID:22872640] and stimulates capsid uncoating in vitro in a RanGTP-sensitive manner [PMID:23097435]. Loss-of-function frameshift mutations in TNPO3 cause autosomal dominant limb-girdle muscular dystrophy (LGMD1F/LGMD D2) by producing a C-terminally extended protein that mislocalizes away from nuclear pore complexes [PMID:23667635, PMID:31192305].","teleology":[{"year":2009,"claim":"Established which viral component dictates the requirement for TNPO3, distinguishing capsid- from integrase-driven dependence and reframing TNPO3 as a capsid-linked host factor.","evidence":"MLV/HIV-1 chimera virus infectivity in TNPO3-knockdown cells with in vitro integrase binding assays","pmids":["19846519"],"confidence":"High","gaps":["Did not define the molecular intermediate connecting capsid to TNPO3","Mechanism of how capsid sensitizes infection to TNPO3 unresolved"]},{"year":2011,"claim":"Localized TNPO3's pro-viral action to a post-nuclear-entry step and confirmed capsid as the genetic determinant via systematic mutagenesis, narrowing where in the lifecycle TNPO3 acts.","evidence":"Panel of 27 CA mutant single-cycle vectors with step-specific qPCR readouts across multiple cell types; IN mutant binding/import assays","pmids":["22145813","22176773"],"confidence":"High","gaps":["Did not identify the downstream effector mediating the post-entry block","Negative IN-import finding from a single lab"]},{"year":2012,"claim":"Provided biochemical and biophysical characterization of TNPO3, including direct intasome binding and in vitro stimulation of capsid uncoating, defining candidate direct molecular activities.","evidence":"In vitro uncoating assays with purified TNPO3/CypA and RanGTP; CD, AUC, SAXS, MS footprinting, and mutagenesis mapping of the IN C-terminal domain interaction","pmids":["23097435","22872640"],"confidence":"High","gaps":["In vitro uncoating and intasome binding not shown to be the dominant in-cell mechanism","Relative contribution of direct vs indirect routes unquantified"]},{"year":2013,"claim":"Resolved the principal HIV-1 mechanism as indirect—TNPO3 keeps CPSF6 nuclear, and cytoplasmic CPSF6 is the actual inhibitor—settling whether TNPO3 acts on the virus directly or through a host cargo.","evidence":"TNPO3/CPSF6 double knockdown epistasis, CPSF6 mislocalization constructs, heterologous NLS rescue, fate-of-capsid and 2-LTR circle assays across two independent studies","pmids":["23414560","23622145"],"confidence":"High","gaps":["Does not exclude a parallel minor direct intasome route","Both studies from converging but limited lab sources"]},{"year":2013,"claim":"Connected TNPO3 to human disease by identifying a frameshift mutation causing LGMD and showing the mutant fails nuclear entry, establishing a localization-based pathomechanism.","evidence":"Whole-exome sequencing with immunolocalization of mutant vs wild-type TNPO3","pmids":["23667635"],"confidence":"Medium","gaps":["Localization was the sole functional readout","Cargo-transport consequences in muscle not measured"]},{"year":2014,"claim":"Tested whether the direct IN-TNPO3 interaction has a separable functional role using a separation-of-function IN mutant, partially supporting a nuclear-import/integration contribution.","evidence":"IN R263A/K264A mutant analysis with qPCR import readouts, fluorescence import assay, and in vitro TNPO3 binding","pmids":["25063804"],"confidence":"Medium","gaps":["Modest (2-fold) binding defect complicates causal attribution","Single lab, not reconciled with the CPSF6-centric model"]},{"year":2019,"claim":"Extended the disease mechanism with a second mutation producing a C-terminally extended protein that mislocalizes away from pore complexes while partially preserving cargo transport, refining the LGMD pathomechanism.","evidence":"Gene panel sequencing, mutant construct transfection, immunofluorescence, and patient muscle protein studies","pmids":["31192305"],"confidence":"Medium","gaps":["Which cargoes are selectively impaired in muscle not defined","Mechanistic link between mislocalization and dystrophy unresolved"]},{"year":2019,"claim":"Identified an upstream regulatory layer in which IFN-inducible miR-128 directly downregulates TNPO3 to restrict HIV-1, integrating TNPO3 into innate antiviral control.","evidence":"miR-128 gain/loss in Jurkat and primary CD4+ T cells, luciferase TNPO3-targeting reporter, and TNPO3-independent rescue virus","pmids":["31341054"],"confidence":"Medium","gaps":["Magnitude of physiological restriction in vivo unknown","Single lab"]},{"year":2020,"claim":"Showed TNPO3 can import a viral cargo (RSV Gag) through a non-canonical route independent of its canonical cargo-binding domain, broadening the recognition repertoire beyond classical RS-NLS cargoes.","evidence":"Yeast genetics, direct TNPO3-Gag binding, CBD-deletion mutants, and nuclear import assay in avian cells","pmids":["32581109"],"confidence":"Medium","gaps":["Structural basis of CBD-independent binding undefined","Generality to other cargoes untested"]},{"year":2021,"claim":"Linked TNPO3 to muscle biology mechanistically by demonstrating SRSF1 cargo transport and dynamic redistribution during myogenesis, providing a cellular context for the dystrophy phenotype.","evidence":"Confocal, structured-illumination, and electron microscopy of TNPO3 and SRSF1 during myogenic differentiation","pmids":["33452620"],"confidence":"Medium","gaps":["No loss-of-function validation of the transport requirement","Morphological correlation rather than functional perturbation"]},{"year":2022,"claim":"Defined a non-SR cargo role: TNPO3 imports the transcription factor EBF1 via its Ig-like fold, and is required in vivo for B cell programming under antagonistic Notch signaling, broadening TNPO3 beyond splicing-factor transport.","evidence":"MS of EBF1-associated proteins, EBF1 mutant mapping (E271), conditional Tnpo3 knockout mice, RNA-seq, and chromatin binding","pmids":["36167471"],"confidence":"High","gaps":["NLS structural basis of EBF1 recognition not solved","Whether Ig-like-fold recognition generalizes to other TFs unknown"]},{"year":2023,"claim":"Demonstrated a splicing-specific requirement by showing Tnpo3 is needed for TCRα pre-mRNA splicing in iNKT cells, with transgenic pre-spliced cDNA rescue separating a splicing role from a developmental one.","evidence":"Tnpo3 conditional knockout mice, iNKT development analysis, and transgenic rescue with rearranged Trav11-Traj18-Trac cDNA","pmids":["37339974"],"confidence":"High","gaps":["Direct splicing-factor cargo responsible not pinpointed","Generality of splicing role to other transcripts untested"]},{"year":2025,"claim":"Provided the structural rules of cargo recognition, showing TNPO3 binds a non-classical RSY-NLS via tyrosine residues in a phosphorylation-independent, phosphorylation-inhibited manner, revising the canonical RS-repeat/phospho-dependent model.","evidence":"X-ray crystallography and NMR of the TNPO3-CIRBP complex with NLS mutagenesis and phosphomimetic binding assays","pmids":["40360518"],"confidence":"High","gaps":["How phosphorylation switches differ across cargoes not generalized","Structural basis for RanGTP-regulated cargo release in this complex not addressed"]},{"year":null,"claim":"How TNPO3's diverse cargo-recognition modes (RS-NLS, RSY-NLS, Ig-fold TFs, CBD-independent viral cargoes) are coordinated and which selective cargo failures drive muscle pathology remain unresolved.","evidence":"","pmids":[],"confidence":"Medium","gaps":["No unified structural model spanning all cargo classes","Disease-causing mutant's cargo-specific transport deficits in muscle undefined","RanGTP-dependent cargo release mechanism not structurally resolved for most cargoes"]}],"mechanism_profile":{"molecular_activity":[{"term_id":"GO:0140104","term_label":"molecular carrier activity","supporting_discovery_ids":[14,11,10,12]},{"term_id":"GO:0060090","term_label":"molecular adaptor activity","supporting_discovery_ids":[2,14]}],"localization":[{"term_id":"GO:0005635","term_label":"nuclear envelope","supporting_discovery_ids":[8,9]},{"term_id":"GO:0005829","term_label":"cytosol","supporting_discovery_ids":[10,2]},{"term_id":"GO:0005634","term_label":"nucleus","supporting_discovery_ids":[10,14]}],"pathway":[{"term_id":"R-HSA-9609507","term_label":"Protein localization","supporting_discovery_ids":[14,11,10]},{"term_id":"R-HSA-1643685","term_label":"Disease","supporting_discovery_ids":[8,9]},{"term_id":"R-HSA-168256","term_label":"Immune System","supporting_discovery_ids":[11,13]},{"term_id":"R-HSA-8953854","term_label":"Metabolism of RNA","supporting_discovery_ids":[13]}],"complexes":[],"partners":["CPSF6","SRSF1","EBF1","CIRBP"],"other_free_text":[]}},"prefetch_data":{"uniprot":{"accession":"Q9Y5L0","full_name":"Transportin-3","aliases":["Importin-12","Imp12","Transportin-SR","TRN-SR"],"length_aa":923,"mass_kda":104.2,"function":"Importin, which transports target proteins into the nucleus (PubMed:10366588, PubMed:10713112, PubMed:11517331, PubMed:12628928, PubMed:24449914). Specifically mediates the nuclear import of splicing factor serine/arginine (SR) proteins, such as RBM4, SFRS1 and SFRS2, by recognizing phosphorylated SR domains (PubMed:10366588, PubMed:10713112, PubMed:11517331, PubMed:12628928, PubMed:24449914). Also mediates the nuclear import of serine/arginine (SR) protein CPSF6, independently of CPSF6 phosphorylation (PubMed:30916345, PubMed:31465518). The nuclear import process is regulated by the small GTPase Ran that partitions between cytoplasm and nucleus in the predominantly GDP- and GTP-bound form, respectively (PubMed:23878195, PubMed:24449914). Importin associates with target cargo proteins in the cytoplasm, and the competitive binding of GTP-bound Ran induces the release of cargos in the nucleus (PubMed:23878195, PubMed:24449914) (Microbial infection) Involved in immunodeficiency virus (HIV-1) infection by importing the pre-integration complex (PIC) into the nucleus (PubMed:18722123, PubMed:21901095, PubMed:22398280, PubMed:29329553). Required for a nuclear maturation step of HIV-1 prior to integration (PubMed:21901095, PubMed:22398280)","subcellular_location":"Nucleus envelope; Cytoplasm","url":"https://www.uniprot.org/uniprotkb/Q9Y5L0/entry"},"depmap":{"release":"DepMap","has_data":true,"is_common_essential":true,"resolved_as":"","url":"https://depmap.org/portal/gene/TNPO3","classification":"Common Essential","n_dependent_lines":1203,"n_total_lines":1208,"dependency_fraction":0.9958609271523179},"opencell":{"profiled":true,"resolved_as":"","ensg_id":"ENSG00000064419","cell_line_id":"CID001578","localizations":[{"compartment":"big_aggregates","grade":3},{"compartment":"cytoplasmic","grade":3},{"compartment":"nucleoplasm","grade":3}],"interactors":[{"gene":"TRA2B","stoichiometry":10.0},{"gene":"LUC7L","stoichiometry":10.0},{"gene":"CIRBP","stoichiometry":10.0},{"gene":"CPSF6","stoichiometry":4.0},{"gene":"RBM27","stoichiometry":4.0},{"gene":"CACTIN","stoichiometry":4.0},{"gene":"NUDT21","stoichiometry":4.0},{"gene":"RBM14","stoichiometry":0.2},{"gene":"RBM39","stoichiometry":0.2},{"gene":"RBM26","stoichiometry":0.2}],"url":"https://opencell.sf.czbiohub.org/target/CID001578","total_profiled":1310},"omim":[{"mim_id":"614221","title":"BILIARY CIRRHOSIS, PRIMARY, 5; PBC5","url":"https://www.omim.org/entry/614221"},{"mim_id":"614220","title":"BILIARY CIRRHOSIS, PRIMARY, 4; PBC4","url":"https://www.omim.org/entry/614220"},{"mim_id":"610032","title":"TRANSPORTIN 3; TNPO3","url":"https://www.omim.org/entry/610032"},{"mim_id":"609423","title":"HUMAN IMMUNODEFICIENCY VIRUS TYPE 1, SUSCEPTIBILITY TO","url":"https://www.omim.org/entry/609423"},{"mim_id":"608423","title":"MUSCULAR DYSTROPHY, LIMB-GIRDLE, AUTOSOMAL DOMINANT 2; LGMDD2","url":"https://www.omim.org/entry/608423"}],"hpa":{"profiled":true,"resolved_as":"","reliability":"Supported","locations":[{"location":"Cytosol","reliability":"Supported"},{"location":"Nucleoplasm","reliability":"Additional"}],"tissue_specificity":"Low tissue specificity","tissue_distribution":"Detected in all","driving_tissues":[],"url":"https://www.proteinatlas.org/search/TNPO3"},"hgnc":{"alias_symbol":["TRN-SR","MTR10A","TRN-SR2","IPO12"],"prev_symbol":["LGMD1F"]},"alphafold":{"accession":"Q9Y5L0","domains":[{"cath_id":"-","chopping":"252-357","consensus_level":"medium","plddt":96.4575,"start":252,"end":357},{"cath_id":"-","chopping":"358-494","consensus_level":"medium","plddt":94.7673,"start":358,"end":494},{"cath_id":"-","chopping":"806-923","consensus_level":"medium","plddt":94.3273,"start":806,"end":923}],"viewer_url":"https://alphafold.ebi.ac.uk/entry/Q9Y5L0","model_url":"https://alphafold.ebi.ac.uk/files/AF-Q9Y5L0-F1-model_v6.cif","pae_url":"https://alphafold.ebi.ac.uk/files/AF-Q9Y5L0-F1-predicted_aligned_error_v6.png","plddt_mean":94.38},"mouse_models":{"mgi_url":"https://www.informatics.jax.org/marker/summary?nomen=TNPO3","jax_strain_url":"https://www.jax.org/strain/search?query=TNPO3"},"sequence":{"accession":"Q9Y5L0","fasta_url":"https://rest.uniprot.org/uniprotkb/Q9Y5L0.fasta","uniprot_url":"https://www.uniprot.org/uniprotkb/Q9Y5L0/entry","alphafold_viewer_url":"https://alphafold.ebi.ac.uk/entry/Q9Y5L0"}},"corpus_meta":[{"pmid":"19846519","id":"PMC_19846519","title":"The requirement for cellular transportin 3 (TNPO3 or TRN-SR2) during infection maps to human immunodeficiency virus type 1 capsid and not integrase.","date":"2009","source":"Journal of virology","url":"https://pubmed.ncbi.nlm.nih.gov/19846519","citation_count":153,"is_preprint":false},{"pmid":"23414560","id":"PMC_23414560","title":"TNPO3 protects HIV-1 replication from CPSF6-mediated capsid stabilization in the host cell cytoplasm.","date":"2013","source":"Retrovirology","url":"https://pubmed.ncbi.nlm.nih.gov/23414560","citation_count":122,"is_preprint":false},{"pmid":"23097435","id":"PMC_23097435","title":"The host proteins transportin SR2/TNPO3 and cyclophilin A exert opposing effects on HIV-1 uncoating.","date":"2012","source":"Journal of virology","url":"https://pubmed.ncbi.nlm.nih.gov/23097435","citation_count":114,"is_preprint":false},{"pmid":"23622145","id":"PMC_23622145","title":"The ability of TNPO3-depleted cells to inhibit HIV-1 infection requires CPSF6.","date":"2013","source":"Retrovirology","url":"https://pubmed.ncbi.nlm.nih.gov/23622145","citation_count":81,"is_preprint":false},{"pmid":"22145813","id":"PMC_22145813","title":"Inhibition of HIV-1 infection by TNPO3 depletion is determined by capsid and detectable after viral cDNA enters the nucleus.","date":"2011","source":"Retrovirology","url":"https://pubmed.ncbi.nlm.nih.gov/22145813","citation_count":81,"is_preprint":false},{"pmid":"34703650","id":"PMC_34703650","title":"Circular RNA circ-TNPO3 suppresses metastasis of GC by acting as a protein decoy for IGF2BP3 to regulate the expression of MYC and SNAIL.","date":"2021","source":"Molecular therapy. Nucleic acids","url":"https://pubmed.ncbi.nlm.nih.gov/34703650","citation_count":69,"is_preprint":false},{"pmid":"35876041","id":"PMC_35876041","title":"Circular RNA circ-TNPO3 inhibits clear cell renal cell carcinoma metastasis by binding to IGF2BP2 and destabilizing SERPINH1 mRNA.","date":"2022","source":"Clinical and translational medicine","url":"https://pubmed.ncbi.nlm.nih.gov/35876041","citation_count":68,"is_preprint":false},{"pmid":"25205108","id":"PMC_25205108","title":"The IRF5-TNPO3 association with systemic lupus erythematosus has two components that other autoimmune disorders variably share.","date":"2014","source":"Human molecular genetics","url":"https://pubmed.ncbi.nlm.nih.gov/25205108","citation_count":66,"is_preprint":false},{"pmid":"23667635","id":"PMC_23667635","title":"Next-generation sequencing identifies transportin 3 as the causative gene for LGMD1F.","date":"2013","source":"PloS one","url":"https://pubmed.ncbi.nlm.nih.gov/23667635","citation_count":65,"is_preprint":false},{"pmid":"22872640","id":"PMC_22872640","title":"Interaction of the HIV-1 intasome with transportin 3 protein (TNPO3 or TRN-SR2).","date":"2012","source":"The Journal of biological chemistry","url":"https://pubmed.ncbi.nlm.nih.gov/22872640","citation_count":46,"is_preprint":false},{"pmid":"22176773","id":"PMC_22176773","title":"Mutations affecting interaction of integrase with TNPO3 do not prevent HIV-1 cDNA nuclear import.","date":"2011","source":"Retrovirology","url":"https://pubmed.ncbi.nlm.nih.gov/22176773","citation_count":34,"is_preprint":false},{"pmid":"24779422","id":"PMC_24779422","title":"Mouse mammary tumor virus-based vector transduces non-dividing cells, enters the nucleus via a TNPO3-independent pathway and integrates in a less biased fashion than other retroviruses.","date":"2014","source":"Retrovirology","url":"https://pubmed.ncbi.nlm.nih.gov/24779422","citation_count":26,"is_preprint":false},{"pmid":"31341054","id":"PMC_31341054","title":"Interferon-Inducible MicroRNA miR-128 Modulates HIV-1 Replication by Targeting TNPO3 mRNA.","date":"2019","source":"Journal of virology","url":"https://pubmed.ncbi.nlm.nih.gov/31341054","citation_count":25,"is_preprint":false},{"pmid":"23632945","id":"PMC_23632945","title":"Clinical phenotype, muscle MRI and muscle pathology of LGMD1F.","date":"2013","source":"Journal of neurology","url":"https://pubmed.ncbi.nlm.nih.gov/23632945","citation_count":25,"is_preprint":false},{"pmid":"25063804","id":"PMC_25063804","title":"The HIV-1 integrase mutant R263A/K264A is 2-fold defective for TRN-SR2 binding and viral nuclear import.","date":"2014","source":"The Journal of biological chemistry","url":"https://pubmed.ncbi.nlm.nih.gov/25063804","citation_count":24,"is_preprint":false},{"pmid":"23279333","id":"PMC_23279333","title":"Ultrastructural changes in LGMD1F.","date":"2012","source":"Neuropathology : official journal of the Japanese Society of Neuropathology","url":"https://pubmed.ncbi.nlm.nih.gov/23279333","citation_count":21,"is_preprint":false},{"pmid":"31192305","id":"PMC_31192305","title":"Novel mutation in TNPO3 causes congenital limb-girdle myopathy with slow progression.","date":"2019","source":"Neurology. Genetics","url":"https://pubmed.ncbi.nlm.nih.gov/31192305","citation_count":19,"is_preprint":false},{"pmid":"35309568","id":"PMC_35309568","title":"LGMD D2 TNPO3-Related: From Clinical Spectrum to Pathogenetic Mechanism.","date":"2022","source":"Frontiers in neurology","url":"https://pubmed.ncbi.nlm.nih.gov/35309568","citation_count":17,"is_preprint":false},{"pmid":"33452620","id":"PMC_33452620","title":"Morphological study of TNPO3 and SRSF1 interaction during myogenesis by combining confocal, structured illumination and electron microscopy analysis.","date":"2021","source":"Molecular and cellular biochemistry","url":"https://pubmed.ncbi.nlm.nih.gov/33452620","citation_count":16,"is_preprint":false},{"pmid":"32690349","id":"PMC_32690349","title":"Transportin 3 (TNPO3) and related proteins in limb girdle muscular dystrophy D2 muscle biopsies: A morphological study and pathogenetic hypothesis.","date":"2020","source":"Neuromuscular disorders : NMD","url":"https://pubmed.ncbi.nlm.nih.gov/32690349","citation_count":14,"is_preprint":false},{"pmid":"36167471","id":"PMC_36167471","title":"Tnpo3 enables EBF1 function in conditions of antagonistic Notch signaling.","date":"2022","source":"Genes & development","url":"https://pubmed.ncbi.nlm.nih.gov/36167471","citation_count":9,"is_preprint":false},{"pmid":"32581109","id":"PMC_32581109","title":"TNPO3-Mediated Nuclear Entry of the Rous Sarcoma Virus Gag Protein Is Independent of the Cargo-Binding Domain.","date":"2020","source":"Journal of virology","url":"https://pubmed.ncbi.nlm.nih.gov/32581109","citation_count":9,"is_preprint":false},{"pmid":"36789274","id":"PMC_36789274","title":"CRISPR-Cas9 editing of a TNPO3 mutation in a muscle cell model of limb-girdle muscular dystrophy type D2.","date":"2023","source":"Molecular therapy. Nucleic acids","url":"https://pubmed.ncbi.nlm.nih.gov/36789274","citation_count":8,"is_preprint":false},{"pmid":"37339974","id":"PMC_37339974","title":"Tnpo3 controls splicing of the pre-mRNA encoding the canonical TCR α chain of iNKT cells.","date":"2023","source":"Nature communications","url":"https://pubmed.ncbi.nlm.nih.gov/37339974","citation_count":3,"is_preprint":false},{"pmid":"40360518","id":"PMC_40360518","title":"Structural basis of phosphorylation-independent nuclear import of CIRBP by TNPO3.","date":"2025","source":"Nature communications","url":"https://pubmed.ncbi.nlm.nih.gov/40360518","citation_count":1,"is_preprint":false},{"pmid":"41291251","id":"PMC_41291251","title":"Novel insights into the molecular mechanisms of LGMDD2: role of TNPO3 in experimental cell and zebrafish models.","date":"2025","source":"Cellular and molecular life sciences : CMLS","url":"https://pubmed.ncbi.nlm.nih.gov/41291251","citation_count":0,"is_preprint":false},{"pmid":"36082569","id":"PMC_36082569","title":"[Analysis of TNPO3 gene variant and clinical phenotype in a neonate with limb-girdle muscular dystrophies form 1F].","date":"2022","source":"Zhonghua yi xue yi chuan xue za zhi = Zhonghua yixue yichuanxue zazhi = Chinese journal of medical genetics","url":"https://pubmed.ncbi.nlm.nih.gov/36082569","citation_count":0,"is_preprint":false},{"pmid":null,"id":"bio_10.1101_2024.12.27.24319460","title":"Deleterious coding variation associated with autism is consistent across populations, as exemplified by admixed Latin American populations","date":"2025-01-06","source":"bioRxiv","url":"https://doi.org/10.1101/2024.12.27.24319460","citation_count":0,"is_preprint":true}],"cost":{"stage1":{"model":"claude-sonnet-4-6","input_tokens":17877,"output_tokens":4355,"usd":0.059478,"stage1_stop_reason":"end_turn"},"stage2":{"model":"claude-opus-4-8","input_tokens":12270,"output_tokens":4551,"usd":0.087563,"stage2_stop_reason":"end_turn"},"total_usd":0.147041,"stage1_batch_id":"msgbatch_017qqVz33Ga63SCDwzb35XPb","stage2_batch_id":"msgbatch_01Scr2Ei9x4HJMiRGxwrmvLK","note":"batch pricing = 50% of standard"},"stage1_raw":"```json\n{\n  \"discoveries\": [\n    {\n      \"year\": 2009,\n      \"finding\": \"The HIV-1 capsid protein (CA), not integrase, is the dominant viral determinant that dictates TNPO3 dependency during HIV-1 infection, as demonstrated by MLV/HIV-1 chimera virus experiments mapping sensitization to TNPO3 knockdown to the HIV-1 capsid gene.\",\n      \"method\": \"MLV/HIV-1 chimera virus infectivity assay in TNPO3 knockdown cells; in vitro pulldown and surface plasmon resonance for integrase binding\",\n      \"journal\": \"Journal of virology\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — genetic chimera approach with direct functional readout, replicated across multiple retrovirus species, combined with orthogonal in vitro binding assays\",\n      \"pmids\": [\"19846519\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2012,\n      \"finding\": \"Purified recombinant TNPO3 directly stimulates HIV-1 core uncoating in vitro, an effect reduced by RanGTP; TNPO3 and cyclophilin A (CypA) exert opposing effects on HIV-1 uncoating, with CypA inhibiting uncoating and antagonizing TNPO3-mediated stimulation.\",\n      \"method\": \"In vitro uncoating assay with purified recombinant TNPO3 and CypA; TNPO3 depletion in target cells combined with cyclosporine treatment and PF74 sensitivity assay\",\n      \"journal\": \"Journal of virology\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 / Moderate — in vitro reconstitution assay with purified protein, mutagenesis-equivalent (RanGTP competition), confirmed in cellular depletion context\",\n      \"pmids\": [\"23097435\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2013,\n      \"finding\": \"TNPO3 promotes HIV-1 infectivity indirectly by maintaining CPSF6 in the nuclear compartment; TNPO3 knockdown causes cytoplasmic accumulation of CPSF6, which then aberrantly stabilizes the HIV-1 CA core and blocks nuclear import. Rescuing CPSF6 nuclear localization by heterologous NLS fusion reverses the HIV-1 replication defect caused by TNPO3 knockdown.\",\n      \"method\": \"TNPO3 knockdown; CPSF6 mislocalization constructs (NLS deletion, NES fusion, heterologous NLS fusion); fate-of-capsid assay; qPCR for 2-LTR circles; correlation analysis of 27 CA mutants\",\n      \"journal\": \"Retrovirology\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — multiple orthogonal approaches (genetic rescue, localization constructs, capsid stability assay, correlation across 27 mutants), single lab but highly rigorous\",\n      \"pmids\": [\"23414560\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2013,\n      \"finding\": \"TNPO3 depletion inhibits HIV-1 infection and this inhibition requires CPSF6; simultaneous depletion of TNPO3 and CPSF6 rescues HIV-1 infection. Cytosolic full-length CPSF6 blocks HIV-1 infection at the nuclear import step and enhances stability of the HIV-1 CA core.\",\n      \"method\": \"siRNA double-knockdown of TNPO3 and CPSF6; overexpression of cytosolic CPSF6; fate-of-capsid assay; 2-LTR circle quantification\",\n      \"journal\": \"Retrovirology\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — epistasis by double knockdown with defined rescue, orthogonal to PMID:23414560, independently replicating CPSF6 mechanism\",\n      \"pmids\": [\"23622145\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2011,\n      \"finding\": \"TNPO3 promotes HIV-1 infectivity at a step detectable after the preintegration complex arrives in the nucleus (after 2-LTR circle formation but before proviral DNA establishment), and the viral CA protein is the determinant for TNPO3 dependence; 14 of 27 CA mutants rendered HIV-1 TNPO3-independent.\",\n      \"method\": \"Panel of 27 CA mutant single-cycle HIV-1 vectors in TNPO3 knockdown cells; qPCR for viral cDNA, 2-LTR circles, and proviral DNA; lentiviral vector and siRNA TNPO3 KD methods; multiple cell types\",\n      \"journal\": \"Retrovirology\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — systematic mutagenesis screen with multiple orthogonal KD methods, multiple cell types, and step-specific molecular readouts\",\n      \"pmids\": [\"22145813\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2012,\n      \"finding\": \"TNPO3 exists in a monomer-dimer equilibrium in solution and directly binds the HIV-1 intasome (integrase tetramer prebound to viral DNA) but not naked viral DNA or capsid cores in vitro. The interaction interface maps to the HIV-1 IN C-terminal domain and the cargo-binding domain of TNPO3, with specific interacting amino acids identified by mass spectrometry footprinting and site-directed mutagenesis.\",\n      \"method\": \"Circular dichroism, analytical ultracentrifugation, small-angle X-ray scattering, homology modeling; in vitro biochemical binding assays; MS-based protein footprinting; site-directed mutagenesis\",\n      \"journal\": \"The Journal of biological chemistry\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 / Moderate — multiple biophysical methods plus mutagenesis mapping of interaction interface in single rigorous study\",\n      \"pmids\": [\"22872640\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2011,\n      \"finding\": \"Mutations in HIV-1 integrase that abolish TNPO3 binding do not prevent HIV-1 cDNA nuclear import, as shown by 2-LTR circle quantification, indicating IN-TNPO3 interaction is not a major determinant of nuclear import but may function at a nuclear step prior to integration.\",\n      \"method\": \"IN mutant viruses (W131A, Q168L, and others) assessed for TNPO3 binding; 2-LTR circle quantification; integration assay\",\n      \"journal\": \"Retrovirology\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — clean mutant virus system with molecular readouts, but negative finding (no nuclear import defect) from single lab\",\n      \"pmids\": [\"22176773\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2014,\n      \"finding\": \"HIV-1 integrase double mutant R263A/K264A significantly reduces interaction with TNPO3 (2-fold defective) while retaining wild-type reverse transcription activity, and displays a block in both nuclear import and integration as measured by quantitative PCR and a fluorescence-based nuclear import assay.\",\n      \"method\": \"Site-directed mutagenesis of HIV-1 IN; quantitative PCR for nuclear import (2-LTR circles, proviral DNA); eGFP-IN-labeled HIV fluorescence-based import assay; in vitro TNPO3 binding assay\",\n      \"journal\": \"The Journal of biological chemistry\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — specific separation-of-function mutant with multiple orthogonal readouts, single lab\",\n      \"pmids\": [\"25063804\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2013,\n      \"finding\": \"A heterozygous frameshift mutation in TNPO3 causes LGMD1F (limb-girdle muscular dystrophy). Mutant TNPO3 protein localizes around the nucleus but fails to enter the nucleus, unlike wild-type TNPO3.\",\n      \"method\": \"Whole-exome sequencing; Sanger validation; immunolocalization of mutant vs. wild-type TNPO3 in transfected cells\",\n      \"journal\": \"PloS one\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 3 / Moderate — genetic identification plus localization experiment, but localization is the only functional readout and single lab\",\n      \"pmids\": [\"23667635\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2019,\n      \"finding\": \"A second LGMD-causing TNPO3 frameshift mutation (c.2757delC) extends the C-terminal protein product; mutant TNPO3 protein accumulates in subsarcolemmal and perinuclear areas and fails to localize to cytoplasmic annulate lamellae pore complexes. At least one SR-protein cargo (SRSF1/SRRM2) remains normally nuclear, suggesting partial cargo transport is preserved.\",\n      \"method\": \"Gene panel sequencing; TNPO3 mutant construct transfection; immunofluorescence localization; protein expression studies in patient muscle\",\n      \"journal\": \"Neurology. Genetics\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 3 / Moderate — transfection of mutant construct with localization readout, corroborated in patient muscle, two families with similar mutations\",\n      \"pmids\": [\"31192305\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2021,\n      \"finding\": \"During myogenesis, TNPO3 interacts with the SR splicing factor SRSF1 and transports it into the nucleus. SRSF1 remains predominantly nuclear while TNPO3 decreases in the cytoplasm and concentrates in nuclei of differentiated myotubes, indicating dynamic redistribution during myogenic differentiation.\",\n      \"method\": \"Confocal, structured illumination, and electron microscopy of TNPO3 and SRSF1 expression during myogenesis in cell culture\",\n      \"journal\": \"Molecular and cellular biochemistry\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 3 / Moderate — morphological interaction study across multiple imaging modalities, no functional loss-of-function validation\",\n      \"pmids\": [\"33452620\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2022,\n      \"finding\": \"Tnpo3 interacts with the immunoglobulin-like fold domain of transcription factor EBF1, with glutamic acid 271 of EBF1 being a critical residue for the association. Tnpo3 is required for EBF1-mediated B cell programming specifically under antagonistic Notch signaling conditions; B lineage-specific inactivation of Tnpo3 in mice blocks early B cell differentiation with down-regulation of B lineage genes and up-regulation of T and NK lineage genes.\",\n      \"method\": \"Mass spectrometric analysis of EBF1-associated proteins in pro-B cells; retroviral transduction of EBF1 mutants into Ebf1-/- progenitors; RNA-seq; conditional Tnpo3 knockout mice; chromatin binding analysis\",\n      \"journal\": \"Genes & development\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — MS identification of interaction, domain mapping by mutagenesis, in vivo conditional KO with defined differentiation phenotype, RNA-seq\",\n      \"pmids\": [\"36167471\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2020,\n      \"finding\": \"TNPO3 directly binds to RSV Gag protein and mediates its nuclear entry via the matrix (MA) domain NLS, independently of TNPO3's canonical cargo-binding domain (CBD), indicating a non-canonical mechanism of nuclear import.\",\n      \"method\": \"Genetic approach in yeast; direct binding assay between TNPO3 and Gag; CBD deletion mutants of TNPO3; nuclear import assay in avian cells\",\n      \"journal\": \"Journal of virology\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — direct binding demonstrated with CBD domain deletion analysis, confirmed in cellular import assay, single lab\",\n      \"pmids\": [\"32581109\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2023,\n      \"finding\": \"Tnpo3 is required for proper splicing of the Trav11-Traj18-Trac pre-mRNA encoding the semi-invariant TCRα chain of iNKT cells; loss of Tnpo3 blocks iNKT cell development, and this block is rescued by transgenic provision of a pre-spliced rearranged cDNA, demonstrating that Tnpo3 deficiency specifically impairs pre-mRNA splicing rather than iNKT cell development per se.\",\n      \"method\": \"Tnpo3 conditional knockout mice; iNKT cell development analysis; transgenic rescue with rearranged Trav11-Traj18-Trac cDNA\",\n      \"journal\": \"Nature communications\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — in vivo KO with defined molecular phenotype (splicing defect) and definitive genetic rescue experiment distinguishing splicing from developmental role\",\n      \"pmids\": [\"37339974\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2025,\n      \"finding\": \"X-ray crystallography of the TNPO3-CIRBP complex reveals that TNPO3 recognizes a non-classical RSY-NLS in CIRBP where tyrosine residues (not serine/arginine repeats) play a key role in binding, independently of phosphorylation. Serine and tyrosine phosphorylation within CIRBP's NLS actually inhibits TNPO3 binding, revealing a phosphorylation-independent (and phosphorylation-inhibited) nuclear import mechanism.\",\n      \"method\": \"X-ray crystallography of TNPO3-CIRBP complex; NMR; mutagenesis of NLS residues; binding assays with phosphomimetic variants\",\n      \"journal\": \"Nature communications\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 / Moderate — crystal structure with functional validation by mutagenesis and binding assays, single lab but Tier 1 method quality\",\n      \"pmids\": [\"40360518\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2019,\n      \"finding\": \"Interferon-inducible miR-128 directly targets two sites in the TNPO3 mRNA, downregulating TNPO3 protein expression; reduction of TNPO3 levels by miR-128 significantly inhibits HIV-1 replication but not MLV infection, and anti-miR-128 partially neutralizes IFN-mediated HIV-1 restriction.\",\n      \"method\": \"miR-128 overexpression/knockdown in Jurkat cells and primary CD4+ T cells; TNPO3-independent HIV-1 virus challenge; luciferase reporter for TNPO3 mRNA targeting\",\n      \"journal\": \"Journal of virology\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — direct targeting of TNPO3 mRNA by miR-128 confirmed with TNPO3-independent rescue virus, single lab\",\n      \"pmids\": [\"31341054\"],\n      \"is_preprint\": false\n    }\n  ],\n  \"current_model\": \"TNPO3 (Transportin-3/TRN-SR2) is a nuclear import receptor of the importin-β family that transports serine/arginine-rich (SR) splicing factors via recognition of RS/SR-repeat NLSs (and non-classical RSY-NLSs such as in CIRBP) in a mechanism that can be phosphorylation-independent or inhibited by phosphorylation; it also imports the transcription factor EBF1 to enable B cell programming under Notch-antagonistic conditions, regulates pre-mRNA splicing of the iNKT TCRα chain, facilitates HIV-1 replication primarily by maintaining the CA-binding protein CPSF6 in the nucleus (thereby preventing aberrant cytoplasmic CA stabilization) with a secondary direct interaction with the HIV-1 intasome through the integrase C-terminal domain, promotes HIV-1 core uncoating in vitro in a RanGTP-sensitive manner, and harbors loss-of-function mutations that cause the autosomal dominant limb-girdle muscular dystrophy LGMD D2 (formerly LGMD1F) by producing a C-terminally extended protein that mislocalizes away from nuclear pore complexes.\"\n}\n```","stage2_raw":"```json\n{\n  \"mechanistic_narrative\": \"TNPO3 (Transportin-3/TRN-SR2) is a nuclear import receptor of the importin-\\u03b2 family that delivers serine/arginine-rich cargoes through the nuclear pore and serves both gene-expression programs and viral biology [#14, #2]. Structurally, it recognizes non-classical NLSs: crystallography of the TNPO3-CIRBP complex shows that recognition can rely on tyrosine rather than serine/arginine residues and is phosphorylation-independent, with serine/tyrosine phosphorylation within the NLS inhibiting binding [#14]. Through this import activity TNPO3 sustains tissue- and lineage-specific programs: it transports the SR splicing factor SRSF1 during myogenesis [#10], imports the transcription factor EBF1 via its immunoglobulin-like fold domain (critical residue E271) to enable B cell programming under antagonistic Notch signaling [#11], and is required for correct splicing of the semi-invariant TCR\\u03b1 pre-mRNA of iNKT cells, a role definitively separated from a developmental one by transgenic rescue with a pre-spliced cDNA [#13]. In HIV-1 infection, the viral capsid\\u2014not integrase\\u2014is the dominant determinant of TNPO3 dependence [#0, #4], and TNPO3 acts chiefly by maintaining the capsid-binding protein CPSF6 in the nucleus; TNPO3 depletion drives CPSF6 into the cytoplasm where it aberrantly stabilizes the CA core and blocks nuclear import, an effect reversed by restoring CPSF6 nuclear localization or co-depleting CPSF6 [#2, #3]. TNPO3 additionally binds the HIV-1 intasome directly through the integrase C-terminal domain [#5] and stimulates capsid uncoating in vitro in a RanGTP-sensitive manner [#1]. Loss-of-function frameshift mutations in TNPO3 cause autosomal dominant limb-girdle muscular dystrophy (LGMD1F/LGMD D2) by producing a C-terminally extended protein that mislocalizes away from nuclear pore complexes [#8, #9].\"\n  ,\n  \"teleology\": [\n    {\n      \"year\": 2009,\n      \"claim\": \"Established which viral component dictates the requirement for TNPO3, distinguishing capsid- from integrase-driven dependence and reframing TNPO3 as a capsid-linked host factor.\",\n      \"evidence\": \"MLV/HIV-1 chimera virus infectivity in TNPO3-knockdown cells with in vitro integrase binding assays\",\n      \"pmids\": [\"19846519\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Did not define the molecular intermediate connecting capsid to TNPO3\", \"Mechanism of how capsid sensitizes infection to TNPO3 unresolved\"]\n    },\n    {\n      \"year\": 2011,\n      \"claim\": \"Localized TNPO3's pro-viral action to a post-nuclear-entry step and confirmed capsid as the genetic determinant via systematic mutagenesis, narrowing where in the lifecycle TNPO3 acts.\",\n      \"evidence\": \"Panel of 27 CA mutant single-cycle vectors with step-specific qPCR readouts across multiple cell types; IN mutant binding/import assays\",\n      \"pmids\": [\"22145813\", \"22176773\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Did not identify the downstream effector mediating the post-entry block\", \"Negative IN-import finding from a single lab\"]\n    },\n    {\n      \"year\": 2012,\n      \"claim\": \"Provided biochemical and biophysical characterization of TNPO3, including direct intasome binding and in vitro stimulation of capsid uncoating, defining candidate direct molecular activities.\",\n      \"evidence\": \"In vitro uncoating assays with purified TNPO3/CypA and RanGTP; CD, AUC, SAXS, MS footprinting, and mutagenesis mapping of the IN C-terminal domain interaction\",\n      \"pmids\": [\"23097435\", \"22872640\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"In vitro uncoating and intasome binding not shown to be the dominant in-cell mechanism\", \"Relative contribution of direct vs indirect routes unquantified\"]\n    },\n    {\n      \"year\": 2013,\n      \"claim\": \"Resolved the principal HIV-1 mechanism as indirect\\u2014TNPO3 keeps CPSF6 nuclear, and cytoplasmic CPSF6 is the actual inhibitor\\u2014settling whether TNPO3 acts on the virus directly or through a host cargo.\",\n      \"evidence\": \"TNPO3/CPSF6 double knockdown epistasis, CPSF6 mislocalization constructs, heterologous NLS rescue, fate-of-capsid and 2-LTR circle assays across two independent studies\",\n      \"pmids\": [\"23414560\", \"23622145\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Does not exclude a parallel minor direct intasome route\", \"Both studies from converging but limited lab sources\"]\n    },\n    {\n      \"year\": 2013,\n      \"claim\": \"Connected TNPO3 to human disease by identifying a frameshift mutation causing LGMD and showing the mutant fails nuclear entry, establishing a localization-based pathomechanism.\",\n      \"evidence\": \"Whole-exome sequencing with immunolocalization of mutant vs wild-type TNPO3\",\n      \"pmids\": [\"23667635\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Localization was the sole functional readout\", \"Cargo-transport consequences in muscle not measured\"]\n    },\n    {\n      \"year\": 2014,\n      \"claim\": \"Tested whether the direct IN-TNPO3 interaction has a separable functional role using a separation-of-function IN mutant, partially supporting a nuclear-import/integration contribution.\",\n      \"evidence\": \"IN R263A/K264A mutant analysis with qPCR import readouts, fluorescence import assay, and in vitro TNPO3 binding\",\n      \"pmids\": [\"25063804\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Modest (2-fold) binding defect complicates causal attribution\", \"Single lab, not reconciled with the CPSF6-centric model\"]\n    },\n    {\n      \"year\": 2019,\n      \"claim\": \"Extended the disease mechanism with a second mutation producing a C-terminally extended protein that mislocalizes away from pore complexes while partially preserving cargo transport, refining the LGMD pathomechanism.\",\n      \"evidence\": \"Gene panel sequencing, mutant construct transfection, immunofluorescence, and patient muscle protein studies\",\n      \"pmids\": [\"31192305\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Which cargoes are selectively impaired in muscle not defined\", \"Mechanistic link between mislocalization and dystrophy unresolved\"]\n    },\n    {\n      \"year\": 2019,\n      \"claim\": \"Identified an upstream regulatory layer in which IFN-inducible miR-128 directly downregulates TNPO3 to restrict HIV-1, integrating TNPO3 into innate antiviral control.\",\n      \"evidence\": \"miR-128 gain/loss in Jurkat and primary CD4+ T cells, luciferase TNPO3-targeting reporter, and TNPO3-independent rescue virus\",\n      \"pmids\": [\"31341054\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Magnitude of physiological restriction in vivo unknown\", \"Single lab\"]\n    },\n    {\n      \"year\": 2020,\n      \"claim\": \"Showed TNPO3 can import a viral cargo (RSV Gag) through a non-canonical route independent of its canonical cargo-binding domain, broadening the recognition repertoire beyond classical RS-NLS cargoes.\",\n      \"evidence\": \"Yeast genetics, direct TNPO3-Gag binding, CBD-deletion mutants, and nuclear import assay in avian cells\",\n      \"pmids\": [\"32581109\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Structural basis of CBD-independent binding undefined\", \"Generality to other cargoes untested\"]\n    },\n    {\n      \"year\": 2021,\n      \"claim\": \"Linked TNPO3 to muscle biology mechanistically by demonstrating SRSF1 cargo transport and dynamic redistribution during myogenesis, providing a cellular context for the dystrophy phenotype.\",\n      \"evidence\": \"Confocal, structured-illumination, and electron microscopy of TNPO3 and SRSF1 during myogenic differentiation\",\n      \"pmids\": [\"33452620\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"No loss-of-function validation of the transport requirement\", \"Morphological correlation rather than functional perturbation\"]\n    },\n    {\n      \"year\": 2022,\n      \"claim\": \"Defined a non-SR cargo role: TNPO3 imports the transcription factor EBF1 via its Ig-like fold, and is required in vivo for B cell programming under antagonistic Notch signaling, broadening TNPO3 beyond splicing-factor transport.\",\n      \"evidence\": \"MS of EBF1-associated proteins, EBF1 mutant mapping (E271), conditional Tnpo3 knockout mice, RNA-seq, and chromatin binding\",\n      \"pmids\": [\"36167471\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"NLS structural basis of EBF1 recognition not solved\", \"Whether Ig-like-fold recognition generalizes to other TFs unknown\"]\n    },\n    {\n      \"year\": 2023,\n      \"claim\": \"Demonstrated a splicing-specific requirement by showing Tnpo3 is needed for TCR\\u03b1 pre-mRNA splicing in iNKT cells, with transgenic pre-spliced cDNA rescue separating a splicing role from a developmental one.\",\n      \"evidence\": \"Tnpo3 conditional knockout mice, iNKT development analysis, and transgenic rescue with rearranged Trav11-Traj18-Trac cDNA\",\n      \"pmids\": [\"37339974\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Direct splicing-factor cargo responsible not pinpointed\", \"Generality of splicing role to other transcripts untested\"]\n    },\n    {\n      \"year\": 2025,\n      \"claim\": \"Provided the structural rules of cargo recognition, showing TNPO3 binds a non-classical RSY-NLS via tyrosine residues in a phosphorylation-independent, phosphorylation-inhibited manner, revising the canonical RS-repeat/phospho-dependent model.\",\n      \"evidence\": \"X-ray crystallography and NMR of the TNPO3-CIRBP complex with NLS mutagenesis and phosphomimetic binding assays\",\n      \"pmids\": [\"40360518\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"How phosphorylation switches differ across cargoes not generalized\", \"Structural basis for RanGTP-regulated cargo release in this complex not addressed\"]\n    },\n    {\n      \"year\": null,\n      \"claim\": \"How TNPO3's diverse cargo-recognition modes (RS-NLS, RSY-NLS, Ig-fold TFs, CBD-independent viral cargoes) are coordinated and which selective cargo failures drive muscle pathology remain unresolved.\",\n      \"evidence\": \"\",\n      \"pmids\": [],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"No unified structural model spanning all cargo classes\", \"Disease-causing mutant's cargo-specific transport deficits in muscle undefined\", \"RanGTP-dependent cargo release mechanism not structurally resolved for most cargoes\"]\n    }\n  ],\n  \"mechanism_profile\": {\n    \"molecular_activity\": [\n      {\"term_id\": \"GO:0140104\", \"supporting_discovery_ids\": [14, 11, 10, 12]},\n      {\"term_id\": \"GO:0060090\", \"supporting_discovery_ids\": [2, 14]}\n    ],\n    \"localization\": [\n      {\"term_id\": \"GO:0005635\", \"supporting_discovery_ids\": [8, 9]},\n      {\"term_id\": \"GO:0005829\", \"supporting_discovery_ids\": [10, 2]},\n      {\"term_id\": \"GO:0005634\", \"supporting_discovery_ids\": [10, 14]}\n    ],\n    \"pathway\": [\n      {\"term_id\": \"GO:0009609507\", \"supporting_discovery_ids\": [14, 11]},\n      {\"term_id\": \"R-HSA-9609507\", \"supporting_discovery_ids\": [14, 11, 10]},\n      {\"term_id\": \"R-HSA-1643685\", \"supporting_discovery_ids\": [8, 9]},\n      {\"term_id\": \"R-HSA-168256\", \"supporting_discovery_ids\": [11, 13]},\n      {\"term_id\": \"R-HSA-8953854\", \"supporting_discovery_ids\": [13]}\n    ],\n    \"complexes\": [],\n    \"partners\": [\"CPSF6\", \"SRSF1\", \"EBF1\", \"CIRBP\"],\n    \"other_free_text\": []\n  }\n}\n```","audit_flag":null,"evaluation":{"pairwise":"win","faith_supported":6,"faith_total":6,"faith_pct":100.0}}