{"gene":"FKBP3","run_date":"2026-06-09T23:54:43","timeline":{"discoveries":[{"year":1992,"finding":"FKBP25 (FKBP3) has peptidylprolyl cis-trans isomerase (PPIase) activity comparable to FKBP12, binds rapamycin with higher affinity than FK506 (IC50 ~50 nM vs ~400 nM), and its PPIase activity is inhibited by rapamycin and FK506.","method":"In vitro PPIase assay with recombinant FKBP25; inhibition assays with rapamycin and FK506","journal":"The Journal of biological chemistry","confidence":"High","confidence_rationale":"Tier 1 / Strong — direct in vitro enzymatic assay with recombinant protein, replicated in subsequent studies","pmids":["1375932"],"is_preprint":false},{"year":1992,"finding":"FKBP25 contains putative nuclear localization sequences in its sequence, distinguishing it from FKBP12 and FKBP13.","method":"cDNA cloning and sequence analysis","journal":"The Journal of biological chemistry","confidence":"Medium","confidence_rationale":"Tier 3 / Moderate — sequence-based prediction confirmed by later fractionation experiments in other papers","pmids":["1375932"],"is_preprint":false},{"year":1993,"finding":"FKBP25 is predominantly localized to the nuclear fraction of T-lymphoma Jurkat cells and has the ability to bind DNA; the FKBP25/DNA complex is dissociable by high salt. FKBP12, which shares C-terminal homology with FKBP25, does not bind DNA.","method":"Cell fractionation, Western blotting with anti-FKBP25 antibodies, DNA-binding assay","journal":"FEBS letters","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — direct fractionation with functional DNA-binding assay, single lab, negative control with FKBP12","pmids":["8422914"],"is_preprint":false},{"year":1999,"finding":"FKBP25 (FKBP3) expression is down-regulated following p53 induction by DNA-damaging stimuli in human and murine cell lines, identifying it as a transcriptional repression target of p53.","method":"Differential gene expression screening following p53 induction; confirmed in multiple cell lines with DNA-damaging stimuli","journal":"Oncogene","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — multiple cell lines and stimuli tested, but no direct mechanistic dissection of repression mechanism","pmids":["10557083"],"is_preprint":false},{"year":2000,"finding":"Endogenous FKBP25 from porcine brain co-isolates with high-mobility group II protein (HMG-II), and a residual pool associates with Rab5, guanylyl kinase, and phosphatidylethanolamine-binding protein.","method":"Immunoblotting, 2D-PAGE, Edman degradation, MALDI-TOF of anti-FKBP25 immunoprecipitates from porcine brain","journal":"Archives of biochemistry and biophysics","confidence":"Low","confidence_rationale":"Tier 3 / Weak — single lab, co-migration/co-purification without reciprocal validation","pmids":["10900128"],"is_preprint":false},{"year":2009,"finding":"FKBP25 interacts with MDM2, stimulates MDM2 auto-ubiquitylation and proteasomal degradation, leading to induction of p53 and p21. Depletion of FKBP25 by siRNA increases MDM2 levels and reduces p53 and p21.","method":"Co-immunoprecipitation, siRNA knockdown, western blotting for MDM2/p53/p21 levels","journal":"FEBS letters","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — Co-IP plus functional siRNA knockdown with multiple readouts, single lab","pmids":["19166840"],"is_preprint":false},{"year":2013,"finding":"Mutagenesis of FKBP25 revealed that certain mutations destabilize the FKBD domain fold entirely, while other 'surgical' mutations specifically ablate PPIase catalytic activity while maintaining domain structure, enabling distinction between catalytic and non-catalytic functions.","method":"Site-directed mutagenesis, in vitro PPIase assays, structural characterization of mutants","journal":"Biochemical Society transactions","confidence":"Medium","confidence_rationale":"Tier 1 / Moderate — mutagenesis with in vitro enzymatic validation, single lab","pmids":["23697935"],"is_preprint":false},{"year":2014,"finding":"FKBP25 interacts with nucleolin in an rRNA-dependent manner and associates with the immature pre-60S ribosomal subunit in nuclear extract but not with mature ribosomes, implicating FKBP25 in ribosome biogenesis.","method":"Proteomic characterization (mass spectrometry of immunoprecipitates), Co-IP with RNase treatment to assess rRNA-dependence","journal":"RNA (New York, N.Y.)","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — Co-IP with MS and orthogonal RNase sensitivity test, single lab","pmids":["24840943"],"is_preprint":false},{"year":2014,"finding":"The N-terminal domain of FKBP25 (residues 1–73) adopts a novel 5-helix bundle structure termed the Basic Tilted Helix Bundle (BTHB) with a positively charged surface patch, which is implicated in DNA binding.","method":"NMR/structural determination of FKBP25(1-73), comparative structural analysis with HectD1","journal":"Biochemical and biophysical research communications","confidence":"Medium","confidence_rationale":"Tier 1 / Moderate — structure determined by NMR, functional inference from conserved surface; single lab","pmids":["24667607"],"is_preprint":false},{"year":2014,"finding":"Endogenous FKBP25 associates with core histones, spliceosomal complexes, ribosomal subunits, and polyribosomes; FKBP25 from polyribosomes can be released by added RNA or high salt, indicating RNA-mediated interaction. Rapamycin or FK506 only partially release FKBP25, suggesting additional protein-mediated binding via the PPIase cavity.","method":"Anti-FKBP25 immunoprecipitation from HeLa, K568, and porcine brain cells; polyribosome fractionation; salt/RNA competition; proteomics","journal":"Biochemical and biophysical research communications","confidence":"Low","confidence_rationale":"Tier 3 / Weak — single lab, fractionation and pulldown without full mechanistic dissection","pmids":["24998444"],"is_preprint":false},{"year":2015,"finding":"FKBP25 associates with TRPC6 (and TRPC3), and its interaction pattern with TRPCs is modified by FK506. Knockdown of FKBP25 significantly inhibits OAG-evoked non-capacitative calcium entry (NCCE) in MEG-01 and HEK293 cells. Biotinylation experiments indicate FKBP25 localizes to the plasma membrane in addition to intracellular compartments.","method":"Co-immunoprecipitation, siRNA knockdown, calcium entry assays, surface biotinylation in platelets and cell lines","journal":"Biochimica et biophysica acta","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — Co-IP plus functional siRNA knockdown with calcium flux readouts, single lab","pmids":["26239116"],"is_preprint":false},{"year":2016,"finding":"FKBP25 forms a rapamycin-induced ternary complex with the FRB domain of mTOR, and the crystal structure of FRB-rapamycin-FKBP25 was determined at 1.67-Å resolution, revealing conformational changes in FRB and covalent metalloid coordination at C2085 of FRB.","method":"Proximity biotin labeling (pBirA), immunoprecipitation, immunofluorescence, X-ray crystallography","journal":"ACS central science","confidence":"High","confidence_rationale":"Tier 1 / Strong — crystal structure at 1.67 Å plus orthogonal biochemical confirmation; multiple methods in one study","pmids":["27610411"],"is_preprint":false},{"year":2016,"finding":"NMR solution structure of full-length human FKBP25 shows the N-terminal HLH domain and C-terminal FKBD interact with each other; both domains participate in DNA binding, with the HLH domain making major-groove contacts and a basic FKBD loop contributing to minor-groove interactions.","method":"NMR structure determination, NMR-monitored DNA titration, mutagenesis, FKBP25-DNA complex modeling","journal":"Nucleic acids research","confidence":"High","confidence_rationale":"Tier 1 / Strong — full-length NMR structure with mutagenesis and functional DNA binding validation in one study","pmids":["26762975"],"is_preprint":false},{"year":2017,"finding":"The N-terminal BTHB domain of FKBP25 selectively binds double-stranded RNA (dsRNA) over DNA or single-stranded oligonucleotides. This RNA-binding activity is required for FKBP25's nucleolar localization and for the vast majority of its protein interactions, including those with pre-60S ribosomes and early ribosome biogenesis factors.","method":"Biochemical dsRNA-binding assays, mutagenesis, immunofluorescence for localization, Co-IP for protein interactions in BTHB mutants","journal":"Nucleic acids research","confidence":"High","confidence_rationale":"Tier 1 / Strong — multiple orthogonal methods (binding assay, mutagenesis, localization, Co-IP) in one study","pmids":["29036638"],"is_preprint":false},{"year":2017,"finding":"FKBP3 promotes NSCLC cell proliferation via a pathway in which FKBP3 inhibits ubiquitination of Sp1, stabilizing Sp1, which drives HDAC2 promoter activity, increasing HDAC2 expression, leading to HDAC2-mediated deacetylation of histone H3K4 at the p27 promoter and reduced p27 expression.","method":"siRNA knockdown, overexpression, ChIP (HDAC2 at p27 promoter), ubiquitination assay for Sp1, western blotting, in vitro and in vivo proliferation assays","journal":"Theranostics","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — multiple mechanistic steps validated with orthogonal methods in one lab","pmids":["28839465"],"is_preprint":false},{"year":2018,"finding":"The FKBP domain of FKBP25 directly binds microtubules in vitro to promote their polymerization and stabilize the MT network. FKBP25 associates with the mitotic spindle, regulates entry into mitosis, and its knockdown leads to increased chromosome instability. Additionally, FKBP25 association with chromatin is regulated by Protein Kinase C (PKC) phosphorylation in a cell-cycle-dependent manner, disrupting FKBP25-DNA contacts during mitosis while maintaining spindle association.","method":"Direct microtubule-binding assay (in vitro), mitotic spindle immunofluorescence, chromosome instability assays in FKBP25 knockdown cells, PKC phosphorylation assays","journal":"Nucleic acids research","confidence":"High","confidence_rationale":"Tier 1 / Strong — in vitro reconstitution of microtubule binding, multiple cellular assays, and phosphorylation mechanism in one study","pmids":["29361176"],"is_preprint":false},{"year":2018,"finding":"FKBP25 translocates from cytoplasm to nucleus in endothelial cells following oxygen-glucose deprivation (OGD) or peroxynitrite treatment, and in the nucleus interacts with 60S ribosomal protein L7a. Overexpression of FKBP25 protects endothelial cells against OGD injury.","method":"Western blot and immunofluorescence (localization), co-immunoprecipitation and FRET (interaction with L7a), overexpression survival assay","journal":"Cellular physiology and biochemistry","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — Co-IP plus FRET plus localization in stress conditions, single lab","pmids":["29969783"],"is_preprint":false},{"year":2019,"finding":"FKBP25's catalytic (PPIase) activity is required for promoting homologous recombination (HR) repair of DNA double-strand breaks (DSBs). FKBP25-depleted cells form fewer Rad51 repair foci and become dependent on Rad52-mediated single-strand annealing (SSA). Rapamycin treatment also impairs HR, at least partly independently of mTOR.","method":"FKBP25 knockdown, catalytic mutant expression, Rad51/Rad52 focus assays (immunofluorescence), genetic epistasis (Rad52 dependence), DSB reporter assays","journal":"Biochemistry and cell biology","confidence":"High","confidence_rationale":"Tier 1 / Strong — catalytic mutant plus multiple orthogonal cellular DNA repair assays, epistasis with Rad52","pmids":["30620620"],"is_preprint":false},{"year":2019,"finding":"FKBP3 downregulation in colorectal cancer cells reduces HDAC2 expression and P-gp levels, reduces p-AKT, and increases PTEN and cleaved caspase-3, sensitizing cells to oxaliplatin. Upregulation of HDAC2 counteracts FKBP3 knockdown-induced sensitization.","method":"siRNA knockdown and overexpression, western blotting, flow cytometry (apoptosis), rescue by HDAC2 overexpression","journal":"Oncology reports","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — functional rescue experiment confirms HDAC2 as downstream effector; single lab","pmids":["31524278"],"is_preprint":false},{"year":2021,"finding":"FKBP3 indirectly binds the HIV-1 LTR through physical interaction with the transcription factor YY1, thereby recruiting HDAC1/2 to the LTR, promoting histone deacetylation and HIV-1 latency. FKBP3 knockout in latently infected cells activates latent HIV-1.","method":"CRISPR knockout in latent HIV-1 cell lines, Co-IP (FKBP3-YY1 interaction), ChIP (HDAC1/2 at LTR), primary latent cell model","journal":"mBio","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — CRISPR KO with ChIP and Co-IP, multiple latent cell line models, single lab","pmids":["34281390"],"is_preprint":false},{"year":2021,"finding":"Esterase D (ESD) interacts with FKBP25 via the N-terminal 1–90 aa domain of FKBP25. ESD reduces K48-linked poly-ubiquitin chains on FKBP25, thereby stabilizing cytoplasmic FKBP25. Stabilized FKBP25 then binds more mTORC1, suppressing mTORC1 activity and promoting autophagy.","method":"Yeast two-hybrid, Co-IP (endogenous and GFP-tagged FKBP25), ubiquitination assay, western blot for mTORC1/P70S6K/4EBP1 phosphorylation","journal":"Cellular & molecular biology letters","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — yeast two-hybrid confirmed by Co-IP, ubiquitination assay, and functional mTORC1 readout; single lab","pmids":["34875997"],"is_preprint":false},{"year":2021,"finding":"FKBP25 depletion in mouse oocytes causes abnormal spindle assembly, chromosome misalignment, and defective kinetochore-microtubule attachment, leading to elevated aneuploidy. Phosphorylation at serine 163 is identified as a key regulatory modification for FKBP25's function in meiotic maturation.","method":"siRNA knockdown in mouse oocytes, immunofluorescence (spindle/chromosome), aneuploidy scoring, site-directed mutagenesis (S163A)","journal":"Frontiers in cell and developmental biology","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — knockdown with multiple cellular readouts plus mutagenesis, single lab","pmids":["33553183"],"is_preprint":false},{"year":2023,"finding":"FKBP3 interacts with PARK7 (DJ-1); knockdown of FKBP3 enhances ubiquitination and degradation of PARK7, reducing Wnt/β-catenin pathway activation in DLBCL cells.","method":"Co-immunoprecipitation (FKBP3-PARK7 interaction), ubiquitination assay, western blotting for β-catenin pathway components, FKBP3 knockdown","journal":"Journal of cellular and molecular medicine","confidence":"Low","confidence_rationale":"Tier 3 / Weak — single Co-IP without reciprocal validation, single lab, limited mechanistic follow-up","pmids":["37987202"],"is_preprint":false},{"year":2023,"finding":"FKBP25 knockdown in C2C12 myoblasts increases cell accumulation/viability and migration in vitro, independently of changes in tubulin dynamics. FKBP25 protein expression increases in hypertrophy (chronic mechanical overload) and muscle regeneration (mdx model) but decreases in denervation atrophy in vivo.","method":"Doxycycline-inducible knockdown in C2C12 cells, viability and migration assays, tubulin dynamics measurements, in vivo muscle adaptation models with western blotting","journal":"The FEBS journal","confidence":"Low","confidence_rationale":"Tier 3 / Weak — single lab, knockdown with phenotypic readout but limited pathway placement","pmids":["37345229"],"is_preprint":false}],"current_model":"FKBP25 (FKBP3) is a nuclear prolyl isomerase with a C-terminal FK506-binding/PPIase domain (which binds rapamycin with high affinity, promotes microtubule polymerization, and is required for homologous recombination repair of DNA double-strand breaks) and an N-terminal BTHB domain (which selectively binds dsRNA to drive nucleolar localization and ribosome biogenesis interactions, and also contacts DNA major-groove); its chromatin association is cell-cycle regulated by PKC-mediated phosphorylation, it controls cell proliferation through an FKBP3→Sp1 ubiquitination→HDAC2→p27 axis, it promotes MDM2 degradation to activate p53, it binds YY1 to recruit HDAC1/2 to gene promoters, and its stability is regulated by ESD-mediated suppression of K48-linked ubiquitination which in turn modulates mTORC1 activity."},"narrative":{"mechanistic_narrative":"FKBP3 (FKBP25) is a predominantly nuclear peptidylprolyl cis-trans isomerase that couples prolyl isomerase activity to nucleic-acid binding to act in ribosome biogenesis, chromatin regulation, genome maintenance, and the mitotic apparatus [PMID:1375932, PMID:26762975]. It is a two-domain protein: a C-terminal FK506-binding/PPIase domain that hydrolyzes peptidyl-prolyl bonds, binds rapamycin with higher affinity than FK506, and is inhibited by both drugs [PMID:1375932]; and an N-terminal Basic Tilted Helix Bundle (BTHB/HLH) domain that, together with a basic FKBD loop, makes major- and minor-groove contacts on DNA [PMID:24667607, PMID:26762975] and selectively binds double-stranded RNA — an activity that drives its nucleolar localization and the bulk of its protein interactions, including with nucleolin and the immature pre-60S ribosomal subunit [PMID:24840943, PMID:29036638]. The catalytic PPIase activity is required for homologous-recombination repair of DNA double-strand breaks, with FKBP3-depleted cells forming fewer Rad51 foci and shifting toward Rad52-dependent single-strand annealing [PMID:30620620]. The FKBD domain directly binds and stabilizes microtubules, and FKBP3 associates with the mitotic spindle and safeguards chromosome stability, with its chromatin association switched off during mitosis by PKC-mediated phosphorylation [PMID:29361176]. FKBP3 also operates as a chromatin regulator and proliferation driver: it stabilizes Sp1 to upregulate HDAC2 and repress p27 [PMID:28839465], and binds the transcription factor YY1 to recruit HDAC1/2 to target promoters, a mechanism it uses to enforce HIV-1 latency at the viral LTR [PMID:34281390]. It additionally tunes the p53 axis by stimulating MDM2 auto-ubiquitylation and degradation [PMID:19166840] and, via ESD-mediated suppression of K48-linked ubiquitination that stabilizes the cytoplasmic pool, suppresses mTORC1 activity to promote autophagy [PMID:34875997]. FKBP3 forms a rapamycin-induced ternary complex with the FRB domain of mTOR, defined at atomic resolution [PMID:27610411].","teleology":[{"year":1992,"claim":"Established that FKBP3 is an enzymatically active prolyl isomerase distinct from FKBP12, defining its core catalytic identity and drug sensitivity.","evidence":"In vitro PPIase assays with recombinant protein and rapamycin/FK506 inhibition; cDNA cloning revealing nuclear localization sequences","pmids":["1375932"],"confidence":"High","gaps":["Physiological substrates of the PPIase activity not identified","Nuclear targeting predicted from sequence, not functionally tested at this stage"]},{"year":1993,"claim":"Showed that FKBP3 is nuclear and binds DNA, a property absent in homologous FKBP12, pointing to a chromatin-associated function beyond isomerase activity.","evidence":"Cell fractionation and DNA-binding assays in Jurkat cells with FKBP12 negative control","pmids":["8422914"],"confidence":"Medium","gaps":["DNA sequence specificity not defined","Domain responsible for DNA binding not mapped"]},{"year":1999,"claim":"Placed FKBP3 downstream of the p53 stress response by identifying it as a transcriptionally repressed p53 target during DNA damage.","evidence":"Differential expression screening after p53 induction across multiple cell lines and damaging stimuli","pmids":["10557083"],"confidence":"Medium","gaps":["Mechanism of p53-mediated repression not dissected","Functional consequence of downregulation not established here"]},{"year":2009,"claim":"Defined a feedback link to p53 by showing FKBP3 stimulates MDM2 auto-ubiquitylation, thereby raising p53 and p21.","evidence":"Co-IP plus siRNA knockdown with MDM2/p53/p21 western readouts","pmids":["19166840"],"confidence":"Medium","gaps":["Whether PPIase activity is required not tested","Single lab, no reciprocal/structural validation of FKBP3-MDM2 contact"]},{"year":2014,"claim":"Connected FKBP3 to ribosome biogenesis and solved the N-terminal domain fold, establishing a structural basis for its nucleic-acid engagement.","evidence":"MS of immunoprecipitates with RNase-sensitivity tests showing nucleolin/pre-60S association; NMR structure of FKBP25(1-73) defining the BTHB fold","pmids":["24840943","24667607"],"confidence":"Medium","gaps":["Direct catalytic role in ribosome assembly not shown","BTHB nucleic-acid selectivity not yet resolved at this stage"]},{"year":2016,"claim":"Provided full-length and complex structures showing inter-domain communication, dual-groove DNA binding, and the rapamycin-bridged FKBP3-mTOR(FRB) ternary complex.","evidence":"NMR of full-length FKBP25 with DNA titration and mutagenesis; X-ray crystallography of FRB-rapamycin-FKBP25 at 1.67 Å","pmids":["26762975","27610411"],"confidence":"High","gaps":["In vivo significance of the mTOR ternary complex not established","Whether DNA binding and PPIase act simultaneously unclear"]},{"year":2017,"claim":"Resolved the molecular logic of FKBP3 localization by showing the BTHB domain selectively binds dsRNA, which drives nucleolar targeting and most of its interactome.","evidence":"dsRNA-binding assays, mutagenesis, immunofluorescence, and Co-IP in BTHB mutants","pmids":["29036638"],"confidence":"High","gaps":["Specific dsRNA targets in cells not identified","Relationship between dsRNA and DNA binding modes not reconciled"]},{"year":2017,"claim":"Defined a proliferation-driving chromatin axis whereby FKBP3 stabilizes Sp1 to upregulate HDAC2 and repress p27.","evidence":"siRNA/overexpression, Sp1 ubiquitination assay, HDAC2-p27 promoter ChIP, and proliferation assays in NSCLC models","pmids":["28839465"],"confidence":"Medium","gaps":["Whether PPIase activity drives Sp1 stabilization not tested","Direct enzyme-substrate relationship with Sp1 not shown"]},{"year":2018,"claim":"Established a structural role at the mitotic apparatus, showing the FKBD domain directly stabilizes microtubules and that cell-cycle PKC phosphorylation toggles chromatin versus spindle engagement.","evidence":"In vitro microtubule-binding assay, mitotic spindle immunofluorescence, chromosome instability assays, and PKC phosphorylation assays","pmids":["29361176"],"confidence":"High","gaps":["PKC site(s) and kinase specificity in vivo not fully mapped","How spindle and DNA-repair roles are temporally coordinated unclear"]},{"year":2019,"claim":"Demonstrated that the catalytic PPIase activity is required for homologous-recombination repair, assigning an enzymatic function to genome maintenance.","evidence":"Catalytic-mutant rescue, Rad51/Rad52 focus assays, Rad52 epistasis, and DSB reporter assays","pmids":["30620620"],"confidence":"High","gaps":["HR substrate isomerized by FKBP3 not identified","Rapamycin's mTOR-independent HR effect not fully delineated"]},{"year":2021,"claim":"Extended the YY1/HDAC repression mechanism to viral chromatin, showing FKBP3 enforces HIV-1 latency by recruiting HDAC1/2 to the LTR via YY1.","evidence":"CRISPR knockout in latent HIV models, FKBP3-YY1 Co-IP, and HDAC1/2 LTR ChIP","pmids":["34281390"],"confidence":"Medium","gaps":["Whether DNA/PPIase activities contribute beyond YY1 bridging unclear","Generality to host gene promoters not defined"]},{"year":2021,"claim":"Identified an ESD-controlled stability switch that links FKBP3 abundance to mTORC1 suppression and autophagy.","evidence":"Yeast two-hybrid, Co-IP, K48-ubiquitination assay, and mTORC1/P70S6K/4EBP1 phospho-readouts","pmids":["34875997"],"confidence":"Medium","gaps":["Identity of the E3 ligase opposed by ESD unknown","Relationship to the rapamycin-induced FRB ternary complex not integrated"]},{"year":2021,"claim":"Confirmed an in vivo meiotic requirement, with FKBP3 needed for proper spindle assembly and kinetochore-microtubule attachment in oocytes.","evidence":"siRNA knockdown in mouse oocytes, spindle/chromosome immunofluorescence, aneuploidy scoring, and S163A mutagenesis","pmids":["33553183"],"confidence":"Medium","gaps":["Mechanism linking S163 phosphorylation to function not resolved","Whether the microtubule-binding FKBD mediates this directly not shown"]},{"year":null,"claim":"The endogenous prolyl-isomerase substrates that underlie FKBP3's roles in HR repair, ribosome biogenesis, and chromatin regulation remain unidentified, leaving the unifying catalytic mechanism open.","evidence":"No direct substrate identification across the available corpus","pmids":[],"confidence":"Low","gaps":["No physiological PPIase substrate validated","How nucleic-acid binding, catalysis, and protein scaffolding are integrated into one mechanism is unresolved"]}],"mechanism_profile":{"molecular_activity":[{"term_id":"GO:0016853","term_label":"isomerase activity","supporting_discovery_ids":[0,6,17]},{"term_id":"GO:0003723","term_label":"RNA binding","supporting_discovery_ids":[7,13]},{"term_id":"GO:0003677","term_label":"DNA binding","supporting_discovery_ids":[2,8,12]},{"term_id":"GO:0008092","term_label":"cytoskeletal protein binding","supporting_discovery_ids":[15]},{"term_id":"GO:0140110","term_label":"transcription regulator activity","supporting_discovery_ids":[14,19]}],"localization":[{"term_id":"GO:0005634","term_label":"nucleus","supporting_discovery_ids":[1,2]},{"term_id":"GO:0005730","term_label":"nucleolus","supporting_discovery_ids":[13]},{"term_id":"GO:0005829","term_label":"cytosol","supporting_discovery_ids":[20]}],"pathway":[{"term_id":"R-HSA-73894","term_label":"DNA Repair","supporting_discovery_ids":[17]},{"term_id":"R-HSA-8953854","term_label":"Metabolism of RNA","supporting_discovery_ids":[7,13]},{"term_id":"R-HSA-1640170","term_label":"Cell Cycle","supporting_discovery_ids":[15,21]},{"term_id":"R-HSA-4839726","term_label":"Chromatin organization","supporting_discovery_ids":[14,19]},{"term_id":"R-HSA-162582","term_label":"Signal Transduction","supporting_discovery_ids":[11,20]}],"complexes":["FKBP25-rapamycin-mTOR(FRB) ternary complex","pre-60S ribosomal subunit"],"partners":["MDM2","NUCLEOLIN","SP1","YY1","HDAC1","HDAC2","MTOR","ESD"],"other_free_text":[]}},"prefetch_data":{"uniprot":{"accession":"Q00688","full_name":"Peptidyl-prolyl cis-trans isomerase FKBP3","aliases":["25 kDa FK506-binding protein","25 kDa FKBP","FKBP-25","FK506-binding protein 3","FKBP-3","Immunophilin FKBP25","Rapamycin-selective 25 kDa immunophilin","Rotamase"],"length_aa":224,"mass_kda":25.2,"function":"FK506- and rapamycin-binding proteins (FKBPs) constitute a family of receptors for the two immunosuppressants which inhibit T-cell proliferation by arresting two distinct cytoplasmic signal transmission pathways. PPIases accelerate the folding of proteins","subcellular_location":"Nucleus","url":"https://www.uniprot.org/uniprotkb/Q00688/entry"},"depmap":{"release":"DepMap","has_data":true,"is_common_essential":false,"resolved_as":"","url":"https://depmap.org/portal/gene/FKBP3","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/FKBP3","total_profiled":1310},"omim":[{"mim_id":"618649","title":"HECT DOMAIN E3 UBIQUITIN PROTEIN LIGASE 1; HECTD1","url":"https://www.omim.org/entry/618649"},{"mim_id":"186947","title":"FK506-BINDING PROTEIN 3; FKBP3","url":"https://www.omim.org/entry/186947"}],"hpa":{"profiled":true,"resolved_as":"","reliability":"Supported","locations":[{"location":"Cytosol","reliability":"Supported"}],"tissue_specificity":"Tissue enhanced","tissue_distribution":"Detected in all","driving_tissues":[{"tissue":"skeletal muscle","ntpm":397.6},{"tissue":"tongue","ntpm":347.3}],"url":"https://www.proteinatlas.org/search/FKBP3"},"hgnc":{"alias_symbol":["FKBP-25","FKBP25"],"prev_symbol":[]},"alphafold":{"accession":"Q00688","domains":[{"cath_id":"1.10.720.80","chopping":"11-68","consensus_level":"high","plddt":83.381,"start":11,"end":68},{"cath_id":"3.10.50.40","chopping":"110-222","consensus_level":"high","plddt":87.379,"start":110,"end":222}],"viewer_url":"https://alphafold.ebi.ac.uk/entry/Q00688","model_url":"https://alphafold.ebi.ac.uk/files/AF-Q00688-F1-model_v6.cif","pae_url":"https://alphafold.ebi.ac.uk/files/AF-Q00688-F1-predicted_aligned_error_v6.png","plddt_mean":77.06},"mouse_models":{"mgi_url":"https://www.informatics.jax.org/marker/summary?nomen=FKBP3","jax_strain_url":"https://www.jax.org/strain/search?query=FKBP3"},"sequence":{"accession":"Q00688","fasta_url":"https://rest.uniprot.org/uniprotkb/Q00688.fasta","uniprot_url":"https://www.uniprot.org/uniprotkb/Q00688/entry","alphafold_viewer_url":"https://alphafold.ebi.ac.uk/entry/Q00688"}},"corpus_meta":[{"pmid":"10557083","id":"PMC_10557083","title":"Down-regulation of the stathmin/Op18 and FKBP25 genes following p53 induction.","date":"1999","source":"Oncogene","url":"https://pubmed.ncbi.nlm.nih.gov/10557083","citation_count":110,"is_preprint":false},{"pmid":"1375932","id":"PMC_1375932","title":"Molecular cloning of a 25-kDa high affinity rapamycin binding protein, FKBP25.","date":"1992","source":"The Journal of biological chemistry","url":"https://pubmed.ncbi.nlm.nih.gov/1375932","citation_count":77,"is_preprint":false},{"pmid":"19166840","id":"PMC_19166840","title":"FKBP25, a novel regulator of the p53 pathway, induces the degradation of MDM2 and activation of p53.","date":"2009","source":"FEBS letters","url":"https://pubmed.ncbi.nlm.nih.gov/19166840","citation_count":58,"is_preprint":false},{"pmid":"28839465","id":"PMC_28839465","title":"FKBP3 Promotes Proliferation of Non-Small Cell Lung Cancer Cells through Regulating Sp1/HDAC2/p27.","date":"2017","source":"Theranostics","url":"https://pubmed.ncbi.nlm.nih.gov/28839465","citation_count":54,"is_preprint":false},{"pmid":"27610411","id":"PMC_27610411","title":"Proximity-Directed Labeling Reveals a New Rapamycin-Induced Heterodimer of FKBP25 and FRB in Live Cells.","date":"2016","source":"ACS central science","url":"https://pubmed.ncbi.nlm.nih.gov/27610411","citation_count":47,"is_preprint":false},{"pmid":"8422914","id":"PMC_8422914","title":"On the localization of FKBP25 in T-lymphocytes.","date":"1993","source":"FEBS letters","url":"https://pubmed.ncbi.nlm.nih.gov/8422914","citation_count":43,"is_preprint":false},{"pmid":"31955008","id":"PMC_31955008","title":"Melatonin Regulates Breast Cancer Progression by the lnc010561/miR-30/FKBP3 Axis.","date":"2019","source":"Molecular therapy. Nucleic acids","url":"https://pubmed.ncbi.nlm.nih.gov/31955008","citation_count":32,"is_preprint":false},{"pmid":"31524278","id":"PMC_31524278","title":"FKBP3 mediates oxaliplatin resistance in colorectal cancer cells by regulating HDAC2 expression.","date":"2019","source":"Oncology reports","url":"https://pubmed.ncbi.nlm.nih.gov/31524278","citation_count":29,"is_preprint":false},{"pmid":"26762975","id":"PMC_26762975","title":"Structural basis of nucleic acid recognition by FK506-binding protein 25 (FKBP25), a nuclear immunophilin.","date":"2016","source":"Nucleic acids research","url":"https://pubmed.ncbi.nlm.nih.gov/26762975","citation_count":26,"is_preprint":false},{"pmid":"24840943","id":"PMC_24840943","title":"The prolyl isomerase, FKBP25, interacts with RNA-engaged nucleolin and the pre-60S ribosomal subunit.","date":"2014","source":"RNA (New York, N.Y.)","url":"https://pubmed.ncbi.nlm.nih.gov/24840943","citation_count":24,"is_preprint":false},{"pmid":"29361176","id":"PMC_29361176","title":"The prolyl isomerase FKBP25 regulates microtubule polymerization impacting cell cycle progression and genomic stability.","date":"2018","source":"Nucleic acids research","url":"https://pubmed.ncbi.nlm.nih.gov/29361176","citation_count":18,"is_preprint":false},{"pmid":"24998444","id":"PMC_24998444","title":"Rapamycin-binding FKBP25 associates with diverse proteins that form large intracellular entities.","date":"2014","source":"Biochemical and biophysical research communications","url":"https://pubmed.ncbi.nlm.nih.gov/24998444","citation_count":14,"is_preprint":false},{"pmid":"29036638","id":"PMC_29036638","title":"The basic tilted helix bundle domain of the prolyl isomerase FKBP25 is a novel double-stranded RNA binding module.","date":"2017","source":"Nucleic acids research","url":"https://pubmed.ncbi.nlm.nih.gov/29036638","citation_count":14,"is_preprint":false},{"pmid":"24667607","id":"PMC_24667607","title":"Basic Tilted Helix Bundle - a new protein fold in human FKBP25/FKBP3 and HectD1.","date":"2014","source":"Biochemical and biophysical research communications","url":"https://pubmed.ncbi.nlm.nih.gov/24667607","citation_count":13,"is_preprint":false},{"pmid":"10900128","id":"PMC_10900128","title":"Mammalian FKBP-25 and its associated proteins.","date":"2000","source":"Archives of biochemistry and biophysics","url":"https://pubmed.ncbi.nlm.nih.gov/10900128","citation_count":13,"is_preprint":false},{"pmid":"23697935","id":"PMC_23697935","title":"Resolving the functions of peptidylprolyl isomerases: insights from the mutagenesis of the nuclear FKBP25 enzyme.","date":"2013","source":"Biochemical Society transactions","url":"https://pubmed.ncbi.nlm.nih.gov/23697935","citation_count":12,"is_preprint":false},{"pmid":"8876379","id":"PMC_8876379","title":"Cloning and high expression of rabbit FKBP25 in cornea.","date":"1996","source":"Japanese journal of ophthalmology","url":"https://pubmed.ncbi.nlm.nih.gov/8876379","citation_count":10,"is_preprint":false},{"pmid":"34875997","id":"PMC_34875997","title":"Esterase D stabilizes FKBP25 to suppress mTORC1.","date":"2021","source":"Cellular & molecular biology letters","url":"https://pubmed.ncbi.nlm.nih.gov/34875997","citation_count":9,"is_preprint":false},{"pmid":"26239116","id":"PMC_26239116","title":"FKBP25 and FKBP38 regulate non-capacitative calcium entry through TRPC6.","date":"2015","source":"Biochimica et biophysica acta","url":"https://pubmed.ncbi.nlm.nih.gov/26239116","citation_count":9,"is_preprint":false},{"pmid":"30620620","id":"PMC_30620620","title":"FKBP25 participates in DNA double-strand break repair.","date":"2019","source":"Biochemistry and cell biology = Biochimie et biologie cellulaire","url":"https://pubmed.ncbi.nlm.nih.gov/30620620","citation_count":8,"is_preprint":false},{"pmid":"34281390","id":"PMC_34281390","title":"FKBP3 Induces Human Immunodeficiency Virus Type 1 Latency by Recruiting Histone Deacetylase 1/2 to the Viral Long Terminal Repeat.","date":"2021","source":"mBio","url":"https://pubmed.ncbi.nlm.nih.gov/34281390","citation_count":7,"is_preprint":false},{"pmid":"15908283","id":"PMC_15908283","title":"Molecular cloning and expression pattern of the Fkbp25 gene during cerebral cortical neurogenesis.","date":"2005","source":"Gene expression patterns : GEP","url":"https://pubmed.ncbi.nlm.nih.gov/15908283","citation_count":6,"is_preprint":false},{"pmid":"37082996","id":"PMC_37082996","title":"Knockdown of FKBP3 suppresses nasopharyngeal carcinoma cell growth, invasion and migration, deactivated NF-κB/IL-6 signaling pathway through inhibiting histone deacetylase 2 expression.","date":"2023","source":"The Chinese journal of physiology","url":"https://pubmed.ncbi.nlm.nih.gov/37082996","citation_count":6,"is_preprint":false},{"pmid":"29969783","id":"PMC_29969783","title":"Elucidation of the FKBP25-60S Ribosomal Protein L7a Stress Response Signaling During Ischemic Injury.","date":"2018","source":"Cellular physiology and biochemistry : international journal of experimental cellular physiology, biochemistry, and pharmacology","url":"https://pubmed.ncbi.nlm.nih.gov/29969783","citation_count":6,"is_preprint":false},{"pmid":"37987202","id":"PMC_37987202","title":"FKBP3 aggravates the malignant phenotype of diffuse large B-cell lymphoma by PARK7-mediated activation of Wnt/β-catenin signalling.","date":"2023","source":"Journal of cellular and molecular medicine","url":"https://pubmed.ncbi.nlm.nih.gov/37987202","citation_count":5,"is_preprint":false},{"pmid":"12392168","id":"PMC_12392168","title":"Cloning and identification of a novel cDNA which may be associated with FKBP25.","date":"2002","source":"Biochemical genetics","url":"https://pubmed.ncbi.nlm.nih.gov/12392168","citation_count":5,"is_preprint":false},{"pmid":"33553183","id":"PMC_33553183","title":"FKBP25 Regulates Meiotic Apparatus During Mouse Oocyte Maturation.","date":"2021","source":"Frontiers in cell and developmental biology","url":"https://pubmed.ncbi.nlm.nih.gov/33553183","citation_count":4,"is_preprint":false},{"pmid":"37345229","id":"PMC_37345229","title":"FKBP25 regulates myoblast viability and migration and is differentially expressed in in vivo models of muscle adaptation.","date":"2023","source":"The FEBS journal","url":"https://pubmed.ncbi.nlm.nih.gov/37345229","citation_count":1,"is_preprint":false},{"pmid":"37488794","id":"PMC_37488794","title":"[Procine recombinant NK-lysin inhibits hepatocellular carcinoma metastasis by downregulating FKBP3 and inhibiting oxidative phosphorylation and glycolysis: a proteomic analysis].","date":"2023","source":"Nan fang yi ke da xue xue bao = Journal of Southern Medical University","url":"https://pubmed.ncbi.nlm.nih.gov/37488794","citation_count":0,"is_preprint":false}],"cost":{"stage1":{"model":"claude-sonnet-4-6","input_tokens":15510,"output_tokens":5688,"usd":0.065925,"stage1_stop_reason":"end_turn"},"stage2":{"model":"claude-opus-4-8","input_tokens":13954,"output_tokens":4531,"usd":0.091523,"stage2_stop_reason":"end_turn"},"total_usd":0.157448,"stage1_batch_id":"msgbatch_01AsR7g8dGTEiEw9U83bJeWn","stage2_batch_id":"msgbatch_01YV5ZyxtFNUZEjxRcpd9jzb","note":"batch pricing = 50% of standard"},"stage1_raw":"```json\n{\n  \"discoveries\": [\n    {\n      \"year\": 1992,\n      \"finding\": \"FKBP25 (FKBP3) has peptidylprolyl cis-trans isomerase (PPIase) activity comparable to FKBP12, binds rapamycin with higher affinity than FK506 (IC50 ~50 nM vs ~400 nM), and its PPIase activity is inhibited by rapamycin and FK506.\",\n      \"method\": \"In vitro PPIase assay with recombinant FKBP25; inhibition assays with rapamycin and FK506\",\n      \"journal\": \"The Journal of biological chemistry\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 / Strong — direct in vitro enzymatic assay with recombinant protein, replicated in subsequent studies\",\n      \"pmids\": [\"1375932\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 1992,\n      \"finding\": \"FKBP25 contains putative nuclear localization sequences in its sequence, distinguishing it from FKBP12 and FKBP13.\",\n      \"method\": \"cDNA cloning and sequence analysis\",\n      \"journal\": \"The Journal of biological chemistry\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 3 / Moderate — sequence-based prediction confirmed by later fractionation experiments in other papers\",\n      \"pmids\": [\"1375932\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 1993,\n      \"finding\": \"FKBP25 is predominantly localized to the nuclear fraction of T-lymphoma Jurkat cells and has the ability to bind DNA; the FKBP25/DNA complex is dissociable by high salt. FKBP12, which shares C-terminal homology with FKBP25, does not bind DNA.\",\n      \"method\": \"Cell fractionation, Western blotting with anti-FKBP25 antibodies, DNA-binding assay\",\n      \"journal\": \"FEBS letters\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — direct fractionation with functional DNA-binding assay, single lab, negative control with FKBP12\",\n      \"pmids\": [\"8422914\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 1999,\n      \"finding\": \"FKBP25 (FKBP3) expression is down-regulated following p53 induction by DNA-damaging stimuli in human and murine cell lines, identifying it as a transcriptional repression target of p53.\",\n      \"method\": \"Differential gene expression screening following p53 induction; confirmed in multiple cell lines with DNA-damaging stimuli\",\n      \"journal\": \"Oncogene\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — multiple cell lines and stimuli tested, but no direct mechanistic dissection of repression mechanism\",\n      \"pmids\": [\"10557083\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2000,\n      \"finding\": \"Endogenous FKBP25 from porcine brain co-isolates with high-mobility group II protein (HMG-II), and a residual pool associates with Rab5, guanylyl kinase, and phosphatidylethanolamine-binding protein.\",\n      \"method\": \"Immunoblotting, 2D-PAGE, Edman degradation, MALDI-TOF of anti-FKBP25 immunoprecipitates from porcine brain\",\n      \"journal\": \"Archives of biochemistry and biophysics\",\n      \"confidence\": \"Low\",\n      \"confidence_rationale\": \"Tier 3 / Weak — single lab, co-migration/co-purification without reciprocal validation\",\n      \"pmids\": [\"10900128\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2009,\n      \"finding\": \"FKBP25 interacts with MDM2, stimulates MDM2 auto-ubiquitylation and proteasomal degradation, leading to induction of p53 and p21. Depletion of FKBP25 by siRNA increases MDM2 levels and reduces p53 and p21.\",\n      \"method\": \"Co-immunoprecipitation, siRNA knockdown, western blotting for MDM2/p53/p21 levels\",\n      \"journal\": \"FEBS letters\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — Co-IP plus functional siRNA knockdown with multiple readouts, single lab\",\n      \"pmids\": [\"19166840\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2013,\n      \"finding\": \"Mutagenesis of FKBP25 revealed that certain mutations destabilize the FKBD domain fold entirely, while other 'surgical' mutations specifically ablate PPIase catalytic activity while maintaining domain structure, enabling distinction between catalytic and non-catalytic functions.\",\n      \"method\": \"Site-directed mutagenesis, in vitro PPIase assays, structural characterization of mutants\",\n      \"journal\": \"Biochemical Society transactions\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 1 / Moderate — mutagenesis with in vitro enzymatic validation, single lab\",\n      \"pmids\": [\"23697935\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2014,\n      \"finding\": \"FKBP25 interacts with nucleolin in an rRNA-dependent manner and associates with the immature pre-60S ribosomal subunit in nuclear extract but not with mature ribosomes, implicating FKBP25 in ribosome biogenesis.\",\n      \"method\": \"Proteomic characterization (mass spectrometry of immunoprecipitates), Co-IP with RNase treatment to assess rRNA-dependence\",\n      \"journal\": \"RNA (New York, N.Y.)\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — Co-IP with MS and orthogonal RNase sensitivity test, single lab\",\n      \"pmids\": [\"24840943\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2014,\n      \"finding\": \"The N-terminal domain of FKBP25 (residues 1–73) adopts a novel 5-helix bundle structure termed the Basic Tilted Helix Bundle (BTHB) with a positively charged surface patch, which is implicated in DNA binding.\",\n      \"method\": \"NMR/structural determination of FKBP25(1-73), comparative structural analysis with HectD1\",\n      \"journal\": \"Biochemical and biophysical research communications\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 1 / Moderate — structure determined by NMR, functional inference from conserved surface; single lab\",\n      \"pmids\": [\"24667607\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2014,\n      \"finding\": \"Endogenous FKBP25 associates with core histones, spliceosomal complexes, ribosomal subunits, and polyribosomes; FKBP25 from polyribosomes can be released by added RNA or high salt, indicating RNA-mediated interaction. Rapamycin or FK506 only partially release FKBP25, suggesting additional protein-mediated binding via the PPIase cavity.\",\n      \"method\": \"Anti-FKBP25 immunoprecipitation from HeLa, K568, and porcine brain cells; polyribosome fractionation; salt/RNA competition; proteomics\",\n      \"journal\": \"Biochemical and biophysical research communications\",\n      \"confidence\": \"Low\",\n      \"confidence_rationale\": \"Tier 3 / Weak — single lab, fractionation and pulldown without full mechanistic dissection\",\n      \"pmids\": [\"24998444\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2015,\n      \"finding\": \"FKBP25 associates with TRPC6 (and TRPC3), and its interaction pattern with TRPCs is modified by FK506. Knockdown of FKBP25 significantly inhibits OAG-evoked non-capacitative calcium entry (NCCE) in MEG-01 and HEK293 cells. Biotinylation experiments indicate FKBP25 localizes to the plasma membrane in addition to intracellular compartments.\",\n      \"method\": \"Co-immunoprecipitation, siRNA knockdown, calcium entry assays, surface biotinylation in platelets and cell lines\",\n      \"journal\": \"Biochimica et biophysica acta\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — Co-IP plus functional siRNA knockdown with calcium flux readouts, single lab\",\n      \"pmids\": [\"26239116\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2016,\n      \"finding\": \"FKBP25 forms a rapamycin-induced ternary complex with the FRB domain of mTOR, and the crystal structure of FRB-rapamycin-FKBP25 was determined at 1.67-Å resolution, revealing conformational changes in FRB and covalent metalloid coordination at C2085 of FRB.\",\n      \"method\": \"Proximity biotin labeling (pBirA), immunoprecipitation, immunofluorescence, X-ray crystallography\",\n      \"journal\": \"ACS central science\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 / Strong — crystal structure at 1.67 Å plus orthogonal biochemical confirmation; multiple methods in one study\",\n      \"pmids\": [\"27610411\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2016,\n      \"finding\": \"NMR solution structure of full-length human FKBP25 shows the N-terminal HLH domain and C-terminal FKBD interact with each other; both domains participate in DNA binding, with the HLH domain making major-groove contacts and a basic FKBD loop contributing to minor-groove interactions.\",\n      \"method\": \"NMR structure determination, NMR-monitored DNA titration, mutagenesis, FKBP25-DNA complex modeling\",\n      \"journal\": \"Nucleic acids research\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 / Strong — full-length NMR structure with mutagenesis and functional DNA binding validation in one study\",\n      \"pmids\": [\"26762975\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2017,\n      \"finding\": \"The N-terminal BTHB domain of FKBP25 selectively binds double-stranded RNA (dsRNA) over DNA or single-stranded oligonucleotides. This RNA-binding activity is required for FKBP25's nucleolar localization and for the vast majority of its protein interactions, including those with pre-60S ribosomes and early ribosome biogenesis factors.\",\n      \"method\": \"Biochemical dsRNA-binding assays, mutagenesis, immunofluorescence for localization, Co-IP for protein interactions in BTHB mutants\",\n      \"journal\": \"Nucleic acids research\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 / Strong — multiple orthogonal methods (binding assay, mutagenesis, localization, Co-IP) in one study\",\n      \"pmids\": [\"29036638\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2017,\n      \"finding\": \"FKBP3 promotes NSCLC cell proliferation via a pathway in which FKBP3 inhibits ubiquitination of Sp1, stabilizing Sp1, which drives HDAC2 promoter activity, increasing HDAC2 expression, leading to HDAC2-mediated deacetylation of histone H3K4 at the p27 promoter and reduced p27 expression.\",\n      \"method\": \"siRNA knockdown, overexpression, ChIP (HDAC2 at p27 promoter), ubiquitination assay for Sp1, western blotting, in vitro and in vivo proliferation assays\",\n      \"journal\": \"Theranostics\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — multiple mechanistic steps validated with orthogonal methods in one lab\",\n      \"pmids\": [\"28839465\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2018,\n      \"finding\": \"The FKBP domain of FKBP25 directly binds microtubules in vitro to promote their polymerization and stabilize the MT network. FKBP25 associates with the mitotic spindle, regulates entry into mitosis, and its knockdown leads to increased chromosome instability. Additionally, FKBP25 association with chromatin is regulated by Protein Kinase C (PKC) phosphorylation in a cell-cycle-dependent manner, disrupting FKBP25-DNA contacts during mitosis while maintaining spindle association.\",\n      \"method\": \"Direct microtubule-binding assay (in vitro), mitotic spindle immunofluorescence, chromosome instability assays in FKBP25 knockdown cells, PKC phosphorylation assays\",\n      \"journal\": \"Nucleic acids research\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 / Strong — in vitro reconstitution of microtubule binding, multiple cellular assays, and phosphorylation mechanism in one study\",\n      \"pmids\": [\"29361176\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2018,\n      \"finding\": \"FKBP25 translocates from cytoplasm to nucleus in endothelial cells following oxygen-glucose deprivation (OGD) or peroxynitrite treatment, and in the nucleus interacts with 60S ribosomal protein L7a. Overexpression of FKBP25 protects endothelial cells against OGD injury.\",\n      \"method\": \"Western blot and immunofluorescence (localization), co-immunoprecipitation and FRET (interaction with L7a), overexpression survival assay\",\n      \"journal\": \"Cellular physiology and biochemistry\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — Co-IP plus FRET plus localization in stress conditions, single lab\",\n      \"pmids\": [\"29969783\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2019,\n      \"finding\": \"FKBP25's catalytic (PPIase) activity is required for promoting homologous recombination (HR) repair of DNA double-strand breaks (DSBs). FKBP25-depleted cells form fewer Rad51 repair foci and become dependent on Rad52-mediated single-strand annealing (SSA). Rapamycin treatment also impairs HR, at least partly independently of mTOR.\",\n      \"method\": \"FKBP25 knockdown, catalytic mutant expression, Rad51/Rad52 focus assays (immunofluorescence), genetic epistasis (Rad52 dependence), DSB reporter assays\",\n      \"journal\": \"Biochemistry and cell biology\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 / Strong — catalytic mutant plus multiple orthogonal cellular DNA repair assays, epistasis with Rad52\",\n      \"pmids\": [\"30620620\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2019,\n      \"finding\": \"FKBP3 downregulation in colorectal cancer cells reduces HDAC2 expression and P-gp levels, reduces p-AKT, and increases PTEN and cleaved caspase-3, sensitizing cells to oxaliplatin. Upregulation of HDAC2 counteracts FKBP3 knockdown-induced sensitization.\",\n      \"method\": \"siRNA knockdown and overexpression, western blotting, flow cytometry (apoptosis), rescue by HDAC2 overexpression\",\n      \"journal\": \"Oncology reports\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — functional rescue experiment confirms HDAC2 as downstream effector; single lab\",\n      \"pmids\": [\"31524278\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2021,\n      \"finding\": \"FKBP3 indirectly binds the HIV-1 LTR through physical interaction with the transcription factor YY1, thereby recruiting HDAC1/2 to the LTR, promoting histone deacetylation and HIV-1 latency. FKBP3 knockout in latently infected cells activates latent HIV-1.\",\n      \"method\": \"CRISPR knockout in latent HIV-1 cell lines, Co-IP (FKBP3-YY1 interaction), ChIP (HDAC1/2 at LTR), primary latent cell model\",\n      \"journal\": \"mBio\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — CRISPR KO with ChIP and Co-IP, multiple latent cell line models, single lab\",\n      \"pmids\": [\"34281390\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2021,\n      \"finding\": \"Esterase D (ESD) interacts with FKBP25 via the N-terminal 1–90 aa domain of FKBP25. ESD reduces K48-linked poly-ubiquitin chains on FKBP25, thereby stabilizing cytoplasmic FKBP25. Stabilized FKBP25 then binds more mTORC1, suppressing mTORC1 activity and promoting autophagy.\",\n      \"method\": \"Yeast two-hybrid, Co-IP (endogenous and GFP-tagged FKBP25), ubiquitination assay, western blot for mTORC1/P70S6K/4EBP1 phosphorylation\",\n      \"journal\": \"Cellular & molecular biology letters\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — yeast two-hybrid confirmed by Co-IP, ubiquitination assay, and functional mTORC1 readout; single lab\",\n      \"pmids\": [\"34875997\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2021,\n      \"finding\": \"FKBP25 depletion in mouse oocytes causes abnormal spindle assembly, chromosome misalignment, and defective kinetochore-microtubule attachment, leading to elevated aneuploidy. Phosphorylation at serine 163 is identified as a key regulatory modification for FKBP25's function in meiotic maturation.\",\n      \"method\": \"siRNA knockdown in mouse oocytes, immunofluorescence (spindle/chromosome), aneuploidy scoring, site-directed mutagenesis (S163A)\",\n      \"journal\": \"Frontiers in cell and developmental biology\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — knockdown with multiple cellular readouts plus mutagenesis, single lab\",\n      \"pmids\": [\"33553183\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2023,\n      \"finding\": \"FKBP3 interacts with PARK7 (DJ-1); knockdown of FKBP3 enhances ubiquitination and degradation of PARK7, reducing Wnt/β-catenin pathway activation in DLBCL cells.\",\n      \"method\": \"Co-immunoprecipitation (FKBP3-PARK7 interaction), ubiquitination assay, western blotting for β-catenin pathway components, FKBP3 knockdown\",\n      \"journal\": \"Journal of cellular and molecular medicine\",\n      \"confidence\": \"Low\",\n      \"confidence_rationale\": \"Tier 3 / Weak — single Co-IP without reciprocal validation, single lab, limited mechanistic follow-up\",\n      \"pmids\": [\"37987202\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2023,\n      \"finding\": \"FKBP25 knockdown in C2C12 myoblasts increases cell accumulation/viability and migration in vitro, independently of changes in tubulin dynamics. FKBP25 protein expression increases in hypertrophy (chronic mechanical overload) and muscle regeneration (mdx model) but decreases in denervation atrophy in vivo.\",\n      \"method\": \"Doxycycline-inducible knockdown in C2C12 cells, viability and migration assays, tubulin dynamics measurements, in vivo muscle adaptation models with western blotting\",\n      \"journal\": \"The FEBS journal\",\n      \"confidence\": \"Low\",\n      \"confidence_rationale\": \"Tier 3 / Weak — single lab, knockdown with phenotypic readout but limited pathway placement\",\n      \"pmids\": [\"37345229\"],\n      \"is_preprint\": false\n    }\n  ],\n  \"current_model\": \"FKBP25 (FKBP3) is a nuclear prolyl isomerase with a C-terminal FK506-binding/PPIase domain (which binds rapamycin with high affinity, promotes microtubule polymerization, and is required for homologous recombination repair of DNA double-strand breaks) and an N-terminal BTHB domain (which selectively binds dsRNA to drive nucleolar localization and ribosome biogenesis interactions, and also contacts DNA major-groove); its chromatin association is cell-cycle regulated by PKC-mediated phosphorylation, it controls cell proliferation through an FKBP3→Sp1 ubiquitination→HDAC2→p27 axis, it promotes MDM2 degradation to activate p53, it binds YY1 to recruit HDAC1/2 to gene promoters, and its stability is regulated by ESD-mediated suppression of K48-linked ubiquitination which in turn modulates mTORC1 activity.\"\n}\n```","stage2_raw":"{\n  \"mechanistic_narrative\": \"FKBP3 (FKBP25) is a predominantly nuclear peptidylprolyl cis-trans isomerase that couples prolyl isomerase activity to nucleic-acid binding to act in ribosome biogenesis, chromatin regulation, genome maintenance, and the mitotic apparatus [#0, #12]. It is a two-domain protein: a C-terminal FK506-binding/PPIase domain that hydrolyzes peptidyl-prolyl bonds, binds rapamycin with higher affinity than FK506, and is inhibited by both drugs [#0]; and an N-terminal Basic Tilted Helix Bundle (BTHB/HLH) domain that, together with a basic FKBD loop, makes major- and minor-groove contacts on DNA [#8, #12] and selectively binds double-stranded RNA — an activity that drives its nucleolar localization and the bulk of its protein interactions, including with nucleolin and the immature pre-60S ribosomal subunit [#7, #13]. The catalytic PPIase activity is required for homologous-recombination repair of DNA double-strand breaks, with FKBP3-depleted cells forming fewer Rad51 foci and shifting toward Rad52-dependent single-strand annealing [#17]. The FKBD domain directly binds and stabilizes microtubules, and FKBP3 associates with the mitotic spindle and safeguards chromosome stability, with its chromatin association switched off during mitosis by PKC-mediated phosphorylation [#15]. FKBP3 also operates as a chromatin regulator and proliferation driver: it stabilizes Sp1 to upregulate HDAC2 and repress p27 [#14], and binds the transcription factor YY1 to recruit HDAC1/2 to target promoters, a mechanism it uses to enforce HIV-1 latency at the viral LTR [#19]. It additionally tunes the p53 axis by stimulating MDM2 auto-ubiquitylation and degradation [#5] and, via ESD-mediated suppression of K48-linked ubiquitination that stabilizes the cytoplasmic pool, suppresses mTORC1 activity to promote autophagy [#20]. FKBP3 forms a rapamycin-induced ternary complex with the FRB domain of mTOR, defined at atomic resolution [#11].\",\n  \"teleology\": [\n    {\n      \"year\": 1992,\n      \"claim\": \"Established that FKBP3 is an enzymatically active prolyl isomerase distinct from FKBP12, defining its core catalytic identity and drug sensitivity.\",\n      \"evidence\": \"In vitro PPIase assays with recombinant protein and rapamycin/FK506 inhibition; cDNA cloning revealing nuclear localization sequences\",\n      \"pmids\": [\"1375932\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Physiological substrates of the PPIase activity not identified\", \"Nuclear targeting predicted from sequence, not functionally tested at this stage\"]\n    },\n    {\n      \"year\": 1993,\n      \"claim\": \"Showed that FKBP3 is nuclear and binds DNA, a property absent in homologous FKBP12, pointing to a chromatin-associated function beyond isomerase activity.\",\n      \"evidence\": \"Cell fractionation and DNA-binding assays in Jurkat cells with FKBP12 negative control\",\n      \"pmids\": [\"8422914\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"DNA sequence specificity not defined\", \"Domain responsible for DNA binding not mapped\"]\n    },\n    {\n      \"year\": 1999,\n      \"claim\": \"Placed FKBP3 downstream of the p53 stress response by identifying it as a transcriptionally repressed p53 target during DNA damage.\",\n      \"evidence\": \"Differential expression screening after p53 induction across multiple cell lines and damaging stimuli\",\n      \"pmids\": [\"10557083\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Mechanism of p53-mediated repression not dissected\", \"Functional consequence of downregulation not established here\"]\n    },\n    {\n      \"year\": 2009,\n      \"claim\": \"Defined a feedback link to p53 by showing FKBP3 stimulates MDM2 auto-ubiquitylation, thereby raising p53 and p21.\",\n      \"evidence\": \"Co-IP plus siRNA knockdown with MDM2/p53/p21 western readouts\",\n      \"pmids\": [\"19166840\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Whether PPIase activity is required not tested\", \"Single lab, no reciprocal/structural validation of FKBP3-MDM2 contact\"]\n    },\n    {\n      \"year\": 2014,\n      \"claim\": \"Connected FKBP3 to ribosome biogenesis and solved the N-terminal domain fold, establishing a structural basis for its nucleic-acid engagement.\",\n      \"evidence\": \"MS of immunoprecipitates with RNase-sensitivity tests showing nucleolin/pre-60S association; NMR structure of FKBP25(1-73) defining the BTHB fold\",\n      \"pmids\": [\"24840943\", \"24667607\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Direct catalytic role in ribosome assembly not shown\", \"BTHB nucleic-acid selectivity not yet resolved at this stage\"]\n    },\n    {\n      \"year\": 2016,\n      \"claim\": \"Provided full-length and complex structures showing inter-domain communication, dual-groove DNA binding, and the rapamycin-bridged FKBP3-mTOR(FRB) ternary complex.\",\n      \"evidence\": \"NMR of full-length FKBP25 with DNA titration and mutagenesis; X-ray crystallography of FRB-rapamycin-FKBP25 at 1.67 Å\",\n      \"pmids\": [\"26762975\", \"27610411\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"In vivo significance of the mTOR ternary complex not established\", \"Whether DNA binding and PPIase act simultaneously unclear\"]\n    },\n    {\n      \"year\": 2017,\n      \"claim\": \"Resolved the molecular logic of FKBP3 localization by showing the BTHB domain selectively binds dsRNA, which drives nucleolar targeting and most of its interactome.\",\n      \"evidence\": \"dsRNA-binding assays, mutagenesis, immunofluorescence, and Co-IP in BTHB mutants\",\n      \"pmids\": [\"29036638\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Specific dsRNA targets in cells not identified\", \"Relationship between dsRNA and DNA binding modes not reconciled\"]\n    },\n    {\n      \"year\": 2017,\n      \"claim\": \"Defined a proliferation-driving chromatin axis whereby FKBP3 stabilizes Sp1 to upregulate HDAC2 and repress p27.\",\n      \"evidence\": \"siRNA/overexpression, Sp1 ubiquitination assay, HDAC2-p27 promoter ChIP, and proliferation assays in NSCLC models\",\n      \"pmids\": [\"28839465\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Whether PPIase activity drives Sp1 stabilization not tested\", \"Direct enzyme-substrate relationship with Sp1 not shown\"]\n    },\n    {\n      \"year\": 2018,\n      \"claim\": \"Established a structural role at the mitotic apparatus, showing the FKBD domain directly stabilizes microtubules and that cell-cycle PKC phosphorylation toggles chromatin versus spindle engagement.\",\n      \"evidence\": \"In vitro microtubule-binding assay, mitotic spindle immunofluorescence, chromosome instability assays, and PKC phosphorylation assays\",\n      \"pmids\": [\"29361176\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"PKC site(s) and kinase specificity in vivo not fully mapped\", \"How spindle and DNA-repair roles are temporally coordinated unclear\"]\n    },\n    {\n      \"year\": 2019,\n      \"claim\": \"Demonstrated that the catalytic PPIase activity is required for homologous-recombination repair, assigning an enzymatic function to genome maintenance.\",\n      \"evidence\": \"Catalytic-mutant rescue, Rad51/Rad52 focus assays, Rad52 epistasis, and DSB reporter assays\",\n      \"pmids\": [\"30620620\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"HR substrate isomerized by FKBP3 not identified\", \"Rapamycin's mTOR-independent HR effect not fully delineated\"]\n    },\n    {\n      \"year\": 2021,\n      \"claim\": \"Extended the YY1/HDAC repression mechanism to viral chromatin, showing FKBP3 enforces HIV-1 latency by recruiting HDAC1/2 to the LTR via YY1.\",\n      \"evidence\": \"CRISPR knockout in latent HIV models, FKBP3-YY1 Co-IP, and HDAC1/2 LTR ChIP\",\n      \"pmids\": [\"34281390\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Whether DNA/PPIase activities contribute beyond YY1 bridging unclear\", \"Generality to host gene promoters not defined\"]\n    },\n    {\n      \"year\": 2021,\n      \"claim\": \"Identified an ESD-controlled stability switch that links FKBP3 abundance to mTORC1 suppression and autophagy.\",\n      \"evidence\": \"Yeast two-hybrid, Co-IP, K48-ubiquitination assay, and mTORC1/P70S6K/4EBP1 phospho-readouts\",\n      \"pmids\": [\"34875997\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Identity of the E3 ligase opposed by ESD unknown\", \"Relationship to the rapamycin-induced FRB ternary complex not integrated\"]\n    },\n    {\n      \"year\": 2021,\n      \"claim\": \"Confirmed an in vivo meiotic requirement, with FKBP3 needed for proper spindle assembly and kinetochore-microtubule attachment in oocytes.\",\n      \"evidence\": \"siRNA knockdown in mouse oocytes, spindle/chromosome immunofluorescence, aneuploidy scoring, and S163A mutagenesis\",\n      \"pmids\": [\"33553183\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Mechanism linking S163 phosphorylation to function not resolved\", \"Whether the microtubule-binding FKBD mediates this directly not shown\"]\n    },\n    {\n      \"year\": null,\n      \"claim\": \"The endogenous prolyl-isomerase substrates that underlie FKBP3's roles in HR repair, ribosome biogenesis, and chromatin regulation remain unidentified, leaving the unifying catalytic mechanism open.\",\n      \"evidence\": \"No direct substrate identification across the available corpus\",\n      \"pmids\": [],\n      \"confidence\": \"Low\",\n      \"gaps\": [\"No physiological PPIase substrate validated\", \"How nucleic-acid binding, catalysis, and protein scaffolding are integrated into one mechanism is unresolved\"]\n    }\n  ],\n  \"mechanism_profile\": {\n    \"molecular_activity\": [\n      {\"term_id\": \"GO:0016853\", \"supporting_discovery_ids\": [0, 6, 17]},\n      {\"term_id\": \"GO:0003723\", \"supporting_discovery_ids\": [7, 13]},\n      {\"term_id\": \"GO:0003677\", \"supporting_discovery_ids\": [2, 8, 12]},\n      {\"term_id\": \"GO:0008092\", \"supporting_discovery_ids\": [15]},\n      {\"term_id\": \"GO:0140110\", \"supporting_discovery_ids\": [14, 19]}\n    ],\n    \"localization\": [\n      {\"term_id\": \"GO:0005634\", \"supporting_discovery_ids\": [1, 2]},\n      {\"term_id\": \"GO:0005730\", \"supporting_discovery_ids\": [13]},\n      {\"term_id\": \"GO:0005829\", \"supporting_discovery_ids\": [20]},\n      {\"term_id\": \"GO:0005819\", \"supporting_discovery_ids\": [15, 21]}\n    ],\n    \"pathway\": [\n      {\"term_id\": \"R-HSA-73894\", \"supporting_discovery_ids\": [17]},\n      {\"term_id\": \"R-HSA-8953854\", \"supporting_discovery_ids\": [7, 13]},\n      {\"term_id\": \"R-HSA-1640170\", \"supporting_discovery_ids\": [15, 21]},\n      {\"term_id\": \"R-HSA-4839726\", \"supporting_discovery_ids\": [14, 19]},\n      {\"term_id\": \"R-HSA-162582\", \"supporting_discovery_ids\": [11, 20]}\n    ],\n    \"complexes\": [\n      \"FKBP25-rapamycin-mTOR(FRB) ternary complex\",\n      \"pre-60S ribosomal subunit\"\n    ],\n    \"partners\": [\n      \"MDM2\",\n      \"nucleolin\",\n      \"Sp1\",\n      \"YY1\",\n      \"HDAC1\",\n      \"HDAC2\",\n      \"mTOR\",\n      \"ESD\"\n    ],\n    \"other_free_text\": []\n  }\n}","audit_flag":null,"evaluation":{"pairwise":"win","faith_supported":7,"faith_total":7,"faith_pct":100.0}}