{"gene":"STK3","run_date":"2026-06-10T10:51:54","timeline":{"discoveries":[{"year":1996,"finding":"STK3 (KRS1/MST2) is a Ste20-related serine/threonine kinase that is activated by a subset of stress conditions and apoptosis-inducing agents but not by mitogenic stimuli, establishing it as a stress-responsive kinase.","method":"Protein purification, cloning, and kinase activity assays","journal":"Proceedings of the National Academy of Sciences of the United States of America","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — purification and cloning with kinase activity characterization, single lab, founding study","pmids":["8816758"],"is_preprint":false},{"year":1998,"finding":"Rat MST2 (STK3 ortholog) phosphorylates thyroid transcription factor-1 (TTF-1) at the same residues previously identified as major in vivo phosphorylation sites, identifying TTF-1 as the first substrate of this kinase class.","method":"In-gel kinase assay, cloning, in vitro kinase assay, in vivo phosphorylation mapping","journal":"The Journal of biological chemistry","confidence":"Medium","confidence_rationale":"Tier 1 / Weak — in vitro kinase assay with site identification, single lab, single study","pmids":["9430685"],"is_preprint":false},{"year":2003,"finding":"MST2 (STK3) kinase activity depends on autophosphorylation at Thr180 via an intermolecular (trans) mechanism; caspase-3 cleaves MST2 generating a truncated form that is resistant to dephosphorylation by PP1 and PP2A, in contrast to full-length MST2 which is rapidly dephosphorylated, explaining constitutive activation of the truncated form during apoptosis.","method":"Site-directed mutagenesis, in vitro kinase assays, phosphatase treatment, cell transfection with truncation mutants","journal":"The Journal of biological chemistry","confidence":"High","confidence_rationale":"Tier 1 / Moderate — reconstitution with mutagenesis and phosphatase assays, multiple orthogonal approaches, single lab","pmids":["12554736"],"is_preprint":false},{"year":2004,"finding":"Raf-1 suppresses MST2 (STK3) activation by preventing its dimerization and activation-loop phosphorylation independently of Raf-1 kinase activity; depletion of MST2 from Raf-1-/- cells abrogates apoptosis hypersensitivity, placing MST2 downstream of Raf-1 in an apoptosis-suppressive pathway.","method":"Proteomic analysis of Raf-1 signaling complexes, Co-IP, siRNA knockdown, genetic epistasis (Raf-1-/- cells), overexpression","journal":"Science","confidence":"High","confidence_rationale":"Tier 2 / Strong — reciprocal Co-IP, genetic epistasis with defined phenotype, replicated in mouse and human cells","pmids":["15618521"],"is_preprint":false},{"year":2005,"finding":"MST2 (STK3) phosphorylates LATS1 at its activation loop (S909) and hydrophobic motif (T1079), activating LATS1 kinase activity; MST2 also directly interacts with hWW45 (Salvador ortholog) via their conserved domains.","method":"In vitro kinase assay, mass spectrometry phosphosite mapping, deletion analysis, Co-IP","journal":"Oncogene","confidence":"High","confidence_rationale":"Tier 1 / Moderate — in vitro reconstitution with phosphosite identification by MS, multiple orthogonal methods, single lab","pmids":["15688006"],"is_preprint":false},{"year":2006,"finding":"MST1 and MST2 (STK3) heterodimerize with hSav (hWW45/Salvador) via conserved C-terminal coiled-coil (SARAH) domains; this interaction leads to hSav phosphorylation by Mst kinases and stabilization of hSav protein, with stabilization requiring the Mst-hSav interaction but not necessarily phosphorylation.","method":"Co-IP, deletion mutagenesis, in vitro phosphorylation, co-expression abundance assays","journal":"The FEBS journal","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — reciprocal Co-IP, in vitro phosphorylation, mutagenesis, single lab","pmids":["16930133"],"is_preprint":false},{"year":2007,"finding":"RASSF1A disrupts the inhibitory Raf-1–MST2 complex, enhancing MST2 interaction with LATS1; RASSF1A-activated LATS1 phosphorylates YAP1, releasing it to translocate to the nucleus and associate with p73, driving transcription of the proapoptotic gene PUMA.","method":"Co-IP, siRNA knockdown, epistasis analysis, reporter assays, apoptosis assays","journal":"Molecular cell","confidence":"High","confidence_rationale":"Tier 2 / Strong — reciprocal Co-IP, genetic epistasis, pathway reconstitution with defined molecular readouts, replicated across multiple cell types","pmids":["17889669"],"is_preprint":false},{"year":2008,"finding":"MST1 and MST2 (STK3) phosphorylate MOBKL1A/MOBKL1B (Mats homologs) as preferred substrates in vitro and in cells during mitosis; MST1/2 activity increases during mitosis (especially in nocodazole arrest); MOBKL1A/B phosphorylation promotes their binding to LATS1 and enables LATS1 activation-loop phosphorylation; replacing endogenous MOBKL1A/B with non-phosphorylatable mutants accelerates cell proliferation through G1/S and mitotic exit.","method":"In vitro kinase assay, cell-based phosphorylation (okadaic acid/H2O2 treatment), Co-IP, dominant-negative replacement, cell cycle analysis","journal":"Current biology","confidence":"High","confidence_rationale":"Tier 1 / Strong — in vitro reconstitution, multiple orthogonal methods (in vitro and in vivo phosphorylation, functional rescue), multiple perturbations","pmids":["18328708"],"is_preprint":false},{"year":2008,"finding":"MST2 (STK3) cooperates with LATS1 kinase to phosphorylate YAP at Ser-127, requiring WW domains of YAP and PPxY motifs in LATS kinases; in HEK293 cells this Hippo pathway reconstitution is anti-apoptotic, partially rescuing cells from YAP2-induced apoptotic death; nuclear localization of YAP2 is negatively regulated by LATS1.","method":"Reconstitution in human cells, Co-IP with deletion/point mutants, subcellular fractionation/localization, apoptosis assays","journal":"The Journal of biological chemistry","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — functional reconstitution with domain mapping, single lab, multiple readouts","pmids":["18640976"],"is_preprint":false},{"year":2008,"finding":"MST2 phosphorylates MOB1 at Thr74 in vitro; this phosphorylation is essential for forming the MOB1–MST2–NDR1 complex and for full activation of NDR1; MOB1 T74A mutant fails to enhance its interaction with NDR1 upon okadaic acid stimulation.","method":"In vitro kinase assay, site-directed mutagenesis (T74A, T181A), Co-IP, NDR1 activation assay","journal":"Oncogene","confidence":"High","confidence_rationale":"Tier 1 / Moderate — in vitro phosphorylation with site mutagenesis validated in cells, multiple orthogonal approaches","pmids":["18362890"],"is_preprint":false},{"year":2009,"finding":"MST2 (STK3) promotes MST2-dependent NDR1 activation and precise chromosome alignment at metaphase; depletion of MST2 or Fry or MOB2 causes mitotic chromosome misalignment; expression of active NDR1 rescues MST2-depletion misalignment; Fry acts as a scaffold binding NDR1, MOB2, and microtubules on the spindle.","method":"siRNA knockdown, live-cell imaging, rescue by active NDR1, biochemical interaction (Co-IP/pulldown)","journal":"Current biology","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — loss-of-function with specific mitotic phenotype and epistasis rescue, single lab","pmids":["19327996"],"is_preprint":false},{"year":2009,"finding":"In mouse liver, Mst1 and Mst2 are cleaved and constitutively activated; combined deficiency results in loss of inhibitory Ser127 phosphorylation of Yap1 and hepatocellular carcinoma; Mst1/2 inactivates Yap1 through an intermediary kinase distinct from Lats1/2 in the liver context.","method":"Conditional knockout mice, phospho-specific western blot, re-expression rescue in HCC cell lines","journal":"Cancer cell","confidence":"High","confidence_rationale":"Tier 2 / Strong — in vivo genetic model with defined molecular (phosphorylation) and cellular (tumor suppression) phenotypes, reinforced by cell-line rescue","pmids":["19878874"],"is_preprint":false},{"year":2009,"finding":"RASSF2 directly binds and stabilizes MST2 (STK3) protein at endogenous levels; RASSF2 co-localizes with MST2 in the cytoplasm (shifting MST2 from nuclear-alone); RASSF2 expression increases MST2 protein levels and protects MST2 from proteolytic degradation; RASSF2 itself is phosphorylated by co-immunoprecipitated MST1/2.","method":"Endogenous Co-IP, co-localization imaging, siRNA knockdown, immunoblotting for protein stability","journal":"Oncogene","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — endogenous reciprocal Co-IP with localization and stability assays, single lab","pmids":["19525978"],"is_preprint":false},{"year":2010,"finding":"Akt phosphorylates MST2 at T117 and T384; these phosphorylations inhibit MST2 by (1) blocking its binding to RASSF1A, (2) promoting its association into the Raf-1 inhibitory complex, and (3) preventing MST2 homodimerization required for activation; Akt thus limits MST2 proapoptotic activity downstream of mitogens and oncogenic Ras.","method":"In vitro kinase assay, site-directed mutagenesis (T117A/T384A), Co-IP, epistasis with Ras effector domain mutants","journal":"Cancer research","confidence":"High","confidence_rationale":"Tier 1 / Moderate — in vitro phosphorylation with mutagenesis validated by Co-IP and pathway readout, multiple orthogonal methods","pmids":["20086174"],"is_preprint":false},{"year":2010,"finding":"Akt phosphorylates MST2 at T117, leading to inhibition of MST2 cleavage, nuclear translocation, autophosphorylation at Thr180, and kinase activity; IGF1-PI3K-Akt signaling suppresses DNA damage-induced MST2 activation.","method":"In vitro and in vivo labeling kinase assays, site-directed mutagenesis, immunoblotting","journal":"PloS one","confidence":"Medium","confidence_rationale":"Tier 1 / Weak — in vitro kinase assay with mutagenesis, single lab, partially overlapping findings with Romano 2010","pmids":["20231902"],"is_preprint":false},{"year":2010,"finding":"A-Raf prevents MST2-mediated apoptosis in cancer cells by sequestering and inactivating MST2; this requires hnRNP H-dependent A-Raf transcription; siRNA knockdown of hnRNP H or A-Raf results in MST2-dependent apoptosis.","method":"siRNA knockdown, Co-IP, apoptosis assays","journal":"Cancer research","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — Co-IP with siRNA epistasis and defined apoptotic phenotype, single lab","pmids":["20145135"],"is_preprint":false},{"year":2011,"finding":"The tumor suppressor RASSF1A prevents PP2A-mediated dephosphorylation of MST1/2 at their activation-loop residues (Thr183/Thr180), maintaining MST2 in an activated, autophosphorylated state; RASSF1A also stabilizes MST2 protein.","method":"Dephosphorylation assays with PP2A, PP2A knockdown, RASSF1A depletion, phospho-specific western blot","journal":"The Journal of biological chemistry","confidence":"High","confidence_rationale":"Tier 1 / Moderate — direct dephosphorylation assays combined with PP2A knockdown and RASSF1A depletion, multiple orthogonal approaches, single lab","pmids":["21199877"],"is_preprint":false},{"year":2011,"finding":"Mutant K-Ras directly binds RASSF1A to activate the apoptotic MST2–LATS1 pathway; activated LATS1 sequesters Mdm2 to stabilize p53 and drive apoptosis; wild-type K-Ras inhibits this pathway via AKT activation.","method":"Co-IP, siRNA knockdown, epistasis analysis with Ras effector domain mutants, apoptosis assays","journal":"Molecular cell","confidence":"High","confidence_rationale":"Tier 2 / Strong — reciprocal Co-IP, genetic epistasis across multiple cell lines, pathway mechanism mapped to p53 stabilization","pmids":["22195963"],"is_preprint":false},{"year":2012,"finding":"Mst1 and Mst2 (STK3) control Rho GTPase activation (Rac1 and RhoA GTP charging) in SP thymocytes; when phosphorylated by Mst1/2, Mob1 binds and activates the Rac1 guanyl nucleotide exchanger Dock8; this pathway drives sphingosine-1-phosphate and CCL21-induced cell migration and thymic egress.","method":"Conditional double knockout mice, Co-IP, GTP-loading assays, migration assays, phospho-Mob1 blotting","journal":"The Journal of experimental medicine","confidence":"High","confidence_rationale":"Tier 2 / Strong — in vivo genetic model with defined molecular mechanism (Mob1 phosphorylation → Dock8 activation → Rac1 GTP charging) and cellular phenotype","pmids":["22412158"],"is_preprint":false},{"year":2013,"finding":"Crystal structure of human MST2 kinase domain alone and in complex with RASSF5 (SARAH domain) reveals that MST2 undergoes activation via trans-autophosphorylation at its activation loop, requiring SARAH domain-mediated homodimerization; RASSF5 binding disrupts MST2 homodimer, blocking autoactivation of unphosphorylated MST2 but not inhibiting already-activated MST2, revealing temporal context-dependent dual roles for RASSF5.","method":"X-ray crystallography, biochemical autophosphorylation assays, mutagenesis","journal":"Structure","confidence":"High","confidence_rationale":"Tier 1 / Strong — crystal structure plus functional biochemical validation with mutagenesis, multiple orthogonal approaches","pmids":["23972470"],"is_preprint":false},{"year":2013,"finding":"Chronic (not transient) K-Ras activation drives MST2–LATS1 complex formation and apoptosis; transient EGF-stimulated K-Ras activation prevents MST2–LATS1 complex formation in an AKT-dependent manner, demonstrating that activation kinetics determine pathway output.","method":"Co-IP, siRNA, AKT inhibition, overexpression of Ras exchange factors, apoptosis assays","journal":"Molecular and cellular biology","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — Co-IP with multiple genetic and pharmacological perturbations, single lab","pmids":["23459937"],"is_preprint":false},{"year":2014,"finding":"c-Abl tyrosine kinase phosphorylates MST2 at the conserved site Y81 within the kinase domain; this phosphorylation disrupts MST2–Raf-1 interaction, enhances MST2 homodimerization and kinase activation, and induces neuronal cell death.","method":"In vitro kinase assay, site-directed mutagenesis (Y81F), Co-IP, cell death assay","journal":"PloS one","confidence":"Medium","confidence_rationale":"Tier 1 / Weak — in vitro kinase assay with mutagenesis validated in cells, single lab","pmids":["22590567"],"is_preprint":false},{"year":2014,"finding":"Competing protein interactions coordinate Raf-1–MST2 crosstalk: Akt phosphorylation of MST2 and LATS1-feedback phosphorylation of Raf-1 Ser259 enable Raf-1 to suppress both MST2 and MEK signaling; mutation of Raf-1 Ser259 simultaneously drives apoptosis and proliferation; concomitant MST2 downregulation switches outcome to proliferation and transformation.","method":"Phosphorylation assays, Co-IP, mutagenesis, mathematical modeling validated by experiment, cell transformation assays","journal":"Nature cell biology","confidence":"High","confidence_rationale":"Tier 1 / Strong — mathematical model-guided mutagenesis with multiple orthogonal experimental validations including functional cell fate readouts","pmids":["24929361"],"is_preprint":false},{"year":2014,"finding":"STK3 (MST2) and STK4 (MST1) phosphorylate LC3 at threonine 50 (Thr50); loss of this phosphorylation blocks autophagosome–lysosome fusion and impairs intracellular bacterial clearance; constitutive phosphomimetic LC3-T50E reverses autophagy block in STK3/STK4-deficient cells.","method":"In vitro kinase assay, site-directed mutagenesis, autophagy flux assays, bacterial clearance assay, genetic rescue","journal":"Molecular cell","confidence":"High","confidence_rationale":"Tier 1 / Strong — in vitro reconstitution with mutagenesis, genetic rescue, conserved across species, multiple functional readouts","pmids":["25544559"],"is_preprint":false},{"year":2014,"finding":"Crystal structure of the MST2 SARAH domain shows it forms an antiparallel homodimeric coiled coil; structure-guided mutagenesis identified interface residues critical for homodimerization; these same mutations also impaired heterodimerization with RAPL (RASSF5); SARAH-mediated homodimerization and RAPL heterointeraction are both required for full MST2 activation and apoptotic functions in T cells.","method":"X-ray crystallography, site-directed mutagenesis, Co-IP, kinase activity assays, apoptosis assays","journal":"Journal of structural biology","confidence":"High","confidence_rationale":"Tier 1 / Strong — crystal structure with mutagenesis validated by functional kinase and apoptosis assays","pmids":["24468289"],"is_preprint":false},{"year":2014,"finding":"MST2 (STK3) kinase suppresses rDNA transcription in response to DNA damage by phosphorylating histone H2B at serine 14 (H2BS14p) in an ATM-dependent manner; MST2 localizes to nucleoli; H2BS14p marks transcriptionally inactive nucleolar chromatin and is necessary for rDNA transcriptional shutdown and genomic integrity maintenance.","method":"ChIP, immunofluorescence localization, siRNA knockdown, kinase assays, ATM inhibition","journal":"The EMBO journal","confidence":"High","confidence_rationale":"Tier 2 / Moderate — direct localization experiment tied to functional consequence (rDNA transcription), ChIP with defined histone phosphorylation mark, multiple orthogonal approaches","pmids":["29789391"],"is_preprint":false},{"year":2015,"finding":"Mst1 and Mst2 (STK3) activate the GTPase Rac to promote TLR-triggered assembly of the TRAF6–ECSIT complex, which recruits mitochondria to phagosomes for synergistic ROS production and bactericidal activity.","method":"Conditional knockout mice, Co-IP, Rac GTP-loading assays, mitochondria trafficking (imaging), ROS measurement, bacterial killing assays","journal":"Nature immunology","confidence":"High","confidence_rationale":"Tier 2 / Strong — in vivo genetic model, Co-IP, mechanistic signaling axis defined with multiple orthogonal readouts","pmids":["26414765"],"is_preprint":false},{"year":2016,"finding":"XMU-MP-1 is a selective, reversible MST1/2 inhibitor confirmed by co-crystal structure; it blocks MST1/2 kinase activities, activates downstream YAP, and promotes tissue repair in mouse intestinal and liver injury models.","method":"Enzyme-based HTS, co-crystal structure, structure-activity relationship, in vivo pharmacology","journal":"Science translational medicine","confidence":"High","confidence_rationale":"Tier 1 / Strong — co-crystal structure with SAR and multiple in vivo models confirming on-target activity","pmids":["27535619"],"is_preprint":false},{"year":2016,"finding":"H-Ras promotes formation of inactive Mst1–Mst2 heterodimers via their SARAH domains in an ERK-dependent manner; Mst1/Mst2 heterodimers have much-reduced kinase activity compared to homodimers; cells lacking Mst1 (unable to form heterodimers) are resistant to H-Ras transformation and maintain active Hippo signaling.","method":"Co-IP, kinase activity assays comparing homo- vs heterodimers, ERK pathway inhibition, Mst1 knockout cells, transformation assays","journal":"Current biology","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — Co-IP with kinase activity comparison and genetic validation, single lab","pmids":["27238285"],"is_preprint":false},{"year":2016,"finding":"CDK1 phosphorylates MST2 at serine 385 during mitosis in vitro and in vivo; this phosphorylation does not affect MST2 kinase activity or Hippo-YAP signaling, but phosphorylation-deficient MST2-S385A has higher activity in suppressing proliferation and tumorigenesis, establishing CDK1-mediated mitotic regulation of MST2 tumor suppression.","method":"In vitro CDK1 kinase assay, phospho-site mutagenesis (S385A), cell proliferation and anchorage-independent growth assays, in vivo xenograft","journal":"Cellular signalling","confidence":"Medium","confidence_rationale":"Tier 1 / Weak — in vitro kinase assay with mutagenesis and in vivo tumor assay, single lab","pmids":["27566175"],"is_preprint":false},{"year":2016,"finding":"A-Raf subcellular localization regulates MST2 activity: A-Raf at mitochondria (in proliferating/cancer cells) efficiently sequesters and inhibits MST2, while A-Raf relocated to the plasma membrane (during differentiation) loses its ability to inhibit MST2; this redistribution is regulated by the scaffold KSR2.","method":"Subcellular fractionation, live-cell imaging, siRNA knockdown of KSR2, apoptosis assays","journal":"Cell death and differentiation","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — direct localization experiments tied to functional consequence on MST2 activity and apoptosis, single lab","pmids":["26891695"],"is_preprint":false},{"year":2018,"finding":"Mst1 and Mst2 amplify IL-2–STAT5 signaling in regulatory T cells by promoting STAT5 activation; Mst1 associates with the cytoskeletal DOCK8–LRCH module; Mst1 deficiency limits Treg cell migration and access to IL-2, and impairs Rac GTPase activity mediating STAT5 activation.","method":"Conditional knockout mice, quantitative proteomics (unbiased Co-IP-MS), phospho-STAT5 assays, migration assays, Rac activation assays","journal":"Immunity","confidence":"High","confidence_rationale":"Tier 2 / Strong — unbiased quantitative proteomics identifying DOCK8-LRCH complex, validated by genetic loss-of-function with multiple mechanistic readouts","pmids":["30413360"],"is_preprint":false},{"year":2019,"finding":"MST2 phosphorylates Runx2 at Ser339 and Ser370 (identified by mass spectrometry); MST2 interacts with Runx2 via SAV1's WW domain binding to a PY motif (aa 292–445) of Runx2; phosphorylation inhibits Runx2 transcriptional activity and osteoblast differentiation; phosphorylation-deficient double mutant (S339A/S370A) is resistant to MST2/SAV1-mediated inhibition.","method":"In vitro kinase assay, mass spectrometry phosphosite mapping, Co-IP domain mapping, reporter assay, differentiation assays, mutagenesis","journal":"Biochemical and biophysical research communications","confidence":"Medium","confidence_rationale":"Tier 1 / Moderate — in vitro kinase assay with MS-identified sites, functional mutagenesis, and cellular differentiation readouts, single lab","pmids":["30910359"],"is_preprint":false},{"year":2020,"finding":"Increasing proximity of adjacent MST2 kinase domains—via SARAH domain homodimerization, membrane recruitment, or complex formation with SAV1—is sufficient to trigger MST2 autophosphorylation; a specific protein assembly is not required; multiple upstream activating events converge on this common molecular mechanism.","method":"Chemically induced dimerization, single-molecule pulldown, in vitro biochemistry, cell-based autophosphorylation assays","journal":"The Journal of biological chemistry","confidence":"High","confidence_rationale":"Tier 1 / Strong — multiple orthogonal biochemical and cell-based methods with chemically induced dimerization as controlled perturbation","pmids":["32994222"],"is_preprint":false},{"year":2021,"finding":"STK3 and STK4 (MST2/MST1) suppress mitochondrial capacity and mitophagy in adipocytes; mechanistically, STK3/4 regulate phosphorylation and dimerization status of the mitophagy receptor BNIP3; genetic inactivation of Stk3/4 increases mitochondrial mass/function and stabilizes UCP1 in beige adipose tissue.","method":"Conditional knockout mice, Co-IP, BNIP3 phosphorylation/dimerization assays, mitochondrial function assays","journal":"Nature metabolism","confidence":"High","confidence_rationale":"Tier 2 / Strong — in vivo genetic model with defined molecular mechanism (BNIP3 phosphorylation/dimerization) and multiple metabolic readouts","pmids":["33758424"],"is_preprint":false},{"year":2022,"finding":"MST2 is ubiquitinated at K473 via K63-linkage by the E3 ligase TRIM21; this ubiquitination promotes MST2 homodimerization and enhances its kinase activity, leading to YAP inactivation and inhibition of EMT in colorectal cancer.","method":"Co-IP, site-directed mutagenesis (K473), in vitro ubiquitination assay, kinase activity assay, tumor organoid models","journal":"Cell chemical biology","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — Co-IP with mutagenesis and functional kinase assay, single lab","pmids":["37354905"],"is_preprint":false},{"year":2022,"finding":"MST2 binds SCFβTrCP E3 ubiquitin ligase via a non-canonical degradation motif and is degraded via proteasomal proteolysis; stiffer extracellular matrix and integrin hyperactivation enhance MST2 degradation through integrin-linked kinase (ILK) and actomyosin stress fibers.","method":"Co-IP, siRNA knockdown of βTrCP, site-directed mutagenesis, proteasome inhibition, molecular dynamics simulation","journal":"Biochimica et biophysica acta. General subjects","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — Co-IP with mutagenesis and functional proteolysis assays, computational validation, single lab","pmids":["36044955"],"is_preprint":false},{"year":2023,"finding":"PRMT5 symmetrically di-methylates MST2 (STK3) at R461 and R467 in its SARAH domain, suppressing MST2 autophosphorylation and kinase activity by blocking homodimerization, thereby inactivating Hippo signaling in pancreatic cancer.","method":"In vitro methylation assay, site-directed mutagenesis (R461/R467), dimerization assays, kinase activity assays, Co-IP, in vivo xenograft with PRMT5 inhibitor","journal":"The EMBO journal","confidence":"High","confidence_rationale":"Tier 1 / Moderate — in vitro biochemical reconstitution with mutagenesis, dimerization assay, functional kinase assay, and in vivo validation","pmids":["37905571"],"is_preprint":false},{"year":2023,"finding":"TRIM69 binds MST2 directly, redistributes it to the perinuclear cytoskeleton, promotes MST2 association with PLK1, and stimulates MST2 phosphorylation at S15 (a PLK1 site critical for centrosome disjunction); TRIM69 ablation causes centrosome scattering and chromosome segregation defects.","method":"Co-IP, immunofluorescence localization, phospho-S15 western blot, siRNA knockdown with centrosome and mitosis phenotype readout","journal":"Nucleic acids research","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — Co-IP with localization experiment tied to functional centrosome phenotype, single lab","pmids":["37739411"],"is_preprint":false},{"year":2024,"finding":"In gastric cancer, STK3 directly interacts with GSK-3β and phosphorylates it, promoting GSK-3β degradation and β-catenin nuclear accumulation, thereby activating Wnt/β-catenin signaling; STK3 is itself a transcriptional target of YAP1, creating a feedback loop.","method":"Co-IP, in vitro kinase assay, ChIP-qPCR, knockout mouse models, functional rescue assays, xenograft models","journal":"Molecular cancer","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — Co-IP with in vitro kinase assay and functional genetic validation, single lab","pmids":["40604818"],"is_preprint":false},{"year":2024,"finding":"STK3 (MST2) activation phosphorylates FOXO1 at Ser212, promoting its nuclear translocation and transcriptional activation of TP53INP1 and P21 to inhibit ESCC cell proliferation and migration; cellular ROS activates STK3 autophosphorylation in ESCC cells.","method":"Co-IP, immunofluorescence, ChIP, western blot, siRNA, xenograft models","journal":"Cellular oncology","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — Co-IP with ChIP and IF confirming nuclear translocation, multiple readouts, single lab","pmids":["38436783"],"is_preprint":false},{"year":2016,"finding":"STK3 (MST2) interacts with apoptosis-inducing factor (AIF); AIF interaction enhances STK3 phosphorylation at Thr180, suggesting AIF is a positive regulator of STK3 activity.","method":"Co-IP, in vitro phosphorylation assay","journal":"PloS one","confidence":"Low","confidence_rationale":"Tier 3 / Weak — single Co-IP and phosphorylation assay, no mechanistic follow-up, single lab","pmids":["24992339"],"is_preprint":false}],"current_model":"STK3 (MST2) is a SARAH domain-containing Ste20-related serine/threonine kinase that is activated by trans-autophosphorylation at Thr180 upon kinase domain proximity (driven by SARAH-mediated homodimerization, membrane recruitment, or SAV1 binding); it is positively regulated by RASSF1A (which protects its activation-loop phosphorylation from PP2A), RASSF2, TRIM21 (K63 ubiquitination at K473), c-Abl (Y81 phosphorylation), and AIF, and negatively regulated by Raf-1/A-Raf (blocking dimerization), Akt (T117/T384 phosphorylation promoting Raf-1 binding and blocking homodimerization), CDK1 (S385 mitotic phosphorylation), PRMT5 (R461/R467 methylation blocking homodimerization), and SCFβTrCP-mediated proteasomal degradation; active STK3 phosphorylates LATS1/2 (S909, T1079), MOBKL1A/B (Thr74), YAP (Ser127), LC3 (Thr50), histone H2B (Ser14), Runx2 (Ser339/Ser370), FOXO1 (Ser212), GSK-3β, and TTF-1, thereby suppressing YAP/TAZ oncogenic transcription, promoting autophagosome–lysosome fusion, suppressing rDNA transcription after DNA damage, regulating mitophagy via BNIP3, and controlling T-cell migration via Mob1–Dock8–Rac1 signaling."},"narrative":{"mechanistic_narrative":"STK3 (MST2) is a stress-responsive Ste20-related serine/threonine kinase and the core kinase of the mammalian Hippo tumor-suppressor pathway, activated by apoptotic and stress stimuli rather than mitogens [PMID:8816758]. Its activity depends on trans-autophosphorylation at Thr180 within the activation loop, a reaction triggered whenever adjacent kinase domains are brought into proximity—by SARAH-domain-mediated antiparallel homodimerization, membrane recruitment, or SAV1 (hWW45/Salvador) complex formation—so that no single dedicated assembly is required for activation [PMID:12554736, PMID:23972470, PMID:24468289, PMID:32994222]. Once active, STK3 propagates Hippo signaling by phosphorylating the adaptor MOBKL1A/B (MOB1) at Thr74 and the downstream kinases LATS1 (S909/T1079) and NDR1, which in turn inactivate the transcriptional coactivator YAP via Ser127 phosphorylation, suppressing YAP/TAZ-driven proliferation and promoting apoptosis [PMID:15688006, PMID:18328708, PMID:18640976, PMID:18362890, PMID:19878874]. This module enforces tumor suppression in vivo, as combined Mst1/Mst2 loss in liver causes loss of YAP Ser127 phosphorylation and hepatocellular carcinoma [PMID:19878874]. STK3 activation is gated by an extensive regulatory network: it is held inactive by Raf-1 and A-Raf (which block dimerization and sequester the kinase) [PMID:15618521, PMID:20145135, PMID:26891695], by Akt phosphorylation at T117/T384 that drives Raf-1 binding and blocks homodimerization [PMID:20086174, PMID:20231902], by CDK1 (S385), by PRMT5 methylation of R461/R467 in the SARAH domain, and by SCFβTrCP-mediated degradation [PMID:27566175, PMID:37905571, PMID:36044955]; it is positively regulated by RASSF1A, which protects activation-loop phosphorylation from PP2A and disrupts the inhibitory Raf-1 complex, by RASSF2 stabilization, by c-Abl phosphorylation at Y81, and by TRIM21 K63-ubiquitination at K473 [PMID:21199877, PMID:17889669, PMID:19525978, PMID:22590567, PMID:37354905]. The K-Ras–RASSF1A–MST2–LATS1 axis couples Ras signaling kinetics to an apoptotic, p53-stabilizing output [PMID:22195963, PMID:23459937, PMID:24929361]. Beyond canonical Hippo signaling, STK3 phosphorylates LC3 at Thr50 to enable autophagosome–lysosome fusion and bacterial clearance [PMID:25544559], phosphorylates histone H2B at Ser14 in nucleoli to shut down rDNA transcription after DNA damage [PMID:29789391], regulates Rac/RhoA GTPase signaling through Mob1–Dock8 to drive thymocyte and Treg migration and TLR-triggered antibacterial ROS production [PMID:22412158, PMID:26414765, PMID:30413360], and controls mitophagy via the receptor BNIP3 in adipocytes [PMID:33758424]. The small-molecule inhibitor XMU-MP-1 confirms these activities pharmacologically and activates YAP to promote tissue repair [PMID:27535619].","teleology":[{"year":1996,"claim":"Established STK3 as a kinase activated by stress and apoptotic stimuli but not mitogens, defining its biological niche before any pathway context existed.","evidence":"Protein purification, cloning, and kinase activity assays","pmids":["8816758"],"confidence":"Medium","gaps":["No substrates or upstream regulators identified","Activation mechanism unknown"]},{"year":1998,"claim":"Identified TTF-1 as the first substrate of this kinase class, providing initial evidence that STK3 directly phosphorylates a defined cellular target.","evidence":"In-gel and in vitro kinase assays with in vivo phosphosite mapping in rat ortholog","pmids":["9430685"],"confidence":"Medium","gaps":["Physiological pathway linking STK3 to TTF-1 not established","Single substrate without broader signaling context"]},{"year":2003,"claim":"Defined the core activation mechanism—trans-autophosphorylation at Thr180—and showed caspase cleavage generates a dephosphorylation-resistant constitutively active form, linking activation to apoptosis.","evidence":"Mutagenesis, in vitro kinase and phosphatase assays, truncation mutants in cells","pmids":["12554736"],"confidence":"High","gaps":["What drives the initial dimerization in cells not yet defined","Downstream effectors of cleaved form unknown"]},{"year":2004,"claim":"Placed STK3 downstream of Raf-1 in an apoptosis-suppressive pathway, revealing a kinase-independent inhibitory mechanism acting on dimerization.","evidence":"Proteomics, reciprocal Co-IP, siRNA, Raf-1-/- genetic epistasis","pmids":["15618521"],"confidence":"High","gaps":["How RASSF proteins relieve this inhibition not yet shown","Structural basis of Raf-1 block undefined"]},{"year":2005,"claim":"Connected STK3 to the Hippo core by showing it phosphorylates and activates LATS1 and binds Salvador, establishing the kinase cascade architecture.","evidence":"In vitro kinase assay, MS phosphosite mapping, deletion analysis, Co-IP","pmids":["15688006","16930133"],"confidence":"High","gaps":["Role of YAP as terminal effector not yet linked","In vivo relevance of LATS activation not tested"]},{"year":2008,"claim":"Defined MOB1 (Thr74) phosphorylation as the adaptor step enabling LATS1/NDR1 activation and showed the module restrains cell-cycle progression, completing the kinase relay logic.","evidence":"In vitro and cellular kinase assays, Co-IP, dominant-negative replacement, cell cycle and NDR1 activation assays","pmids":["18328708","18362890","18640976"],"confidence":"High","gaps":["Whether MOB1 phosphorylation is the sole route to LATS activation unclear","Mitotic versus interphase roles not fully separated"]},{"year":2007,"claim":"Showed RASSF1A relieves Raf-1 inhibition to drive a proapoptotic MST2–LATS1–YAP–p73 axis, establishing how upstream signals switch the pathway toward apoptosis.","evidence":"Co-IP, siRNA, epistasis, reporter and apoptosis assays","pmids":["17889669"],"confidence":"High","gaps":["How RASSF1A physically displaces Raf-1 not resolved","Tissue specificity of p73 versus YAP outputs unclear"]},{"year":2009,"claim":"Validated STK3 as a bona fide tumor suppressor in vivo and identified RASSF2 as a stabilizing partner, distinguishing context-dependent (liver) effector usage.","evidence":"Conditional knockout mice, phospho-specific blotting, endogenous Co-IP, localization and stability assays","pmids":["19878874","19525978"],"confidence":"High","gaps":["Identity of the liver intermediary kinase distinct from LATS unknown","Mechanism of RASSF2-mediated stabilization undefined"]},{"year":2010,"claim":"Mapped the inhibitory inputs Akt (T117/T384) and A-Raf, showing how mitogenic and oncogenic signaling restrain STK3 proapoptotic activity.","evidence":"In vitro kinase assays, mutagenesis, Co-IP, Ras effector epistasis, apoptosis assays","pmids":["20086174","20231902","20145135"],"confidence":"High","gaps":["Quantitative balance between activating and inhibitory inputs unclear","Whether these regulators act simultaneously not resolved"]},{"year":2011,"claim":"Established that RASSF1A protects STK3 activation-loop phosphorylation from PP2A and that mutant K-Ras engages this axis to stabilize p53, linking Ras mutation status to apoptotic output.","evidence":"PP2A dephosphorylation assays, knockdowns, Co-IP, Ras effector epistasis, apoptosis assays","pmids":["21199877","22195963"],"confidence":"High","gaps":["How wild-type versus mutant K-Ras diverge mechanistically only partly defined","PP2A targeting specificity not fully mapped"]},{"year":2012,"claim":"Revealed a non-Hippo function: STK3-phosphorylated Mob1 activates the Rac-GEF Dock8 to drive GTPase signaling and lymphocyte migration, broadening STK3 into cytoskeletal regulation.","evidence":"Conditional double-knockout mice, Co-IP, GTP-loading and migration assays","pmids":["22412158"],"confidence":"High","gaps":["How the same Mob1 phosphorylation switches between LATS and Dock8 outputs unclear","Direct STK3-Dock8 contact not defined"]},{"year":2013,"claim":"Provided structural proof that SARAH-mediated homodimerization enables trans-autophosphorylation and that RASSF5 binding has temporally dual roles, resolving the activation switch at atomic resolution.","evidence":"X-ray crystallography of kinase and SARAH-RASSF5 complexes, autophosphorylation assays, mutagenesis; plus Co-IP/AKT epistasis of Ras kinetics","pmids":["23972470","23459937"],"confidence":"High","gaps":["How membrane recruitment substitutes for homodimerization not addressed here","Dynamics of dimer-to-monomer transitions unresolved"]},{"year":2014,"claim":"Expanded STK3 substrate repertoire (LC3 Thr50 for autophagy, H2B Ser14 for rDNA shutdown) and defined SARAH homodimer interface, c-Abl Y81 activation, and Raf-1 crosstalk circuitry, showing STK3 acts well beyond canonical Hippo.","evidence":"Crystallography, in vitro kinase assays, mutagenesis, autophagy/bacterial clearance and ChIP assays, mathematical modeling","pmids":["25544559","24468289","29789391","22590567","24929361"],"confidence":"High","gaps":["Whether autophagy and Hippo functions are mechanistically separable unclear","Coordination among multiple substrate programs undefined"]},{"year":2015,"claim":"Showed STK3 activates Rac to assemble the TRAF6–ECSIT complex and recruit mitochondria to phagosomes, establishing a role in innate antibacterial ROS production.","evidence":"Conditional knockout mice, Co-IP, Rac GTP-loading, mitochondrial imaging, ROS and killing assays","pmids":["26414765"],"confidence":"High","gaps":["Direct STK3 substrate in this axis not defined","Relationship to Mob1-Dock8 module unclear"]},{"year":2016,"claim":"Layered additional regulation—CDK1 mitotic phosphorylation, ERK-dependent inactive Mst1/Mst2 heterodimers, A-Raf subcellular relocalization, and the XMU-MP-1 inhibitor—refining how STK3 activity is tuned spatially and temporally.","evidence":"In vitro kinase assays, mutagenesis, Co-IP, fractionation, co-crystal structure, in vivo pharmacology","pmids":["27566175","27238285","26891695","27535619"],"confidence":"High","gaps":["Functional weight of heterodimerization in vivo unclear","How CDK1 site affects non-Hippo functions untested"]},{"year":2018,"claim":"Extended the cytoskeletal-migration role to Treg cells via a DOCK8–LRCH module amplifying IL-2–STAT5 signaling, reinforcing STK3 control of immune cell positioning.","evidence":"Conditional knockout, unbiased Co-IP-MS, phospho-STAT5, migration and Rac assays","pmids":["30413360"],"confidence":"High","gaps":["Direct kinase substrate in STAT5 amplification not defined","Mst1 versus Mst2 specific contributions unresolved"]},{"year":2020,"claim":"Unified the activation mechanism by demonstrating that kinase-domain proximity alone—via homodimerization, membrane recruitment, or SAV1—is sufficient to trigger autophosphorylation, with no dedicated assembly required.","evidence":"Chemically induced dimerization, single-molecule pulldown, in vitro and cell-based autophosphorylation","pmids":["32994222"],"confidence":"High","gaps":["How distinct upstream events select downstream output not addressed","Endogenous trigger frequency in vivo unknown"]},{"year":2021,"claim":"Established STK3 control of mitophagy via the receptor BNIP3 in adipocytes, defining a metabolic function in mitochondrial quality control.","evidence":"Conditional knockout mice, Co-IP, BNIP3 phosphorylation/dimerization and mitochondrial function assays","pmids":["33758424"],"confidence":"High","gaps":["Whether BNIP3 is a direct STK3 substrate not firmly established","Link to canonical Hippo signaling in adipocytes unclear"]},{"year":2022,"claim":"Identified opposing ubiquitin-system regulators—TRIM21 K63-ubiquitination (K473) activating, SCFβTrCP degradation (ECM-stiffness-driven) inactivating—showing post-translational and mechanical control of STK3 abundance and activity.","evidence":"Co-IP, mutagenesis, in vitro ubiquitination, proteasome inhibition, organoid and MD simulation","pmids":["37354905","36044955"],"confidence":"Medium","gaps":["Reciprocal validation of ubiquitin marks limited to single labs","Interplay between the two ligases not tested"]},{"year":2023,"claim":"Defined PRMT5 arginine methylation (R461/R467) of the SARAH domain as a homodimerization-blocking off-switch, and TRIM69 as a spatial regulator coupling STK3 to PLK1 at centrosomes, broadening regulatory and mitotic functions.","evidence":"In vitro methylation/kinase/dimerization assays, mutagenesis, Co-IP, localization, centrosome phenotype, in vivo xenograft","pmids":["37905571","37739411"],"confidence":"High","gaps":["Whether methylation and phosphorylation compete on the same SARAH surface unclear","TRIM69-S15/PLK1 axis confirmed by single lab only"]},{"year":2024,"claim":"Added context-specific substrates—GSK-3β (activating Wnt/β-catenin in a YAP1 feedback loop) and FOXO1 Ser212 (driving tumor-suppressive transcription)—illustrating STK3 outputs that diverge from canonical YAP suppression.","evidence":"Co-IP, in vitro kinase, ChIP, knockout mice, IF, xenograft assays","pmids":["40604818","38436783"],"confidence":"Medium","gaps":["Apparent context-dependent oncogenic versus suppressive roles unreconciled","Single-lab findings without independent replication"]},{"year":null,"claim":"How divergent upstream activating events and the diverse substrate repertoire are selected to produce specific cellular outputs (apoptosis, autophagy, migration, mitophagy, transcription) within a single cell remains unresolved.","evidence":"","pmids":[],"confidence":"High","gaps":["No unifying model linking activation context to output choice","Quantitative integration of competing regulators not established","Tissue-specific substrate selection mechanisms undefined"]}],"mechanism_profile":{"molecular_activity":[{"term_id":"GO:0140096","term_label":"catalytic activity, acting on a protein","supporting_discovery_ids":[4,7,8,9,23,25,32,39,40]},{"term_id":"GO:0016740","term_label":"transferase activity","supporting_discovery_ids":[2,4,7,19,33]},{"term_id":"GO:0042393","term_label":"histone binding","supporting_discovery_ids":[25]}],"localization":[{"term_id":"GO:0005730","term_label":"nucleolus","supporting_discovery_ids":[25]},{"term_id":"GO:0005829","term_label":"cytosol","supporting_discovery_ids":[12]},{"term_id":"GO:0005815","term_label":"microtubule organizing center","supporting_discovery_ids":[38]}],"pathway":[{"term_id":"R-HSA-162582","term_label":"Signal Transduction","supporting_discovery_ids":[4,6,7,8,11,17]},{"term_id":"R-HSA-5357801","term_label":"Programmed Cell Death","supporting_discovery_ids":[0,3,6,17]},{"term_id":"R-HSA-9612973","term_label":"Autophagy","supporting_discovery_ids":[23,34]},{"term_id":"R-HSA-1640170","term_label":"Cell Cycle","supporting_discovery_ids":[7,10,29,38]},{"term_id":"R-HSA-168256","term_label":"Immune System","supporting_discovery_ids":[18,26,31]},{"term_id":"R-HSA-74160","term_label":"Gene expression (Transcription)","supporting_discovery_ids":[25,32,40]}],"complexes":["MST2–SAV1 (hWW45/Salvador)","MST2–Raf-1 inhibitory complex","MOB1–MST2–NDR1 complex"],"partners":["SAV1","RASSF1A","RAF1","LATS1","MOB1","RASSF5","AKT1","TRIM21"],"other_free_text":[]}},"prefetch_data":{"uniprot":{"accession":"Q9Y6E0","full_name":"Serine/threonine-protein kinase 24","aliases":["Mammalian STE20-like protein kinase 3","MST-3","STE20-like kinase MST3"],"length_aa":443,"mass_kda":49.3,"function":"Serine/threonine-protein kinase that acts on both serine and threonine residues and promotes apoptosis in response to stress stimuli and caspase activation. Mediates oxidative-stress-induced cell death by modulating phosphorylation of JNK1-JNK2 (MAPK8 and MAPK9), p38 (MAPK11, MAPK12, MAPK13 and MAPK14) during oxidative stress. Plays a role in a staurosporine-induced caspase-independent apoptotic pathway by regulating the nuclear translocation of AIFM1 and ENDOG and the DNase activity associated with ENDOG. Phosphorylates STK38L on 'Thr-442' and stimulates its kinase activity. In association with STK26 negatively regulates Golgi reorientation in polarized cell migration upon RHO activation (PubMed:27807006). Also regulates cellular migration with alteration of PTPN12 activity and PXN phosphorylation: phosphorylates PTPN12 and inhibits its activity and may regulate PXN phosphorylation through PTPN12. May act as a key regulator of axon regeneration in the optic nerve and radial nerve. Part of the striatin-interacting phosphatase and kinase (STRIPAK) complexes. STRIPAK complexes have critical roles in protein (de)phosphorylation and are regulators of multiple signaling pathways including Hippo, MAPK, nuclear receptor and cytoskeleton remodeling. Different types of STRIPAK complexes are involved in a variety of biological processes such as cell growth, differentiation, apoptosis, metabolism and immune regulation (PubMed:18782753)","subcellular_location":"Cytoplasm; Nucleus; Membrane","url":"https://www.uniprot.org/uniprotkb/Q9Y6E0/entry"},"depmap":{"release":"DepMap","has_data":true,"is_common_essential":false,"resolved_as":"","url":"https://depmap.org/portal/gene/STK3","classification":"Not Classified","n_dependent_lines":48,"n_total_lines":1208,"dependency_fraction":0.039735099337748346},"opencell":{"profiled":false,"resolved_as":"","ensg_id":"","cell_line_id":"","localizations":[],"interactors":[{"gene":"STK4","stoichiometry":4.0},{"gene":"PHAX","stoichiometry":0.2},{"gene":"RBM39","stoichiometry":0.2},{"gene":"VAPA","stoichiometry":0.2}],"url":"https://opencell.sf.czbiohub.org/search/STK3","total_profiled":1310},"omim":[{"mim_id":"620929","title":"MOB KINASE ACTIVATOR 3A; MOB3A","url":"https://www.omim.org/entry/620929"},{"mim_id":"620110","title":"WW AND C2 DOMAINS-CONTAINING PROTEIN 2; WWC2","url":"https://www.omim.org/entry/620110"},{"mim_id":"616711","title":"TAO KINASE 3; TAOK3","url":"https://www.omim.org/entry/616711"},{"mim_id":"616563","title":"STE20-LIKE PROTEIN KINASE; SLK","url":"https://www.omim.org/entry/616563"},{"mim_id":"615931","title":"PROLINE-RICH PROTEIN 16; PRR16","url":"https://www.omim.org/entry/615931"}],"hpa":{"profiled":true,"resolved_as":"","reliability":"Approved","locations":[{"location":"Intermediate filaments","reliability":"Approved"},{"location":"Rods & Rings","reliability":"Approved"}],"tissue_specificity":"Low tissue specificity","tissue_distribution":"Detected in all","driving_tissues":[],"url":"https://www.proteinatlas.org/search/STK3"},"hgnc":{"alias_symbol":["MST2","KRS1"],"prev_symbol":[]},"alphafold":{"accession":"Q9Y6E0","domains":[{"cath_id":"3.30.200.20","chopping":"2-6_23-111","consensus_level":"high","plddt":85.6376,"start":2,"end":111},{"cath_id":"1.10.510.10","chopping":"116-177_194-302","consensus_level":"high","plddt":94.937,"start":116,"end":302},{"cath_id":"1.10.12.70","chopping":"373-433","consensus_level":"high","plddt":83.8779,"start":373,"end":433}],"viewer_url":"https://alphafold.ebi.ac.uk/entry/Q9Y6E0","model_url":"https://alphafold.ebi.ac.uk/files/AF-Q9Y6E0-F1-model_v6.cif","pae_url":"https://alphafold.ebi.ac.uk/files/AF-Q9Y6E0-F1-predicted_aligned_error_v6.png","plddt_mean":77.62},"mouse_models":{"mgi_url":"https://www.informatics.jax.org/marker/summary?nomen=STK3","jax_strain_url":"https://www.jax.org/strain/search?query=STK3"},"sequence":{"accession":"Q9Y6E0","fasta_url":"https://rest.uniprot.org/uniprotkb/Q9Y6E0.fasta","uniprot_url":"https://www.uniprot.org/uniprotkb/Q9Y6E0/entry","alphafold_viewer_url":"https://alphafold.ebi.ac.uk/entry/Q9Y6E0"}},"corpus_meta":[{"pmid":"19878874","id":"PMC_19878874","title":"Mst1 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caspase-3 cleaves MST2 generating a truncated form that is resistant to dephosphorylation by PP1 and PP2A, in contrast to full-length MST2 which is rapidly dephosphorylated, explaining constitutive activation of the truncated form during apoptosis.\",\n      \"method\": \"Site-directed mutagenesis, in vitro kinase assays, phosphatase treatment, cell transfection with truncation mutants\",\n      \"journal\": \"The Journal of biological chemistry\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 / Moderate — reconstitution with mutagenesis and phosphatase assays, multiple orthogonal approaches, single lab\",\n      \"pmids\": [\"12554736\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2004,\n      \"finding\": \"Raf-1 suppresses MST2 (STK3) activation by preventing its dimerization and activation-loop phosphorylation independently of Raf-1 kinase activity; depletion of MST2 from Raf-1-/- cells abrogates apoptosis hypersensitivity, placing MST2 downstream of Raf-1 in an apoptosis-suppressive pathway.\",\n      \"method\": \"Proteomic analysis of Raf-1 signaling complexes, Co-IP, siRNA knockdown, genetic epistasis (Raf-1-/- cells), overexpression\",\n      \"journal\": \"Science\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — reciprocal Co-IP, genetic epistasis with defined phenotype, replicated in mouse and human cells\",\n      \"pmids\": [\"15618521\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2005,\n      \"finding\": \"MST2 (STK3) phosphorylates LATS1 at its activation loop (S909) and hydrophobic motif (T1079), activating LATS1 kinase activity; MST2 also directly interacts with hWW45 (Salvador ortholog) via their conserved domains.\",\n      \"method\": \"In vitro kinase assay, mass spectrometry phosphosite mapping, deletion analysis, Co-IP\",\n      \"journal\": \"Oncogene\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 / Moderate — in vitro reconstitution with phosphosite identification by MS, multiple orthogonal methods, single lab\",\n      \"pmids\": [\"15688006\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2006,\n      \"finding\": \"MST1 and MST2 (STK3) heterodimerize with hSav (hWW45/Salvador) via conserved C-terminal coiled-coil (SARAH) domains; this interaction leads to hSav phosphorylation by Mst kinases and stabilization of hSav protein, with stabilization requiring the Mst-hSav interaction but not necessarily phosphorylation.\",\n      \"method\": \"Co-IP, deletion mutagenesis, in vitro phosphorylation, co-expression abundance assays\",\n      \"journal\": \"The FEBS journal\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — reciprocal Co-IP, in vitro phosphorylation, mutagenesis, single lab\",\n      \"pmids\": [\"16930133\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2007,\n      \"finding\": \"RASSF1A disrupts the inhibitory Raf-1–MST2 complex, enhancing MST2 interaction with LATS1; RASSF1A-activated LATS1 phosphorylates YAP1, releasing it to translocate to the nucleus and associate with p73, driving transcription of the proapoptotic gene PUMA.\",\n      \"method\": \"Co-IP, siRNA knockdown, epistasis analysis, reporter assays, apoptosis assays\",\n      \"journal\": \"Molecular cell\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — reciprocal Co-IP, genetic epistasis, pathway reconstitution with defined molecular readouts, replicated across multiple cell types\",\n      \"pmids\": [\"17889669\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2008,\n      \"finding\": \"MST1 and MST2 (STK3) phosphorylate MOBKL1A/MOBKL1B (Mats homologs) as preferred substrates in vitro and in cells during mitosis; MST1/2 activity increases during mitosis (especially in nocodazole arrest); MOBKL1A/B phosphorylation promotes their binding to LATS1 and enables LATS1 activation-loop phosphorylation; replacing endogenous MOBKL1A/B with non-phosphorylatable mutants accelerates cell proliferation through G1/S and mitotic exit.\",\n      \"method\": \"In vitro kinase assay, cell-based phosphorylation (okadaic acid/H2O2 treatment), Co-IP, dominant-negative replacement, cell cycle analysis\",\n      \"journal\": \"Current biology\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 / Strong — in vitro reconstitution, multiple orthogonal methods (in vitro and in vivo phosphorylation, functional rescue), multiple perturbations\",\n      \"pmids\": [\"18328708\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2008,\n      \"finding\": \"MST2 (STK3) cooperates with LATS1 kinase to phosphorylate YAP at Ser-127, requiring WW domains of YAP and PPxY motifs in LATS kinases; in HEK293 cells this Hippo pathway reconstitution is anti-apoptotic, partially rescuing cells from YAP2-induced apoptotic death; nuclear localization of YAP2 is negatively regulated by LATS1.\",\n      \"method\": \"Reconstitution in human cells, Co-IP with deletion/point mutants, subcellular fractionation/localization, apoptosis assays\",\n      \"journal\": \"The Journal of biological chemistry\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — functional reconstitution with domain mapping, single lab, multiple readouts\",\n      \"pmids\": [\"18640976\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2008,\n      \"finding\": \"MST2 phosphorylates MOB1 at Thr74 in vitro; this phosphorylation is essential for forming the MOB1–MST2–NDR1 complex and for full activation of NDR1; MOB1 T74A mutant fails to enhance its interaction with NDR1 upon okadaic acid stimulation.\",\n      \"method\": \"In vitro kinase assay, site-directed mutagenesis (T74A, T181A), Co-IP, NDR1 activation assay\",\n      \"journal\": \"Oncogene\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 / Moderate — in vitro phosphorylation with site mutagenesis validated in cells, multiple orthogonal approaches\",\n      \"pmids\": [\"18362890\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2009,\n      \"finding\": \"MST2 (STK3) promotes MST2-dependent NDR1 activation and precise chromosome alignment at metaphase; depletion of MST2 or Fry or MOB2 causes mitotic chromosome misalignment; expression of active NDR1 rescues MST2-depletion misalignment; Fry acts as a scaffold binding NDR1, MOB2, and microtubules on the spindle.\",\n      \"method\": \"siRNA knockdown, live-cell imaging, rescue by active NDR1, biochemical interaction (Co-IP/pulldown)\",\n      \"journal\": \"Current biology\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — loss-of-function with specific mitotic phenotype and epistasis rescue, single lab\",\n      \"pmids\": [\"19327996\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2009,\n      \"finding\": \"In mouse liver, Mst1 and Mst2 are cleaved and constitutively activated; combined deficiency results in loss of inhibitory Ser127 phosphorylation of Yap1 and hepatocellular carcinoma; Mst1/2 inactivates Yap1 through an intermediary kinase distinct from Lats1/2 in the liver context.\",\n      \"method\": \"Conditional knockout mice, phospho-specific western blot, re-expression rescue in HCC cell lines\",\n      \"journal\": \"Cancer cell\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — in vivo genetic model with defined molecular (phosphorylation) and cellular (tumor suppression) phenotypes, reinforced by cell-line rescue\",\n      \"pmids\": [\"19878874\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2009,\n      \"finding\": \"RASSF2 directly binds and stabilizes MST2 (STK3) protein at endogenous levels; RASSF2 co-localizes with MST2 in the cytoplasm (shifting MST2 from nuclear-alone); RASSF2 expression increases MST2 protein levels and protects MST2 from proteolytic degradation; RASSF2 itself is phosphorylated by co-immunoprecipitated MST1/2.\",\n      \"method\": \"Endogenous Co-IP, co-localization imaging, siRNA knockdown, immunoblotting for protein stability\",\n      \"journal\": \"Oncogene\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — endogenous reciprocal Co-IP with localization and stability assays, single lab\",\n      \"pmids\": [\"19525978\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2010,\n      \"finding\": \"Akt phosphorylates MST2 at T117 and T384; these phosphorylations inhibit MST2 by (1) blocking its binding to RASSF1A, (2) promoting its association into the Raf-1 inhibitory complex, and (3) preventing MST2 homodimerization required for activation; Akt thus limits MST2 proapoptotic activity downstream of mitogens and oncogenic Ras.\",\n      \"method\": \"In vitro kinase assay, site-directed mutagenesis (T117A/T384A), Co-IP, epistasis with Ras effector domain mutants\",\n      \"journal\": \"Cancer research\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 / Moderate — in vitro phosphorylation with mutagenesis validated by Co-IP and pathway readout, multiple orthogonal methods\",\n      \"pmids\": [\"20086174\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2010,\n      \"finding\": \"Akt phosphorylates MST2 at T117, leading to inhibition of MST2 cleavage, nuclear translocation, autophosphorylation at Thr180, and kinase activity; IGF1-PI3K-Akt signaling suppresses DNA damage-induced MST2 activation.\",\n      \"method\": \"In vitro and in vivo labeling kinase assays, site-directed mutagenesis, immunoblotting\",\n      \"journal\": \"PloS one\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 1 / Weak — in vitro kinase assay with mutagenesis, single lab, partially overlapping findings with Romano 2010\",\n      \"pmids\": [\"20231902\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2010,\n      \"finding\": \"A-Raf prevents MST2-mediated apoptosis in cancer cells by sequestering and inactivating MST2; this requires hnRNP H-dependent A-Raf transcription; siRNA knockdown of hnRNP H or A-Raf results in MST2-dependent apoptosis.\",\n      \"method\": \"siRNA knockdown, Co-IP, apoptosis assays\",\n      \"journal\": \"Cancer research\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — Co-IP with siRNA epistasis and defined apoptotic phenotype, single lab\",\n      \"pmids\": [\"20145135\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2011,\n      \"finding\": \"The tumor suppressor RASSF1A prevents PP2A-mediated dephosphorylation of MST1/2 at their activation-loop residues (Thr183/Thr180), maintaining MST2 in an activated, autophosphorylated state; RASSF1A also stabilizes MST2 protein.\",\n      \"method\": \"Dephosphorylation assays with PP2A, PP2A knockdown, RASSF1A depletion, phospho-specific western blot\",\n      \"journal\": \"The Journal of biological chemistry\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 / Moderate — direct dephosphorylation assays combined with PP2A knockdown and RASSF1A depletion, multiple orthogonal approaches, single lab\",\n      \"pmids\": [\"21199877\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2011,\n      \"finding\": \"Mutant K-Ras directly binds RASSF1A to activate the apoptotic MST2–LATS1 pathway; activated LATS1 sequesters Mdm2 to stabilize p53 and drive apoptosis; wild-type K-Ras inhibits this pathway via AKT activation.\",\n      \"method\": \"Co-IP, siRNA knockdown, epistasis analysis with Ras effector domain mutants, apoptosis assays\",\n      \"journal\": \"Molecular cell\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — reciprocal Co-IP, genetic epistasis across multiple cell lines, pathway mechanism mapped to p53 stabilization\",\n      \"pmids\": [\"22195963\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2012,\n      \"finding\": \"Mst1 and Mst2 (STK3) control Rho GTPase activation (Rac1 and RhoA GTP charging) in SP thymocytes; when phosphorylated by Mst1/2, Mob1 binds and activates the Rac1 guanyl nucleotide exchanger Dock8; this pathway drives sphingosine-1-phosphate and CCL21-induced cell migration and thymic egress.\",\n      \"method\": \"Conditional double knockout mice, Co-IP, GTP-loading assays, migration assays, phospho-Mob1 blotting\",\n      \"journal\": \"The Journal of experimental medicine\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — in vivo genetic model with defined molecular mechanism (Mob1 phosphorylation → Dock8 activation → Rac1 GTP charging) and cellular phenotype\",\n      \"pmids\": [\"22412158\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2013,\n      \"finding\": \"Crystal structure of human MST2 kinase domain alone and in complex with RASSF5 (SARAH domain) reveals that MST2 undergoes activation via trans-autophosphorylation at its activation loop, requiring SARAH domain-mediated homodimerization; RASSF5 binding disrupts MST2 homodimer, blocking autoactivation of unphosphorylated MST2 but not inhibiting already-activated MST2, revealing temporal context-dependent dual roles for RASSF5.\",\n      \"method\": \"X-ray crystallography, biochemical autophosphorylation assays, mutagenesis\",\n      \"journal\": \"Structure\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 / Strong — crystal structure plus functional biochemical validation with mutagenesis, multiple orthogonal approaches\",\n      \"pmids\": [\"23972470\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2013,\n      \"finding\": \"Chronic (not transient) K-Ras activation drives MST2–LATS1 complex formation and apoptosis; transient EGF-stimulated K-Ras activation prevents MST2–LATS1 complex formation in an AKT-dependent manner, demonstrating that activation kinetics determine pathway output.\",\n      \"method\": \"Co-IP, siRNA, AKT inhibition, overexpression of Ras exchange factors, apoptosis assays\",\n      \"journal\": \"Molecular and cellular biology\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — Co-IP with multiple genetic and pharmacological perturbations, single lab\",\n      \"pmids\": [\"23459937\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2014,\n      \"finding\": \"c-Abl tyrosine kinase phosphorylates MST2 at the conserved site Y81 within the kinase domain; this phosphorylation disrupts MST2–Raf-1 interaction, enhances MST2 homodimerization and kinase activation, and induces neuronal cell death.\",\n      \"method\": \"In vitro kinase assay, site-directed mutagenesis (Y81F), Co-IP, cell death assay\",\n      \"journal\": \"PloS one\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 1 / Weak — in vitro kinase assay with mutagenesis validated in cells, single lab\",\n      \"pmids\": [\"22590567\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2014,\n      \"finding\": \"Competing protein interactions coordinate Raf-1–MST2 crosstalk: Akt phosphorylation of MST2 and LATS1-feedback phosphorylation of Raf-1 Ser259 enable Raf-1 to suppress both MST2 and MEK signaling; mutation of Raf-1 Ser259 simultaneously drives apoptosis and proliferation; concomitant MST2 downregulation switches outcome to proliferation and transformation.\",\n      \"method\": \"Phosphorylation assays, Co-IP, mutagenesis, mathematical modeling validated by experiment, cell transformation assays\",\n      \"journal\": \"Nature cell biology\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 / Strong — mathematical model-guided mutagenesis with multiple orthogonal experimental validations including functional cell fate readouts\",\n      \"pmids\": [\"24929361\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2014,\n      \"finding\": \"STK3 (MST2) and STK4 (MST1) phosphorylate LC3 at threonine 50 (Thr50); loss of this phosphorylation blocks autophagosome–lysosome fusion and impairs intracellular bacterial clearance; constitutive phosphomimetic LC3-T50E reverses autophagy block in STK3/STK4-deficient cells.\",\n      \"method\": \"In vitro kinase assay, site-directed mutagenesis, autophagy flux assays, bacterial clearance assay, genetic rescue\",\n      \"journal\": \"Molecular cell\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 / Strong — in vitro reconstitution with mutagenesis, genetic rescue, conserved across species, multiple functional readouts\",\n      \"pmids\": [\"25544559\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2014,\n      \"finding\": \"Crystal structure of the MST2 SARAH domain shows it forms an antiparallel homodimeric coiled coil; structure-guided mutagenesis identified interface residues critical for homodimerization; these same mutations also impaired heterodimerization with RAPL (RASSF5); SARAH-mediated homodimerization and RAPL heterointeraction are both required for full MST2 activation and apoptotic functions in T cells.\",\n      \"method\": \"X-ray crystallography, site-directed mutagenesis, Co-IP, kinase activity assays, apoptosis assays\",\n      \"journal\": \"Journal of structural biology\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 / Strong — crystal structure with mutagenesis validated by functional kinase and apoptosis assays\",\n      \"pmids\": [\"24468289\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2014,\n      \"finding\": \"MST2 (STK3) kinase suppresses rDNA transcription in response to DNA damage by phosphorylating histone H2B at serine 14 (H2BS14p) in an ATM-dependent manner; MST2 localizes to nucleoli; H2BS14p marks transcriptionally inactive nucleolar chromatin and is necessary for rDNA transcriptional shutdown and genomic integrity maintenance.\",\n      \"method\": \"ChIP, immunofluorescence localization, siRNA knockdown, kinase assays, ATM inhibition\",\n      \"journal\": \"The EMBO journal\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — direct localization experiment tied to functional consequence (rDNA transcription), ChIP with defined histone phosphorylation mark, multiple orthogonal approaches\",\n      \"pmids\": [\"29789391\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2015,\n      \"finding\": \"Mst1 and Mst2 (STK3) activate the GTPase Rac to promote TLR-triggered assembly of the TRAF6–ECSIT complex, which recruits mitochondria to phagosomes for synergistic ROS production and bactericidal activity.\",\n      \"method\": \"Conditional knockout mice, Co-IP, Rac GTP-loading assays, mitochondria trafficking (imaging), ROS measurement, bacterial killing assays\",\n      \"journal\": \"Nature immunology\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — in vivo genetic model, Co-IP, mechanistic signaling axis defined with multiple orthogonal readouts\",\n      \"pmids\": [\"26414765\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2016,\n      \"finding\": \"XMU-MP-1 is a selective, reversible MST1/2 inhibitor confirmed by co-crystal structure; it blocks MST1/2 kinase activities, activates downstream YAP, and promotes tissue repair in mouse intestinal and liver injury models.\",\n      \"method\": \"Enzyme-based HTS, co-crystal structure, structure-activity relationship, in vivo pharmacology\",\n      \"journal\": \"Science translational medicine\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 / Strong — co-crystal structure with SAR and multiple in vivo models confirming on-target activity\",\n      \"pmids\": [\"27535619\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2016,\n      \"finding\": \"H-Ras promotes formation of inactive Mst1–Mst2 heterodimers via their SARAH domains in an ERK-dependent manner; Mst1/Mst2 heterodimers have much-reduced kinase activity compared to homodimers; cells lacking Mst1 (unable to form heterodimers) are resistant to H-Ras transformation and maintain active Hippo signaling.\",\n      \"method\": \"Co-IP, kinase activity assays comparing homo- vs heterodimers, ERK pathway inhibition, Mst1 knockout cells, transformation assays\",\n      \"journal\": \"Current biology\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — Co-IP with kinase activity comparison and genetic validation, single lab\",\n      \"pmids\": [\"27238285\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2016,\n      \"finding\": \"CDK1 phosphorylates MST2 at serine 385 during mitosis in vitro and in vivo; this phosphorylation does not affect MST2 kinase activity or Hippo-YAP signaling, but phosphorylation-deficient MST2-S385A has higher activity in suppressing proliferation and tumorigenesis, establishing CDK1-mediated mitotic regulation of MST2 tumor suppression.\",\n      \"method\": \"In vitro CDK1 kinase assay, phospho-site mutagenesis (S385A), cell proliferation and anchorage-independent growth assays, in vivo xenograft\",\n      \"journal\": \"Cellular signalling\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 1 / Weak — in vitro kinase assay with mutagenesis and in vivo tumor assay, single lab\",\n      \"pmids\": [\"27566175\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2016,\n      \"finding\": \"A-Raf subcellular localization regulates MST2 activity: A-Raf at mitochondria (in proliferating/cancer cells) efficiently sequesters and inhibits MST2, while A-Raf relocated to the plasma membrane (during differentiation) loses its ability to inhibit MST2; this redistribution is regulated by the scaffold KSR2.\",\n      \"method\": \"Subcellular fractionation, live-cell imaging, siRNA knockdown of KSR2, apoptosis assays\",\n      \"journal\": \"Cell death and differentiation\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — direct localization experiments tied to functional consequence on MST2 activity and apoptosis, single lab\",\n      \"pmids\": [\"26891695\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2018,\n      \"finding\": \"Mst1 and Mst2 amplify IL-2–STAT5 signaling in regulatory T cells by promoting STAT5 activation; Mst1 associates with the cytoskeletal DOCK8–LRCH module; Mst1 deficiency limits Treg cell migration and access to IL-2, and impairs Rac GTPase activity mediating STAT5 activation.\",\n      \"method\": \"Conditional knockout mice, quantitative proteomics (unbiased Co-IP-MS), phospho-STAT5 assays, migration assays, Rac activation assays\",\n      \"journal\": \"Immunity\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — unbiased quantitative proteomics identifying DOCK8-LRCH complex, validated by genetic loss-of-function with multiple mechanistic readouts\",\n      \"pmids\": [\"30413360\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2019,\n      \"finding\": \"MST2 phosphorylates Runx2 at Ser339 and Ser370 (identified by mass spectrometry); MST2 interacts with Runx2 via SAV1's WW domain binding to a PY motif (aa 292–445) of Runx2; phosphorylation inhibits Runx2 transcriptional activity and osteoblast differentiation; phosphorylation-deficient double mutant (S339A/S370A) is resistant to MST2/SAV1-mediated inhibition.\",\n      \"method\": \"In vitro kinase assay, mass spectrometry phosphosite mapping, Co-IP domain mapping, reporter assay, differentiation assays, mutagenesis\",\n      \"journal\": \"Biochemical and biophysical research communications\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 1 / Moderate — in vitro kinase assay with MS-identified sites, functional mutagenesis, and cellular differentiation readouts, single lab\",\n      \"pmids\": [\"30910359\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2020,\n      \"finding\": \"Increasing proximity of adjacent MST2 kinase domains—via SARAH domain homodimerization, membrane recruitment, or complex formation with SAV1—is sufficient to trigger MST2 autophosphorylation; a specific protein assembly is not required; multiple upstream activating events converge on this common molecular mechanism.\",\n      \"method\": \"Chemically induced dimerization, single-molecule pulldown, in vitro biochemistry, cell-based autophosphorylation assays\",\n      \"journal\": \"The Journal of biological chemistry\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 / Strong — multiple orthogonal biochemical and cell-based methods with chemically induced dimerization as controlled perturbation\",\n      \"pmids\": [\"32994222\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2021,\n      \"finding\": \"STK3 and STK4 (MST2/MST1) suppress mitochondrial capacity and mitophagy in adipocytes; mechanistically, STK3/4 regulate phosphorylation and dimerization status of the mitophagy receptor BNIP3; genetic inactivation of Stk3/4 increases mitochondrial mass/function and stabilizes UCP1 in beige adipose tissue.\",\n      \"method\": \"Conditional knockout mice, Co-IP, BNIP3 phosphorylation/dimerization assays, mitochondrial function assays\",\n      \"journal\": \"Nature metabolism\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — in vivo genetic model with defined molecular mechanism (BNIP3 phosphorylation/dimerization) and multiple metabolic readouts\",\n      \"pmids\": [\"33758424\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2022,\n      \"finding\": \"MST2 is ubiquitinated at K473 via K63-linkage by the E3 ligase TRIM21; this ubiquitination promotes MST2 homodimerization and enhances its kinase activity, leading to YAP inactivation and inhibition of EMT in colorectal cancer.\",\n      \"method\": \"Co-IP, site-directed mutagenesis (K473), in vitro ubiquitination assay, kinase activity assay, tumor organoid models\",\n      \"journal\": \"Cell chemical biology\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — Co-IP with mutagenesis and functional kinase assay, single lab\",\n      \"pmids\": [\"37354905\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2022,\n      \"finding\": \"MST2 binds SCFβTrCP E3 ubiquitin ligase via a non-canonical degradation motif and is degraded via proteasomal proteolysis; stiffer extracellular matrix and integrin hyperactivation enhance MST2 degradation through integrin-linked kinase (ILK) and actomyosin stress fibers.\",\n      \"method\": \"Co-IP, siRNA knockdown of βTrCP, site-directed mutagenesis, proteasome inhibition, molecular dynamics simulation\",\n      \"journal\": \"Biochimica et biophysica acta. General subjects\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — Co-IP with mutagenesis and functional proteolysis assays, computational validation, single lab\",\n      \"pmids\": [\"36044955\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2023,\n      \"finding\": \"PRMT5 symmetrically di-methylates MST2 (STK3) at R461 and R467 in its SARAH domain, suppressing MST2 autophosphorylation and kinase activity by blocking homodimerization, thereby inactivating Hippo signaling in pancreatic cancer.\",\n      \"method\": \"In vitro methylation assay, site-directed mutagenesis (R461/R467), dimerization assays, kinase activity assays, Co-IP, in vivo xenograft with PRMT5 inhibitor\",\n      \"journal\": \"The EMBO journal\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 / Moderate — in vitro biochemical reconstitution with mutagenesis, dimerization assay, functional kinase assay, and in vivo validation\",\n      \"pmids\": [\"37905571\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2023,\n      \"finding\": \"TRIM69 binds MST2 directly, redistributes it to the perinuclear cytoskeleton, promotes MST2 association with PLK1, and stimulates MST2 phosphorylation at S15 (a PLK1 site critical for centrosome disjunction); TRIM69 ablation causes centrosome scattering and chromosome segregation defects.\",\n      \"method\": \"Co-IP, immunofluorescence localization, phospho-S15 western blot, siRNA knockdown with centrosome and mitosis phenotype readout\",\n      \"journal\": \"Nucleic acids research\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — Co-IP with localization experiment tied to functional centrosome phenotype, single lab\",\n      \"pmids\": [\"37739411\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2024,\n      \"finding\": \"In gastric cancer, STK3 directly interacts with GSK-3β and phosphorylates it, promoting GSK-3β degradation and β-catenin nuclear accumulation, thereby activating Wnt/β-catenin signaling; STK3 is itself a transcriptional target of YAP1, creating a feedback loop.\",\n      \"method\": \"Co-IP, in vitro kinase assay, ChIP-qPCR, knockout mouse models, functional rescue assays, xenograft models\",\n      \"journal\": \"Molecular cancer\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — Co-IP with in vitro kinase assay and functional genetic validation, single lab\",\n      \"pmids\": [\"40604818\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2024,\n      \"finding\": \"STK3 (MST2) activation phosphorylates FOXO1 at Ser212, promoting its nuclear translocation and transcriptional activation of TP53INP1 and P21 to inhibit ESCC cell proliferation and migration; cellular ROS activates STK3 autophosphorylation in ESCC cells.\",\n      \"method\": \"Co-IP, immunofluorescence, ChIP, western blot, siRNA, xenograft models\",\n      \"journal\": \"Cellular oncology\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — Co-IP with ChIP and IF confirming nuclear translocation, multiple readouts, single lab\",\n      \"pmids\": [\"38436783\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2016,\n      \"finding\": \"STK3 (MST2) interacts with apoptosis-inducing factor (AIF); AIF interaction enhances STK3 phosphorylation at Thr180, suggesting AIF is a positive regulator of STK3 activity.\",\n      \"method\": \"Co-IP, in vitro phosphorylation assay\",\n      \"journal\": \"PloS one\",\n      \"confidence\": \"Low\",\n      \"confidence_rationale\": \"Tier 3 / Weak — single Co-IP and phosphorylation assay, no mechanistic follow-up, single lab\",\n      \"pmids\": [\"24992339\"],\n      \"is_preprint\": false\n    }\n  ],\n  \"current_model\": \"STK3 (MST2) is a SARAH domain-containing Ste20-related serine/threonine kinase that is activated by trans-autophosphorylation at Thr180 upon kinase domain proximity (driven by SARAH-mediated homodimerization, membrane recruitment, or SAV1 binding); it is positively regulated by RASSF1A (which protects its activation-loop phosphorylation from PP2A), RASSF2, TRIM21 (K63 ubiquitination at K473), c-Abl (Y81 phosphorylation), and AIF, and negatively regulated by Raf-1/A-Raf (blocking dimerization), Akt (T117/T384 phosphorylation promoting Raf-1 binding and blocking homodimerization), CDK1 (S385 mitotic phosphorylation), PRMT5 (R461/R467 methylation blocking homodimerization), and SCFβTrCP-mediated proteasomal degradation; active STK3 phosphorylates LATS1/2 (S909, T1079), MOBKL1A/B (Thr74), YAP (Ser127), LC3 (Thr50), histone H2B (Ser14), Runx2 (Ser339/Ser370), FOXO1 (Ser212), GSK-3β, and TTF-1, thereby suppressing YAP/TAZ oncogenic transcription, promoting autophagosome–lysosome fusion, suppressing rDNA transcription after DNA damage, regulating mitophagy via BNIP3, and controlling T-cell migration via Mob1–Dock8–Rac1 signaling.\"\n}\n```","stage2_raw":"{\n  \"mechanistic_narrative\": \"STK3 (MST2) is a stress-responsive Ste20-related serine/threonine kinase and the core kinase of the mammalian Hippo tumor-suppressor pathway, activated by apoptotic and stress stimuli rather than mitogens [#0]. Its activity depends on trans-autophosphorylation at Thr180 within the activation loop, a reaction triggered whenever adjacent kinase domains are brought into proximity—by SARAH-domain-mediated antiparallel homodimerization, membrane recruitment, or SAV1 (hWW45/Salvador) complex formation—so that no single dedicated assembly is required for activation [#2, #19, #24, #33]. Once active, STK3 propagates Hippo signaling by phosphorylating the adaptor MOBKL1A/B (MOB1) at Thr74 and the downstream kinases LATS1 (S909/T1079) and NDR1, which in turn inactivate the transcriptional coactivator YAP via Ser127 phosphorylation, suppressing YAP/TAZ-driven proliferation and promoting apoptosis [#4, #7, #8, #9, #11]. This module enforces tumor suppression in vivo, as combined Mst1/Mst2 loss in liver causes loss of YAP Ser127 phosphorylation and hepatocellular carcinoma [#11]. STK3 activation is gated by an extensive regulatory network: it is held inactive by Raf-1 and A-Raf (which block dimerization and sequester the kinase) [#3, #15, #30], by Akt phosphorylation at T117/T384 that drives Raf-1 binding and blocks homodimerization [#13, #14], by CDK1 (S385), by PRMT5 methylation of R461/R467 in the SARAH domain, and by SCF\\u03b2TrCP-mediated degradation [#29, #37, #36]; it is positively regulated by RASSF1A, which protects activation-loop phosphorylation from PP2A and disrupts the inhibitory Raf-1 complex, by RASSF2 stabilization, by c-Abl phosphorylation at Y81, and by TRIM21 K63-ubiquitination at K473 [#16, #6, #12, #21, #35]. The K-Ras\\u2013RASSF1A\\u2013MST2\\u2013LATS1 axis couples Ras signaling kinetics to an apoptotic, p53-stabilizing output [#17, #20, #22]. Beyond canonical Hippo signaling, STK3 phosphorylates LC3 at Thr50 to enable autophagosome\\u2013lysosome fusion and bacterial clearance [#23], phosphorylates histone H2B at Ser14 in nucleoli to shut down rDNA transcription after DNA damage [#25], regulates Rac/RhoA GTPase signaling through Mob1\\u2013Dock8 to drive thymocyte and Treg migration and TLR-triggered antibacterial ROS production [#18, #26, #31], and controls mitophagy via the receptor BNIP3 in adipocytes [#34]. The small-molecule inhibitor XMU-MP-1 confirms these activities pharmacologically and activates YAP to promote tissue repair [#27].\",\n  \"teleology\": [\n    {\n      \"year\": 1996,\n      \"claim\": \"Established STK3 as a kinase activated by stress and apoptotic stimuli but not mitogens, defining its biological niche before any pathway context existed.\",\n      \"evidence\": \"Protein purification, cloning, and kinase activity assays\",\n      \"pmids\": [\"8816758\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"No substrates or upstream regulators identified\", \"Activation mechanism unknown\"]\n    },\n    {\n      \"year\": 1998,\n      \"claim\": \"Identified TTF-1 as the first substrate of this kinase class, providing initial evidence that STK3 directly phosphorylates a defined cellular target.\",\n      \"evidence\": \"In-gel and in vitro kinase assays with in vivo phosphosite mapping in rat ortholog\",\n      \"pmids\": [\"9430685\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Physiological pathway linking STK3 to TTF-1 not established\", \"Single substrate without broader signaling context\"]\n    },\n    {\n      \"year\": 2003,\n      \"claim\": \"Defined the core activation mechanism—trans-autophosphorylation at Thr180—and showed caspase cleavage generates a dephosphorylation-resistant constitutively active form, linking activation to apoptosis.\",\n      \"evidence\": \"Mutagenesis, in vitro kinase and phosphatase assays, truncation mutants in cells\",\n      \"pmids\": [\"12554736\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"What drives the initial dimerization in cells not yet defined\", \"Downstream effectors of cleaved form unknown\"]\n    },\n    {\n      \"year\": 2004,\n      \"claim\": \"Placed STK3 downstream of Raf-1 in an apoptosis-suppressive pathway, revealing a kinase-independent inhibitory mechanism acting on dimerization.\",\n      \"evidence\": \"Proteomics, reciprocal Co-IP, siRNA, Raf-1-/- genetic epistasis\",\n      \"pmids\": [\"15618521\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"How RASSF proteins relieve this inhibition not yet shown\", \"Structural basis of Raf-1 block undefined\"]\n    },\n    {\n      \"year\": 2005,\n      \"claim\": \"Connected STK3 to the Hippo core by showing it phosphorylates and activates LATS1 and binds Salvador, establishing the kinase cascade architecture.\",\n      \"evidence\": \"In vitro kinase assay, MS phosphosite mapping, deletion analysis, Co-IP\",\n      \"pmids\": [\"15688006\", \"16930133\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Role of YAP as terminal effector not yet linked\", \"In vivo relevance of LATS activation not tested\"]\n    },\n    {\n      \"year\": 2008,\n      \"claim\": \"Defined MOB1 (Thr74) phosphorylation as the adaptor step enabling LATS1/NDR1 activation and showed the module restrains cell-cycle progression, completing the kinase relay logic.\",\n      \"evidence\": \"In vitro and cellular kinase assays, Co-IP, dominant-negative replacement, cell cycle and NDR1 activation assays\",\n      \"pmids\": [\"18328708\", \"18362890\", \"18640976\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Whether MOB1 phosphorylation is the sole route to LATS activation unclear\", \"Mitotic versus interphase roles not fully separated\"]\n    },\n    {\n      \"year\": 2007,\n      \"claim\": \"Showed RASSF1A relieves Raf-1 inhibition to drive a proapoptotic MST2\\u2013LATS1\\u2013YAP\\u2013p73 axis, establishing how upstream signals switch the pathway toward apoptosis.\",\n      \"evidence\": \"Co-IP, siRNA, epistasis, reporter and apoptosis assays\",\n      \"pmids\": [\"17889669\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"How RASSF1A physically displaces Raf-1 not resolved\", \"Tissue specificity of p73 versus YAP outputs unclear\"]\n    },\n    {\n      \"year\": 2009,\n      \"claim\": \"Validated STK3 as a bona fide tumor suppressor in vivo and identified RASSF2 as a stabilizing partner, distinguishing context-dependent (liver) effector usage.\",\n      \"evidence\": \"Conditional knockout mice, phospho-specific blotting, endogenous Co-IP, localization and stability assays\",\n      \"pmids\": [\"19878874\", \"19525978\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Identity of the liver intermediary kinase distinct from LATS unknown\", \"Mechanism of RASSF2-mediated stabilization undefined\"]\n    },\n    {\n      \"year\": 2010,\n      \"claim\": \"Mapped the inhibitory inputs Akt (T117/T384) and A-Raf, showing how mitogenic and oncogenic signaling restrain STK3 proapoptotic activity.\",\n      \"evidence\": \"In vitro kinase assays, mutagenesis, Co-IP, Ras effector epistasis, apoptosis assays\",\n      \"pmids\": [\"20086174\", \"20231902\", \"20145135\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Quantitative balance between activating and inhibitory inputs unclear\", \"Whether these regulators act simultaneously not resolved\"]\n    },\n    {\n      \"year\": 2011,\n      \"claim\": \"Established that RASSF1A protects STK3 activation-loop phosphorylation from PP2A and that mutant K-Ras engages this axis to stabilize p53, linking Ras mutation status to apoptotic output.\",\n      \"evidence\": \"PP2A dephosphorylation assays, knockdowns, Co-IP, Ras effector epistasis, apoptosis assays\",\n      \"pmids\": [\"21199877\", \"22195963\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"How wild-type versus mutant K-Ras diverge mechanistically only partly defined\", \"PP2A targeting specificity not fully mapped\"]\n    },\n    {\n      \"year\": 2012,\n      \"claim\": \"Revealed a non-Hippo function: STK3-phosphorylated Mob1 activates the Rac-GEF Dock8 to drive GTPase signaling and lymphocyte migration, broadening STK3 into cytoskeletal regulation.\",\n      \"evidence\": \"Conditional double-knockout mice, Co-IP, GTP-loading and migration assays\",\n      \"pmids\": [\"22412158\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"How the same Mob1 phosphorylation switches between LATS and Dock8 outputs unclear\", \"Direct STK3-Dock8 contact not defined\"]\n    },\n    {\n      \"year\": 2013,\n      \"claim\": \"Provided structural proof that SARAH-mediated homodimerization enables trans-autophosphorylation and that RASSF5 binding has temporally dual roles, resolving the activation switch at atomic resolution.\",\n      \"evidence\": \"X-ray crystallography of kinase and SARAH-RASSF5 complexes, autophosphorylation assays, mutagenesis; plus Co-IP/AKT epistasis of Ras kinetics\",\n      \"pmids\": [\"23972470\", \"23459937\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"How membrane recruitment substitutes for homodimerization not addressed here\", \"Dynamics of dimer-to-monomer transitions unresolved\"]\n    },\n    {\n      \"year\": 2014,\n      \"claim\": \"Expanded STK3 substrate repertoire (LC3 Thr50 for autophagy, H2B Ser14 for rDNA shutdown) and defined SARAH homodimer interface, c-Abl Y81 activation, and Raf-1 crosstalk circuitry, showing STK3 acts well beyond canonical Hippo.\",\n      \"evidence\": \"Crystallography, in vitro kinase assays, mutagenesis, autophagy/bacterial clearance and ChIP assays, mathematical modeling\",\n      \"pmids\": [\"25544559\", \"24468289\", \"29789391\", \"22590567\", \"24929361\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Whether autophagy and Hippo functions are mechanistically separable unclear\", \"Coordination among multiple substrate programs undefined\"]\n    },\n    {\n      \"year\": 2015,\n      \"claim\": \"Showed STK3 activates Rac to assemble the TRAF6\\u2013ECSIT complex and recruit mitochondria to phagosomes, establishing a role in innate antibacterial ROS production.\",\n      \"evidence\": \"Conditional knockout mice, Co-IP, Rac GTP-loading, mitochondrial imaging, ROS and killing assays\",\n      \"pmids\": [\"26414765\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Direct STK3 substrate in this axis not defined\", \"Relationship to Mob1-Dock8 module unclear\"]\n    },\n    {\n      \"year\": 2016,\n      \"claim\": \"Layered additional regulation—CDK1 mitotic phosphorylation, ERK-dependent inactive Mst1/Mst2 heterodimers, A-Raf subcellular relocalization, and the XMU-MP-1 inhibitor—refining how STK3 activity is tuned spatially and temporally.\",\n      \"evidence\": \"In vitro kinase assays, mutagenesis, Co-IP, fractionation, co-crystal structure, in vivo pharmacology\",\n      \"pmids\": [\"27566175\", \"27238285\", \"26891695\", \"27535619\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Functional weight of heterodimerization in vivo unclear\", \"How CDK1 site affects non-Hippo functions untested\"]\n    },\n    {\n      \"year\": 2018,\n      \"claim\": \"Extended the cytoskeletal-migration role to Treg cells via a DOCK8\\u2013LRCH module amplifying IL-2\\u2013STAT5 signaling, reinforcing STK3 control of immune cell positioning.\",\n      \"evidence\": \"Conditional knockout, unbiased Co-IP-MS, phospho-STAT5, migration and Rac assays\",\n      \"pmids\": [\"30413360\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Direct kinase substrate in STAT5 amplification not defined\", \"Mst1 versus Mst2 specific contributions unresolved\"]\n    },\n    {\n      \"year\": 2020,\n      \"claim\": \"Unified the activation mechanism by demonstrating that kinase-domain proximity alone—via homodimerization, membrane recruitment, or SAV1—is sufficient to trigger autophosphorylation, with no dedicated assembly required.\",\n      \"evidence\": \"Chemically induced dimerization, single-molecule pulldown, in vitro and cell-based autophosphorylation\",\n      \"pmids\": [\"32994222\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"How distinct upstream events select downstream output not addressed\", \"Endogenous trigger frequency in vivo unknown\"]\n    },\n    {\n      \"year\": 2021,\n      \"claim\": \"Established STK3 control of mitophagy via the receptor BNIP3 in adipocytes, defining a metabolic function in mitochondrial quality control.\",\n      \"evidence\": \"Conditional knockout mice, Co-IP, BNIP3 phosphorylation/dimerization and mitochondrial function assays\",\n      \"pmids\": [\"33758424\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Whether BNIP3 is a direct STK3 substrate not firmly established\", \"Link to canonical Hippo signaling in adipocytes unclear\"]\n    },\n    {\n      \"year\": 2022,\n      \"claim\": \"Identified opposing ubiquitin-system regulators—TRIM21 K63-ubiquitination (K473) activating, SCF\\u03b2TrCP degradation (ECM-stiffness-driven) inactivating—showing post-translational and mechanical control of STK3 abundance and activity.\",\n      \"evidence\": \"Co-IP, mutagenesis, in vitro ubiquitination, proteasome inhibition, organoid and MD simulation\",\n      \"pmids\": [\"37354905\", \"36044955\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Reciprocal validation of ubiquitin marks limited to single labs\", \"Interplay between the two ligases not tested\"]\n    },\n    {\n      \"year\": 2023,\n      \"claim\": \"Defined PRMT5 arginine methylation (R461/R467) of the SARAH domain as a homodimerization-blocking off-switch, and TRIM69 as a spatial regulator coupling STK3 to PLK1 at centrosomes, broadening regulatory and mitotic functions.\",\n      \"evidence\": \"In vitro methylation/kinase/dimerization assays, mutagenesis, Co-IP, localization, centrosome phenotype, in vivo xenograft\",\n      \"pmids\": [\"37905571\", \"37739411\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Whether methylation and phosphorylation compete on the same SARAH surface unclear\", \"TRIM69-S15/PLK1 axis confirmed by single lab only\"]\n    },\n    {\n      \"year\": 2024,\n      \"claim\": \"Added context-specific substrates—GSK-3\\u03b2 (activating Wnt/\\u03b2-catenin in a YAP1 feedback loop) and FOXO1 Ser212 (driving tumor-suppressive transcription)—illustrating STK3 outputs that diverge from canonical YAP suppression.\",\n      \"evidence\": \"Co-IP, in vitro kinase, ChIP, knockout mice, IF, xenograft assays\",\n      \"pmids\": [\"40604818\", \"38436783\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Apparent context-dependent oncogenic versus suppressive roles unreconciled\", \"Single-lab findings without independent replication\"]\n    },\n    {\n      \"year\": null,\n      \"claim\": \"How divergent upstream activating events and the diverse substrate repertoire are selected to produce specific cellular outputs (apoptosis, autophagy, migration, mitophagy, transcription) within a single cell remains unresolved.\",\n      \"evidence\": \"\",\n      \"pmids\": [],\n      \"confidence\": \"High\",\n      \"gaps\": [\"No unifying model linking activation context to output choice\", \"Quantitative integration of competing regulators not established\", \"Tissue-specific substrate selection mechanisms undefined\"]\n    }\n  ],\n  \"mechanism_profile\": {\n    \"molecular_activity\": [\n      {\"term_id\": \"GO:0140096\", \"supporting_discovery_ids\": [4, 7, 8, 9, 23, 25, 32, 39, 40]},\n      {\"term_id\": \"GO:0016740\", \"supporting_discovery_ids\": [2, 4, 7, 19, 33]},\n      {\"term_id\": \"GO:0042393\", \"supporting_discovery_ids\": [25]}\n    ],\n    \"localization\": [\n      {\"term_id\": \"GO:0005730\", \"supporting_discovery_ids\": [25]},\n      {\"term_id\": \"GO:0005829\", \"supporting_discovery_ids\": [12]},\n      {\"term_id\": \"GO:0005815\", \"supporting_discovery_ids\": [38]}\n    ],\n    \"pathway\": [\n      {\"term_id\": \"R-HSA-162582\", \"supporting_discovery_ids\": [4, 6, 7, 8, 11, 17]},\n      {\"term_id\": \"R-HSA-5357801\", \"supporting_discovery_ids\": [0, 3, 6, 17]},\n      {\"term_id\": \"R-HSA-9612973\", \"supporting_discovery_ids\": [23, 34]},\n      {\"term_id\": \"R-HSA-1640170\", \"supporting_discovery_ids\": [7, 10, 29, 38]},\n      {\"term_id\": \"R-HSA-168256\", \"supporting_discovery_ids\": [18, 26, 31]},\n      {\"term_id\": \"R-HSA-74160\", \"supporting_discovery_ids\": [25, 32, 40]}\n    ],\n    \"complexes\": [\n      \"MST2\\u2013SAV1 (hWW45/Salvador)\",\n      \"MST2\\u2013Raf-1 inhibitory complex\",\n      \"MOB1\\u2013MST2\\u2013NDR1 complex\"\n    ],\n    \"partners\": [\n      \"SAV1\",\n      \"RASSF1A\",\n      \"RAF1\",\n      \"LATS1\",\n      \"MOB1\",\n      \"RASSF5\",\n      \"AKT1\",\n      \"TRIM21\"\n    ],\n    \"other_free_text\": []\n  }\n}","audit_flag":null,"evaluation":{"pairwise":"win","faith_supported":8,"faith_total":8,"faith_pct":100.0}}