{"gene":"MAP4K4","run_date":"2026-06-10T02:59:50","timeline":{"discoveries":[{"year":1999,"finding":"MAP4K4 (HGK) is a serine/threonine kinase that specifically activates the JNK signaling pathway but not ERK or p38. HGK-induced JNK activation is inhibited by dominant-negative MKK4 and MKK7 mutants, and dominant-negative TAK1 (but not MEKK1 or MUK) strongly inhibits HGK-induced JNK activation, positioning HGK upstream of TAK1→MKK4/MKK7→JNK. TNF-α activates HGK in 293T cells and dominant-negative HGK mutants inhibit TNF-α-induced JNK activation.","method":"Transient transfection of 293T cells with wild-type and dominant-negative kinase constructs, kinase pathway specificity assays, epistasis with dominant-negative pathway components","journal":"The Journal of biological chemistry","confidence":"High","confidence_rationale":"Tier 2 / Strong — multiple orthogonal epistasis experiments, dominant-negative constructs for multiple pathway components, independently replicated concept in subsequent work","pmids":["9890973"],"is_preprint":false},{"year":2006,"finding":"MAP4K4 knockdown by siRNA inhibits migration and invasion of multiple carcinoma cell lines, and this pro-migratory effect is mediated through JNK signaling independently of AP-1 transcriptional activation.","method":"siRNA knockdown screen (10,996 siRNAs / 5,234 genes), 384-well wound-healing assay, validation in multiple cell lines, small-molecule JNK inhibition","journal":"Proceedings of the National Academy of Sciences of the United States of America","confidence":"High","confidence_rationale":"Tier 2 / Strong — genome-scale siRNA screen confirmed with multiple independent siRNAs, validated across multiple cell lines, pathway mechanism confirmed with small-molecule inhibitor","pmids":["16537454"],"is_preprint":false},{"year":2006,"finding":"MAP4K4/NIK is a negative regulator of PPARγ expression, adipogenesis, and insulin-stimulated glucose transport in adipocytes. MAP4K4 suppresses expression of C/EBPα, C/EBPβ, PPARγ, and GLUT4. TNF-α-induced GLUT4 downregulation requires MAP4K4 expression.","method":"RNAi screen of protein kinases in adipocytes, siRNA knockdown, gene expression analysis, glucose transport assays","journal":"Proceedings of the National Academy of Sciences of the United States of America","confidence":"High","confidence_rationale":"Tier 2 / Strong — functional RNAi screen with mechanistic follow-up across multiple readouts, replicated by multiple subsequent studies","pmids":["16461467"],"is_preprint":false},{"year":2007,"finding":"MAP4K4 mediates TNF-α-induced insulin resistance in human skeletal muscle by promoting JNK and ERK-1/2 phosphorylation and IRS-1 serine phosphorylation. Silencing MAP4K4 prevents TNF-α-induced inhibition of glucose uptake and rescues Akt and AS160 signaling. Silencing downstream MAP2K1 and MAP2K4 recapitulates MAP4K4 siRNA effects, placing MAP4K4 upstream of these kinases.","method":"siRNA knockdown in primary human skeletal muscle cells, glucose uptake assays, phosphorylation analysis by western blot, epistasis with MAP2K1/MAP2K4 siRNA","journal":"The Journal of biological chemistry","confidence":"High","confidence_rationale":"Tier 2 / Strong — primary human cells, multiple orthogonal readouts, epistasis experiments positioning MAP4K4 in pathway","pmids":["17227768"],"is_preprint":false},{"year":2007,"finding":"TNF-α selectively upregulates Map4k4 mRNA expression through TNFR1 (not TNFR2), and this requires the transcription factors c-Jun and ATF2. siRNA depletion of c-Jun and ATF2 attenuates TNF-α-induced Map4k4 expression.","method":"Agonistic/antagonistic antibodies and siRNA against TNFR1/TNFR2, siRNA depletion of c-Jun and ATF2, mRNA quantification in cultured adipocytes","journal":"The Journal of biological chemistry","confidence":"High","confidence_rationale":"Tier 2 / Strong — multiple independent siRNA and antibody approaches, receptor specificity clearly established with multiple orthogonal methods","pmids":["17500068"],"is_preprint":false},{"year":2009,"finding":"Map4k4 negatively regulates PPARγ at the post-transcriptional level by inhibiting mTOR signaling (reducing 4E-BP1 phosphorylation), thereby decreasing PPARγ protein translation rather than altering mRNA levels or PPARγ degradation. Map4k4 depletion requires mTOR and 4E-BP1 to increase PPARγ protein levels.","method":"siRNA knockdown, [35S]methionine/cysteine labeling to measure protein synthesis, cycloheximide chase for degradation measurement, adenovirus-mediated MAP4K4 overexpression, mTOR/4E-BP1 pathway analysis","journal":"The Journal of biological chemistry","confidence":"High","confidence_rationale":"Tier 1-2 / Strong — metabolic labeling, cycloheximide chase, gain- and loss-of-function, multiple orthogonal methods in single study","pmids":["20038583"],"is_preprint":false},{"year":2009,"finding":"Map4k4 in macrophages is a mediator of cytokine (TNF-α and IL-1β) expression. Silencing Map4k4 in macrophages in vivo protected mice from LPS-induced lethality by inhibiting TNF-α and IL-1β production.","method":"GeRP-mediated siRNA delivery to macrophages in vivo, LPS-induced lethality model, cytokine measurement","journal":"Nature","confidence":"High","confidence_rationale":"Tier 2 / Strong — in vivo siRNA delivery validated with multiple tissue macrophage populations, functional lethality endpoint, replicated by subsequent studies","pmids":["19407801"],"is_preprint":false},{"year":2012,"finding":"Map4k4 is a negative regulator of skeletal muscle differentiation. Its kinase activity is essential for this inhibitory function, as a kinase-inactive mutant enhances myotube formation. Map4k4 acts through upregulation of the myogenic factor Myf5: Map4k4 silencing upregulates Myf5, and Myf5 is required for the enhanced myogenesis seen with Map4k4 depletion. This mechanism is independent of ERK, p38, or JNK signaling.","method":"siRNA knockdown and adenovirus-mediated overexpression in C2C12 cells, kinase-inactive mutant expression, Myf5 rescue experiments, myogenic gene expression and myotube morphology analysis","journal":"Molecular and cellular biology","confidence":"High","confidence_rationale":"Tier 2 / Strong — bidirectional gain/loss of function, kinase-dead mutant, epistasis with Myf5, multiple orthogonal readouts in single study","pmids":["23207904"],"is_preprint":false},{"year":2013,"finding":"Map4k4 suppresses adipocyte lipogenesis by inhibiting mTORC1 via AMPK-mediated phosphorylation of raptor (Ser792), leading to suppression of Srebp-1 protein expression and cleavage. This mechanism is independent of JNK signaling: Map4k4 silencing enhances lipogenesis and Srebp-1 upregulation without affecting JNK, and Map4k4 overexpression fails to activate endogenous JNK.","method":"siRNA knockdown, full-genome microarray, 14C-glucose and 14C-acetate incorporation into TGs/fatty acids, Srebp-1 rescue experiments, rapamycin treatment, JNK depletion comparison","journal":"Journal of lipid research","confidence":"High","confidence_rationale":"Tier 1-2 / Strong — metabolic isotope labeling, multiple genetic perturbations, pharmacological inhibitor controls, mechanistic pathway dissection in single rigorous study","pmids":["23924694"],"is_preprint":false},{"year":2014,"finding":"MAP4K4 phosphorylates moesin in retracting membranes of migrating endothelial cells at sites of focal adhesion disassembly. Moesin functions downstream of MAP4K4 to inactivate β1-integrin by competing with talin for binding to the β1-integrin intracellular domain, thereby promoting membrane retraction and focal adhesion disassembly. Epistasis analyses established moesin downstream of MAP4K4; α5β1-integrin blockade reversed membrane retraction defects caused by MAP4K4 loss.","method":"siRNA and chemical inhibitor screens, phosphorylation analysis, epistasis analysis, α5β1-integrin blocking, in vitro and in vivo angiogenesis assays, inducible Map4k4 knockout mice","journal":"Nature","confidence":"High","confidence_rationale":"Tier 1-2 / Strong — multiple orthogonal methods (biochemical, genetic epistasis, in vivo KO), substrate phosphorylation demonstrated, competitive binding mechanism established","pmids":["25799996"],"is_preprint":false},{"year":2014,"finding":"MAP4K4 associates with microtubule plus-end binding protein EB2 and the Arf6 GEF IQSEC1. MAP4K4 is delivered to focal adhesions via EB2-bound microtubules, where it activates Arf6 through IQSEC1, promoting integrin internalization and focal adhesion disassembly. MAP4K4 knockout stabilizes focal adhesions and impairs cell migration.","method":"Quantitative proteomics (MAP4K4 FA-associated complex), knockout, co-immunoprecipitation, Arf6 activation assay, microtubule association studies","journal":"Developmental cell","confidence":"High","confidence_rationale":"Tier 1-2 / Strong — quantitative proteomics for complex identification, knockout phenotype, biochemical validation of IQSEC1 interaction and Arf6 activation","pmids":["25490267"],"is_preprint":false},{"year":2014,"finding":"HGK/MAP4K4 directly phosphorylates TRAF2, promoting its lysosomal degradation and thereby inhibiting IL-6 production in T cells. In T-cell-specific MAP4K4 knockout mice, TRAF2 is stabilized, IL-6 is overproduced, and this drives Th17 differentiation in adipose tissue leading to insulin resistance.","method":"T-cell-specific conditional knockout mice, direct phosphorylation assay, IL-6 and IL-17 neutralization, CCL20 neutralization, CCR6 knockout epistasis, leptin receptor knockout epistasis","journal":"Nature communications","confidence":"High","confidence_rationale":"Tier 2 / Strong — direct phosphorylation demonstrated, multiple genetic epistasis experiments in vivo, T-cell-specific conditional KO with multi-readout phenotype","pmids":["25098764"],"is_preprint":false},{"year":2015,"finding":"MAP4K4 directly phosphorylates and coimmunoprecipitates with FARP1 (FERM, RhoGEF, and pleckstrin domain-containing protein 1). MAP4K4 favors a pTL motif in vitro based on kinase specificity profiling. MAP4K4 inhibition in SH-SY5Y cells increases neurite outgrowth, a process known to involve FARP1.","method":"Phosphoproteomic analysis of MAP4K4 inhibition, in vitro kinase specificity assay, co-immunoprecipitation, direct phosphorylation assay, neurite outgrowth assay","journal":"ACS chemical biology","confidence":"High","confidence_rationale":"Tier 1 / Moderate — in vitro kinase assay with substrate identification, Co-IP, phosphoproteomics, functional cellular readout, single lab","pmids":["26422651"],"is_preprint":false},{"year":2015,"finding":"MAP4K4 is identified as the protein target of militarinone-inspired 4-hydroxy-2-pyridone neuritogenic compounds. The most potent compound is a selective ATP-competitive inhibitor of MAP4K4, confirmed by biochemical analysis and a crystal structure of the inhibitor in complex with MAP4K4.","method":"Target identification via activity-based profiling, biochemical kinase assays, X-ray crystallography of inhibitor-MAP4K4 complex","journal":"Angewandte Chemie (International ed. in English)","confidence":"High","confidence_rationale":"Tier 1 / Moderate — crystal structure with bound inhibitor, ATP-competitive mechanism confirmed biochemically","pmids":["25908259"],"is_preprint":false},{"year":2015,"finding":"MAP4K4 promotes vascular inflammation and atherosclerosis in endothelial cells by activating NF-κB (reducing its nuclear localization and activity when MAP4K4 is silenced), leading to decreased cell surface adhesion molecule expression and reduced macrophage recruitment.","method":"Endothelial-specific MAP4K4 siRNA and gene ablation in Apoe−/− mice, small-molecule MAP4K4 inhibitor in Apoe−/− and Ldlr−/− mice, NF-κB nuclear localization and activity assays in cultured ECs","journal":"Nature communications","confidence":"High","confidence_rationale":"Tier 2 / Strong — endothelial-specific genetic ablation in vivo, pharmacological inhibitor in two mouse models, NF-κB mechanistic readout in vitro","pmids":["26688060"],"is_preprint":false},{"year":2015,"finding":"Inducible whole-body Map4k4 deletion in obese mice improves insulin sensitivity in liver and adipose tissue but not skeletal muscle. Deletion of Map4k4 specifically in Myf5-positive tissues (skeletal muscle) is sufficient to protect from obesity-induced glucose intolerance, and also unexpectedly increases insulin sensitivity in adipose tissue.","method":"Tamoxifen-inducible whole-body Cre/ERT2 Map4k4 floxed knockout, tissue-specific conditional knockouts (adiponectin-Cre, albumin-Cre, Myf5-Cre), insulin signaling (AKT phosphorylation) in multiple tissues","journal":"Molecular and cellular biology","confidence":"High","confidence_rationale":"Tier 2 / Strong — multiple tissue-specific conditional knockout mouse models with metabolic phenotyping, distinguishes cell-autonomous from systemic effects","pmids":["25918248"],"is_preprint":false},{"year":2017,"finding":"MAP4K4, MINK1 (MAP4K6), and TNIK (MAP4K7) act redundantly upstream of DLK/JNK signaling in neurons. Targeting all three (but not any one individually) is sufficient to protect neurons from degeneration. MAP4K4/MINK1/TNIK pharmacological inhibition blocks DLK stabilization/phosphorylation within axons and prevents retrograde translocation of the JNK signaling complex to the nucleus.","method":"siRNA/pharmacological inhibition in mouse DRG neurons (trophic factor withdrawal model), DLK phosphorylation assays, retrograde signaling analysis, cell viability assays","journal":"The Journal of neuroscience","confidence":"High","confidence_rationale":"Tier 2 / Strong — genetic and pharmacological epistasis, redundancy dissected with combinatorial knockdown, mechanistic readout of DLK phosphorylation and retrograde transport","pmids":["28993483"],"is_preprint":false},{"year":2017,"finding":"Alternative splicing of MAP4K4 exon 16, regulated by SRSF3 in a sequence-dependent manner (with RBM4 as an upstream regulator of SRSF3 splicing), generates MAP4K4 variants with differential effects on JNK1 phosphorylation, which modulates E-cadherin, N-cadherin, and vimentin expression and alters migration/invasion of colorectal cancer cells.","method":"Transcriptome analysis of CRC tissues, SRSF3 splicing manipulation, MAP4K4 exon 16 inclusion/exclusion constructs, JNK1 phosphorylation assays, EMT marker expression, migration/invasion assays","journal":"Biochimica et biophysica acta. Molecular cell research","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — splice variant functional differences demonstrated with JNK1 phosphorylation readout, RBM4-SRSF3-MAP4K4 cascade established by splicing assays, single lab","pmids":["29138007"],"is_preprint":false},{"year":2017,"finding":"MAP4K4 is a novel positive regulator of MAPK/ERK signaling in lung adenocarcinoma that activates ERK through inhibiting protein phosphatase 2 (PP2A) activity.","method":"MAP4K4 knockdown in lung adenocarcinoma cells, PP2A activity assay, ERK phosphorylation analysis, xenograft growth and metastasis assays","journal":"Molecular oncology","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — PP2A activity directly measured, ERK phosphorylation as mechanistic readout, loss-of-function with multiple cellular phenotypes, single lab","pmids":["28306189"],"is_preprint":false},{"year":2019,"finding":"MAP4K4 is activated in failing human hearts and by oxidative stress. MAP4K4 is required for oxidative stress-induced cardiomyocyte death. Pharmacological inhibition with DMX-5804 rescues cell survival, mitochondrial function, and calcium cycling in hiPSC-CMs, and reduces ischemia-reperfusion injury in mice by >50%.","method":"MAP4K4 gene silencing in hiPSC-CMs, small-molecule inhibitor (DMX-5804), oxidative stress assays, mitochondrial function, calcium cycling measurement, mouse ischemia-reperfusion model","journal":"Cell stem cell","confidence":"High","confidence_rationale":"Tier 2 / Strong — genetic and pharmacological inhibition in human stem cell-derived cardiomyocytes and in vivo mouse model, multiple orthogonal functional readouts","pmids":["30853557"],"is_preprint":false},{"year":2019,"finding":"MAP4K4 activation regulates motor neuron degeneration in ALS via phosphorylated JNK3 and activation of the canonical c-Jun apoptotic pathway. MAP4K4 suppression promotes survival, prevents neurite degeneration, and decreases mutant SOD1 levels through autophagy (FoxO1-mediated) activation.","method":"MAP4K4 knockdown and pharmacological inhibition in ESC/iPSC-derived motor neurons and primary mouse spinal cord MNs, JNK3 phosphorylation assays, c-Jun pathway analysis, autophagy assays, SOD1 aggregate quantification","journal":"Cell reports","confidence":"High","confidence_rationale":"Tier 2 / Strong — multiple cell models (ESC, iPSC, primary), genetic and pharmacological loss-of-function, mechanistic pathway dissection with JNK3/c-Jun and autophagy readouts","pmids":["30699345"],"is_preprint":false},{"year":2020,"finding":"STRIPAK complex member STRN4 is required for SV40 Small T antigen (ST)-PP2A-induced cell transformation. ST recruits STRIPAK (via striatin B''' subunits), which directs PP2A-mediated dephosphorylation of MAP4K4, activating the Hippo pathway effector YAP1 and promoting oncogenic transformation.","method":"ST-PP2A interaction proteomics, STRN4 knockdown transformation assays, MAP4K4 phosphorylation analysis, YAP1 activity assays","journal":"eLife","confidence":"High","confidence_rationale":"Tier 1-2 / Strong — biochemical reconstitution of PP2A-STRIPAK-MAP4K4 axis, direct dephosphorylation demonstrated, genetic epistasis with STRN4 and YAP1","pmids":["31913126"],"is_preprint":false},{"year":2020,"finding":"MAP4K4 negatively regulates LFA-1 activation on CD8 T cells through ERM (ezrin, radixin, moesin) proteins, which serve as mediators between MAP4K4 and LFA-1. MAP4K4 deletion or inhibition enhances LFA-1 activation, improves T cell adherence to APCs, and increases CD8 T cell priming, proliferation, cytokine production, and cytotoxic activity. LFA-1 inhibition reverses these phenotypes.","method":"Genetic deletion (inducible whole-body Map4k4 KO), pharmacological inhibition, LFA-1 activation assays, T cell-APC conjugation assays, antitumor and antiviral in vivo models, LFA-1 inhibitor epistasis","journal":"Science immunology","confidence":"High","confidence_rationale":"Tier 2 / Strong — genetic and pharmacological evidence, LFA-1 inhibitor epistasis defining ERM as mediator, multiple functional in vivo readouts","pmids":["32220977"],"is_preprint":false},{"year":2021,"finding":"MAP4K4 directly phosphorylates MLK3 on Thr738, increasing MLK3 kinase activity and downstream signaling, thereby promoting pancreatic cancer cell proliferation, migration, and colony formation. MAP4K4 and MLK3 co-immunoprecipitate.","method":"Co-immunoprecipitation, direct phosphorylation assay (Thr738 site-specific), cell proliferation and migration assays, pharmacological MAP4K4 inhibitor (GNE-495) in KPC mouse model","journal":"Oncogene","confidence":"High","confidence_rationale":"Tier 1-2 / Strong — direct phosphorylation with site identification, Co-IP, gain-of-function rescue experiments, in vivo KPC mouse model validation","pmids":["34511598"],"is_preprint":false},{"year":2021,"finding":"MAP4K4 is regulated within STRIPAK complexes in cardiomyocytes. FLAG-MAP4K4 associates with all three striatins (STRN1, STRN3, STRN4) and with myosin. Calyculin A (Ser/Thr phosphatase inhibitor) induces MAP4K4 hyperphosphorylation of the activation loop and linker region in a kinase-activity-dependent manner. Surprisingly, FLAG-MAP4K4 inhibits JNK activation by H2O2 in cardiomyocytes and increases myofibrillar organization.","method":"5'-RACE and full-length cloning of rat Map4k4, FLAG-MAP4K4 co-immunoprecipitation with striatins and myosin, calyculin A treatment, phosphorylation mapping, kinase-dead mutant, JNK activation assay, myofibrillar staining","journal":"The Biochemical journal","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — biochemical Co-IP with all three striatins, activation loop phosphorylation demonstrated, kinase-dead control, unexpected finding (MAP4K4 inhibits JNK in this context) with appropriate caveats, single lab","pmids":["34032269"],"is_preprint":false},{"year":2022,"finding":"MAP4K4 interacts with striatin 3 (STRN3) via its CNH domain; STRN3 couples MAP4K4 to PP2A, which inactivates growth-repressing activities of MAP4K4. STRN3 also enables growth factor-induced PKCθ activation and direct phosphorylation of VASP-S157 by MAP4K4, which is required for efficient tumor cell invasion. Loss of STRN3 reactivates Hippo signaling.","method":"Co-immunoprecipitation (MAP4K4-STRN3/4), CNH domain deletion constructs, STRN3 and MAP4K4 depletion, VASP-S157 phosphorylation assay, PKCθ activation assay, Hippo pathway (YAP) activity, invasion assays in cerebellar tissue","journal":"Communications biology","confidence":"High","confidence_rationale":"Tier 1-2 / Strong — direct phosphorylation of VASP-S157 by MAP4K4 demonstrated, CNH domain interaction mapped, PP2A-STRIPAK regulation confirmed, multiple orthogonal functional assays","pmids":["35941177"],"is_preprint":false},{"year":2022,"finding":"RhoA counteracts abdominal aortic aneurysm formation by inhibiting MAP4K4 activity. In the absence of RhoA, the PP2A inhibitor Set sequesters PP2A, releasing MAP4K4 from inhibition and activating MAP kinase signaling including MAP4K4. MAP4K4 inhibitor (DMX-5804) decreases AAA formation in RhoA cKO mice.","method":"VSMC-specific RhoA conditional knockout mice, MAP4K4 activity measurement, Set-PP2A interaction assays, DMX-5804 pharmacological inhibition, aortic ring contractility, gene expression","journal":"Communications biology","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — genetic (RhoA cKO) and pharmacological (DMX-5804) evidence, Set-PP2A-MAP4K4 axis proposed with biochemical support, single lab","pmids":["36207400"],"is_preprint":false},{"year":2023,"finding":"MAP4K4 promotes ovarian cancer metastasis by phosphorylating ADAM10 at Ser436, which suppresses ADAM10-mediated cleavage of N-cadherin, leading to N-cadherin stabilization and enhanced cancer cell adhesion, migration, and invasion.","method":"MAP4K4 knockdown/overexpression, ADAM10 phosphorylation at Ser436 (site-specific), N-cadherin cleavage assays, cell adhesion/migration/invasion assays, pharmacological MAP4K4 inhibition in peritoneal metastasis model","journal":"Oncogene","confidence":"High","confidence_rationale":"Tier 1-2 / Strong — direct substrate phosphorylation with site identification, cleavage assay showing functional consequence, in vivo metastasis model","pmids":["36922678"],"is_preprint":false},{"year":2023,"finding":"MAP4K4 regulates forces at cell-cell and cell-substrate adhesions to promote collective cell migration. MAP4K4 promotes focal adhesion disassembly through moesin phosphorylation but disassembles adherens junctions through a moesin-independent mechanism. Loss of MAP4K4 increases traction forces and tension at cell-cell adhesions, causing tensional disequilibrium in migrating clusters.","method":"MAP4K4 inactivation and overexpression in A431 carcinoma cells, traction force microscopy, intercellular force measurement, moesin phosphorylation assay, focal adhesion and adherens junction dynamics","journal":"Life science alliance","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — biophysical force measurements, moesin phosphorylation mechanistic link established, but moesin-independent adherens junction mechanism not fully resolved, single lab","pmids":["37369604"],"is_preprint":false},{"year":2024,"finding":"MAP4K4 promotes S-nitrosylation of Drp1 (SNO-Drp1) at Cys644 (human C644/mouse C650) in cardiac microvascular endothelial cells by inhibiting GPX4 expression, which stimulates endothelial ferroptosis and cardiac microvascular dysfunction in diabetes. MAP4K4 inhibition (DMX-5804) reduces SNO-Drp1, alleviates microvascular dysfunction, and improves cardiac function in db/db mice; C650A Drp1 mutation abolishes SNO-Drp1 and its pathological effects.","method":"MAP4K4 overexpression/inhibition in CMECs, SNO-Drp1 site-specific analysis (C644/C650A mutation), GPX4 expression assay, ferroptosis assays, DMX-5804 in db/db mice, cardiac functional readouts","journal":"Cardiovascular diabetology","confidence":"High","confidence_rationale":"Tier 1-2 / Strong — site-specific Cys→Ala mutation abolishing SNO-Drp1, GPX4 mechanistic link, pharmacological and genetic validation in vivo, multiple orthogonal methods","pmids":["38724987"],"is_preprint":false},{"year":2018,"finding":"HGK (MAP4K4) activation by the diterpenoid Tanshinone IIA triggers JNK1-dependent c-Jun activation, leading to increased c-Jun recruitment to the AP-1-binding site in the sestrin 2 (SESN2) promoter and upregulation of SESN2 transcription. This SESN2-dependent (but Beclin 1-independent) autophagy mediates the antitumor effects of TIIA in osteosarcoma.","method":"ChIP assay (c-Jun at SESN2 promoter), HGK/JNK1/Jun pathway dissection, autophagy assays (SESN2-dependent vs Beclin 1-dependent), in vitro and in vivo (NOD/SCID) osteosarcoma models","journal":"Cell death & disease","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — ChIP confirms c-Jun at SESN2 promoter, SESN2 specificity established by depletion, single lab","pmids":["30258193"],"is_preprint":false},{"year":2016,"finding":"MAP4K4 promotes NFκB signaling and MAP4K4 silencing in hepatocellular carcinoma cells represses JNK, NFκB, and toll-like receptor signaling pathways. In vitro, MAP4K4 knockdown reduces proliferation, blocks cell cycle at S phase, and increases apoptosis.","method":"shRNA knockdown in HCC cell lines, quantitative real-time PCR arrays for signaling pathway analysis, cell cycle analysis, apoptosis assay, xenograft tumor growth","journal":"Clinical cancer research","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — shRNA KD with pathway-level PCR array readout, multiple cellular phenotype readouts, in vivo xenograft, but signaling pathway assignment is broad, single lab","pmids":["21196414"],"is_preprint":false},{"year":2013,"finding":"MAP4K4 is required for KSHV lytic reactivation from latency and promotes invasiveness of KSHV-infected endothelial cells by regulating expression of COX-2, MMP-7, and MMP-13.","method":"Small molecule library screening and siRNA silencing in KSHV-infected endothelial cells, COX-2/MMP expression assays, invasion assays","journal":"PLoS pathogens","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — siRNA and small-molecule screening convergent result, multiple downstream target analysis, single lab","pmids":["24244164"],"is_preprint":false},{"year":2013,"finding":"MAP4K4 interacts with Pyk2 via the Pyk2 FERM domain, co-immunoprecipitates with Pyk2 (but not FAK), and is a substrate for Pyk2. MAP4K4 knockdown inhibits glioma cell migration and blocks Pyk2 stimulation of cell migration; MAP4K4 overexpression-stimulated migration is blocked by Pyk2 knockdown, indicating mutual dependence.","method":"Yeast two-hybrid screen, co-immunoprecipitation, in vitro substrate assay (Pyk2 phosphorylates MAP4K4), siRNA knockdown, cell migration assays","journal":"Journal of signal transduction","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — yeast 2-hybrid validated by Co-IP, substrate relationship confirmed in vitro, bidirectional knockdown epistasis, single lab","pmids":["24163766"],"is_preprint":false},{"year":2021,"finding":"MAP4K4 directly controls integrin β1 activation and c-Met endocytosis in medulloblastoma cells, promoting receptor recycling that accumulates activated c-Met in cytosolic vesicles and sustains downstream signaling. MAP4K4 depletion restricts HGF-driven matrix invasion in vitro and brain tissue infiltration ex vivo.","method":"MAP4K4 siRNA depletion, HGF-stimulated invasion and 3D migration assays, integrin β1 activation assay, c-Met endocytosis and recycling measurement, ex vivo brain slice infiltration","journal":"Oncotarget","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — multiple functional assays, endocytic recycling mechanism directly measured, single lab","pmids":["29796184"],"is_preprint":false},{"year":2024,"finding":"Dipeptide transport by PEPT1 activates the MAP4K4/G3BP2 signaling pathway in hepatocellular carcinoma cells, leading to phosphorylation of G3BP2 at Thr227 by MAP4K4, which facilitates HCC metastasis.","method":"PEPT1 overexpression/knockdown, MAP4K4/G3BP2 pathway analysis, G3BP2-Thr227 phosphorylation assay, migration/invasion assays, lung metastasis mouse model","journal":"Advanced science","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — direct G3BP2 phosphorylation at Thr227 demonstrated, in vivo metastasis model, dipeptide-to-kinase activation link established, single lab","pmids":["38639383"],"is_preprint":false}],"current_model":"MAP4K4 is a Ste20-family serine/threonine kinase that functions as a multi-context stress and inflammatory signaling hub: it activates JNK via a TAK1→MKK4/MKK7 cascade downstream of TNF-α/TNFR1, phosphorylates moesin to promote focal adhesion disassembly and integrin inactivation during cell migration (in a MAP4K4→moesin→talin/β1-integrin pathway), delivers to focal adhesions via EB2-microtubule tracks to activate Arf6 through IQSEC1, directly phosphorylates substrates including TRAF2 (promoting lysosomal degradation), MLK3-Thr738, ADAM10-Ser436, VASP-Ser157, FARP1, and Drp1 at Cys644 (facilitating S-nitrosylation via GPX4 suppression), inhibits mTORC1/AMPK signaling to suppress PPARγ translation and adipocyte lipogenesis, is regulated by STRIPAK complexes (striatins tether PP2A to keep MAP4K4 dephosphorylated/inactive), and acts redundantly with MINK1/TNIK upstream of DLK to drive retrograde JNK stress signaling in neurons."},"narrative":{"mechanistic_narrative":"MAP4K4 is a Ste20-family serine/threonine kinase that functions as a stress- and inflammation-responsive signaling hub coupling extracellular cues to JNK-pathway activation, cytoskeletal remodeling, and metabolic control [PMID:9890973, PMID:19407801]. Classically, MAP4K4 acts upstream of a TAK1→MKK4/MKK7→JNK cascade and is itself activated by TNF-α through TNFR1, with TNF-α also transcriptionally inducing Map4k4 via c-Jun and ATF2 to create a feed-forward inflammatory loop [PMID:9890973, PMID:17500068]. In cell migration, MAP4K4 directly phosphorylates moesin at sites of focal adhesion disassembly, where moesin competes with talin to inactivate β1-integrin and promote membrane retraction; MAP4K4 is delivered to focal adhesions along EB2-bound microtubules to activate Arf6 through IQSEC1, driving integrin internalization and adhesion turnover [PMID:25799996, PMID:25490267, PMID:37369604]. Beyond moesin, MAP4K4 directly phosphorylates a defined set of substrates to control distinct outputs: TRAF2 (promoting its lysosomal degradation to limit IL-6) [PMID:25098764], MLK3 at Thr738 [PMID:34511598], ADAM10 at Ser436 (stabilizing N-cadherin) [PMID:36922678], VASP at Ser157 [PMID:35941177], FARP1 [PMID:26422651], and Drp1 (facilitating its S-nitrosylation at Cys644 via GPX4 suppression to drive endothelial ferroptosis) [PMID:38724987]. Metabolically, MAP4K4 suppresses adipogenesis and lipogenesis by inhibiting mTOR/mTORC1—reducing PPARγ protein translation and, via AMPK-mediated raptor phosphorylation, suppressing SREBP-1—and mediates TNF-α-induced insulin resistance [PMID:16461467, PMID:20038583, PMID:23924694, PMID:25918248]. MAP4K4 activity is restrained by STRIPAK: striatins (STRN1/3/4) tether PP2A to keep MAP4K4 dephosphorylated and inactive, linking it to Hippo/YAP and growth control [PMID:31913126, PMID:35941177]. Functionally, MAP4K4 promotes neuronal degeneration acting redundantly with MINK1 and TNIK upstream of DLK-driven retrograde JNK stress signaling [PMID:28993483, PMID:30699345], drives oxidative-stress cardiomyocyte death and ischemia-reperfusion injury [PMID:30853557], and restrains CD8 T-cell LFA-1 activation through ERM proteins [PMID:32220977].","teleology":[{"year":1999,"claim":"Established MAP4K4's core signaling identity by placing it as a JNK-selective upstream kinase responsive to TNF-α, defining the pathway architecture all later work builds on.","evidence":"Dominant-negative epistasis with TAK1/MKK4/MKK7 and TNF-α stimulation in 293T cells","pmids":["9890973"],"confidence":"High","gaps":["Did not identify direct physiological substrates","Mechanism of TNF-α-induced MAP4K4 activation left unresolved"]},{"year":2006,"claim":"Linked MAP4K4 to physiological outputs—cancer cell migration via JNK and metabolic regulation of adipogenesis/glucose transport—revealing it as more than a stress kinase.","evidence":"Genome-scale siRNA wound-healing screen across carcinoma lines, and RNAi kinase screen in adipocytes with glucose transport readouts","pmids":["16537454","16461467"],"confidence":"High","gaps":["Migration mechanism downstream of JNK undefined","Direct adipocyte substrates not identified"]},{"year":2007,"claim":"Defined a feed-forward inflammatory circuit and the insulin-resistance mechanism: TNFR1 induces Map4k4 transcription via c-Jun/ATF2, and MAP4K4 mediates TNF-α-driven IRS-1 serine phosphorylation.","evidence":"Receptor-specific antibodies/siRNA, c-Jun/ATF2 depletion, and epistasis with MAP2K1/MAP2K4 in primary human muscle","pmids":["17500068","17227768"],"confidence":"High","gaps":["Whether MAP4K4 directly phosphorylates IRS-1 not established","In vivo relevance to systemic insulin resistance not yet tested"]},{"year":2009,"claim":"Resolved the mechanism of metabolic suppression: MAP4K4 inhibits mTOR signaling to reduce PPARγ translation and acts in macrophages to drive cytokine production, with systemic protective effects.","evidence":"Metabolic labeling/cycloheximide chase with mTOR-4E-BP1 analysis in adipocytes, and in vivo macrophage siRNA in an LPS lethality model","pmids":["20038583","19407801"],"confidence":"High","gaps":["How MAP4K4 inhibits mTOR mechanistically not fully defined","Direct kinase substrate in the mTOR axis unknown"]},{"year":2012,"claim":"Showed MAP4K4 kinase activity inhibits myogenesis via Myf5, expanding its developmental roles beyond canonical MAPK cascades.","evidence":"Bidirectional gain/loss-of-function and kinase-dead mutant with Myf5 rescue in C2C12 cells","pmids":["23207904"],"confidence":"High","gaps":["Direct substrate linking MAP4K4 to Myf5 unidentified","ERK/p38/JNK-independent mechanism not molecularly defined"]},{"year":2013,"claim":"Connected MAP4K4 to adhesion-receptor signaling through Pyk2 interaction and demonstrated roles in viral (KSHV) reactivation and invasion.","evidence":"Yeast two-hybrid/Co-IP with Pyk2 FERM domain plus migration epistasis, and siRNA/small-molecule screening in KSHV-infected endothelial cells","pmids":["24163766","24244164"],"confidence":"Medium","gaps":["Pyk2-MAP4K4 phosphorylation directionality and sites incompletely mapped","KSHV regulation is single-lab, mechanism upstream of COX-2/MMPs unclear"]},{"year":2013,"claim":"Dissected the mTORC1/AMPK arm of metabolic control, showing MAP4K4 suppresses lipogenesis via AMPK-raptor-Ser792 and SREBP-1 independently of JNK.","evidence":"Isotope incorporation assays, genetic perturbation, and rapamycin/JNK-depletion controls in adipocytes","pmids":["23924694"],"confidence":"High","gaps":["Whether MAP4K4 directly phosphorylates AMPK not shown","Tissue-level lipogenic relevance not yet tested in vivo"]},{"year":2014,"claim":"Defined the direct cytoskeletal mechanism of migration: MAP4K4 phosphorylates moesin to inactivate β1-integrin and is trafficked to focal adhesions via EB2 microtubules to activate Arf6 through IQSEC1.","evidence":"Substrate phosphorylation, genetic epistasis, integrin blockade, inducible KO mice, and FA-complex proteomics with Arf6 activation assays","pmids":["25799996","25490267"],"confidence":"High","gaps":["How MAP4K4 selects moesin among ERM proteins at adhesions unclear","Coordination of microtubule delivery with kinase activation not resolved"]},{"year":2014,"claim":"Identified TRAF2 as a direct degradative substrate linking MAP4K4 to immune cytokine control, with in vivo consequences for adipose inflammation and insulin resistance.","evidence":"Direct phosphorylation assay and T-cell-specific conditional KO with multiple cytokine/genetic epistasis readouts","pmids":["25098764"],"confidence":"High","gaps":["TRAF2 phosphosite(s) and lysosomal targeting mechanism not fully mapped"]},{"year":2015,"claim":"Broadened the direct substrate catalog (FARP1) and provided structural/chemical tools, while clarifying tissue-specific metabolic genetics in vivo.","evidence":"Phosphoproteomics/in vitro kinase profiling for FARP1, X-ray crystallography of inhibitor-bound MAP4K4, and tissue-specific conditional KO metabolic phenotyping","pmids":["26422651","25908259","25918248"],"confidence":"High","gaps":["FARP1 phosphosite physiological role undefined","Why Myf5-tissue deletion improves adipose insulin sensitivity unexplained"]},{"year":2015,"claim":"Extended MAP4K4 to vascular inflammation through NF-κB regulation in endothelium, with atheroprotection upon inhibition.","evidence":"Endothelial-specific gene ablation and small-molecule inhibition in Apoe-/- and Ldlr-/- mice with NF-κB readouts","pmids":["26688060"],"confidence":"High","gaps":["Direct MAP4K4 substrate in the NF-κB pathway not identified"]},{"year":2016,"claim":"Implicated MAP4K4 in hepatocellular carcinoma proliferation and survival through broad JNK/NF-κB/TLR pathway engagement.","evidence":"shRNA knockdown with PCR-array pathway profiling, cell cycle/apoptosis assays, and xenografts","pmids":["21196414"],"confidence":"Medium","gaps":["Signaling assignment broad and correlative","No direct substrate identified, single-lab"]},{"year":2017,"claim":"Established neuronal degeneration roles: MAP4K4 acts redundantly with MINK1/TNIK upstream of DLK-driven retrograde JNK stress signaling.","evidence":"Combinatorial siRNA/pharmacological inhibition in DRG neurons with DLK phosphorylation and retrograde-transport readouts","pmids":["28993483"],"confidence":"High","gaps":["Whether MAP4K4 directly phosphorylates DLK not established","Relative contribution of each MAP4K family member in vivo unresolved"]},{"year":2017,"claim":"Revealed regulatory complexity: alternative splicing of exon 16 (SRSF3/RBM4) tunes JNK1 signaling and EMT, and MAP4K4 can positively regulate ERK by inhibiting PP2A in lung cancer.","evidence":"Splice-variant constructs with JNK1/EMT readouts, and knockdown with PP2A activity/ERK phosphorylation assays plus xenografts","pmids":["29138007","28306189"],"confidence":"Medium","gaps":["Functional difference between splice isoforms mechanistically incomplete","How MAP4K4 inhibits PP2A unclear, both single-lab"]},{"year":2018,"claim":"Linked MAP4K4-JNK1-c-Jun signaling to SESN2-dependent autophagy as an antitumor mechanism, and showed MAP4K4 controls c-Met recycling/integrin β1 in medulloblastoma invasion.","evidence":"ChIP of c-Jun at the SESN2 promoter with autophagy assays in osteosarcoma, and endocytosis/recycling measurements in medulloblastoma","pmids":["30258193","29796184"],"confidence":"Medium","gaps":["Pharmacological MAP4K4 activation specificity not fully controlled","Direct substrate controlling c-Met endocytosis unidentified"]},{"year":2019,"claim":"Demonstrated MAP4K4 as a driver of oxidative-stress cardiomyocyte death and motor-neuron degeneration, nominating it as a therapeutic target in heart and ALS contexts.","evidence":"Genetic/pharmacological inhibition (DMX-5804) in hiPSC-CMs and mouse ischemia-reperfusion; knockdown/inhibition in ESC/iPSC/primary motor neurons with JNK3/c-Jun and autophagy readouts","pmids":["30853557","30699345"],"confidence":"High","gaps":["Direct substrate mediating cardiomyocyte death not identified","Link between MAP4K4 and FoxO1-mediated autophagy mechanistically incomplete"]},{"year":2020,"claim":"Defined STRIPAK-PP2A as the inhibitory regulator that keeps MAP4K4 dephosphorylated, and showed MAP4K4 negatively regulates CD8 T-cell LFA-1 activation via ERM proteins.","evidence":"ST-PP2A proteomics with STRN4 knockdown/dephosphorylation/YAP1 assays, and genetic/pharmacological MAP4K4 perturbation with LFA-1 epistasis in tumor/viral models","pmids":["31913126","32220977"],"confidence":"High","gaps":["Which MAP4K4 substrate(s) gate Hippo/YAP downstream of STRIPAK not fully defined"]},{"year":2021,"claim":"Expanded the direct substrate repertoire (MLK3-Thr738) in pancreatic cancer and mapped CNH-domain-dependent STRN3 binding coupling MAP4K4 to PP2A and VASP-S157 phosphorylation.","evidence":"Co-IP, site-specific phosphorylation assays, CNH-deletion constructs, and in vivo KPC model","pmids":["34511598","34032269"],"confidence":"High","gaps":["How STRN3 binding switches between inhibition and substrate selection unresolved","Context-dependent MAP4K4 inhibition of JNK in cardiomyocytes (single-lab) not generalized"]},{"year":2022,"claim":"Clarified STRN3-PP2A control of MAP4K4 growth-repressing activity and VASP-driven invasion, and positioned RhoA-Set-PP2A as an upstream restraint relevant to aortic aneurysm.","evidence":"Co-IP/CNH-domain mapping with VASP-S157 and Hippo assays, and VSMC-specific RhoA cKO with Set-PP2A analysis and DMX-5804","pmids":["35941177","36207400"],"confidence":"Medium","gaps":["Set-PP2A-MAP4K4 axis is proposed with limited reconstitution","RhoA-to-MAP4K4 link is single-lab"]},{"year":2023,"claim":"Added ADAM10-Ser436 as a direct substrate stabilizing N-cadherin in ovarian cancer, and resolved that MAP4K4 disassembles adherens junctions by a moesin-independent route in collective migration.","evidence":"Site-specific ADAM10 phosphorylation/cleavage assays with in vivo metastasis, and traction-force microscopy with moesin phosphorylation in A431 cells","pmids":["36922678","37369604"],"confidence":"Medium","gaps":["Moesin-independent adherens-junction mechanism not molecularly defined","ADAM10 finding single-lab"]},{"year":2024,"claim":"Identified Drp1-Cys644 S-nitrosylation (via GPX4 suppression) as a MAP4K4-driven ferroptotic mechanism in diabetic cardiac microvasculature, and PEPT1-dipeptide activation of MAP4K4-G3BP2-Thr227 in HCC metastasis.","evidence":"Site-specific Cys-to-Ala mutation with GPX4/ferroptosis assays and DMX-5804 in db/db mice; PEPT1 perturbation with G3BP2-Thr227 phosphorylation and lung metastasis model","pmids":["38724987","38639383"],"confidence":"High","gaps":["How MAP4K4 suppresses GPX4 expression mechanistically unclear","G3BP2-Thr227 phosphorylation is single-lab"]},{"year":null,"claim":"It remains unresolved how a single kinase integrates its many context-specific substrates and opposite outputs (e.g., JNK activation vs inhibition; growth promotion vs repression), and what governs substrate selection downstream of STRIPAK-PP2A regulation.","evidence":"","pmids":[],"confidence":"Medium","gaps":["No unifying structural/regulatory model for substrate selection","Context-dependent reversal of JNK output unexplained","Endogenous activating signal that overrides STRIPAK inhibition undefined"]}],"mechanism_profile":{"molecular_activity":[{"term_id":"GO:0140096","term_label":"catalytic activity, acting on a protein","supporting_discovery_ids":[9,11,12,23,25,27,29,35]},{"term_id":"GO:0016740","term_label":"transferase activity","supporting_discovery_ids":[0,9,23,27]},{"term_id":"GO:0140110","term_label":"transcription regulator 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HGK-induced JNK activation is inhibited by dominant-negative MKK4 and MKK7 mutants, and dominant-negative TAK1 (but not MEKK1 or MUK) strongly inhibits HGK-induced JNK activation, positioning HGK upstream of TAK1→MKK4/MKK7→JNK. TNF-α activates HGK in 293T cells and dominant-negative HGK mutants inhibit TNF-α-induced JNK activation.\",\n      \"method\": \"Transient transfection of 293T cells with wild-type and dominant-negative kinase constructs, kinase pathway specificity assays, epistasis with dominant-negative pathway components\",\n      \"journal\": \"The Journal of biological chemistry\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — multiple orthogonal epistasis experiments, dominant-negative constructs for multiple pathway components, independently replicated concept in subsequent work\",\n      \"pmids\": [\"9890973\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2006,\n      \"finding\": \"MAP4K4 knockdown by siRNA inhibits migration and invasion of multiple carcinoma cell lines, and this pro-migratory effect is mediated through JNK signaling independently of AP-1 transcriptional activation.\",\n      \"method\": \"siRNA knockdown screen (10,996 siRNAs / 5,234 genes), 384-well wound-healing assay, validation in multiple cell lines, small-molecule JNK inhibition\",\n      \"journal\": \"Proceedings of the National Academy of Sciences of the United States of America\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — genome-scale siRNA screen confirmed with multiple independent siRNAs, validated across multiple cell lines, pathway mechanism confirmed with small-molecule inhibitor\",\n      \"pmids\": [\"16537454\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2006,\n      \"finding\": \"MAP4K4/NIK is a negative regulator of PPARγ expression, adipogenesis, and insulin-stimulated glucose transport in adipocytes. MAP4K4 suppresses expression of C/EBPα, C/EBPβ, PPARγ, and GLUT4. TNF-α-induced GLUT4 downregulation requires MAP4K4 expression.\",\n      \"method\": \"RNAi screen of protein kinases in adipocytes, siRNA knockdown, gene expression analysis, glucose transport assays\",\n      \"journal\": \"Proceedings of the National Academy of Sciences of the United States of America\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — functional RNAi screen with mechanistic follow-up across multiple readouts, replicated by multiple subsequent studies\",\n      \"pmids\": [\"16461467\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2007,\n      \"finding\": \"MAP4K4 mediates TNF-α-induced insulin resistance in human skeletal muscle by promoting JNK and ERK-1/2 phosphorylation and IRS-1 serine phosphorylation. Silencing MAP4K4 prevents TNF-α-induced inhibition of glucose uptake and rescues Akt and AS160 signaling. Silencing downstream MAP2K1 and MAP2K4 recapitulates MAP4K4 siRNA effects, placing MAP4K4 upstream of these kinases.\",\n      \"method\": \"siRNA knockdown in primary human skeletal muscle cells, glucose uptake assays, phosphorylation analysis by western blot, epistasis with MAP2K1/MAP2K4 siRNA\",\n      \"journal\": \"The Journal of biological chemistry\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — primary human cells, multiple orthogonal readouts, epistasis experiments positioning MAP4K4 in pathway\",\n      \"pmids\": [\"17227768\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2007,\n      \"finding\": \"TNF-α selectively upregulates Map4k4 mRNA expression through TNFR1 (not TNFR2), and this requires the transcription factors c-Jun and ATF2. siRNA depletion of c-Jun and ATF2 attenuates TNF-α-induced Map4k4 expression.\",\n      \"method\": \"Agonistic/antagonistic antibodies and siRNA against TNFR1/TNFR2, siRNA depletion of c-Jun and ATF2, mRNA quantification in cultured adipocytes\",\n      \"journal\": \"The Journal of biological chemistry\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — multiple independent siRNA and antibody approaches, receptor specificity clearly established with multiple orthogonal methods\",\n      \"pmids\": [\"17500068\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2009,\n      \"finding\": \"Map4k4 negatively regulates PPARγ at the post-transcriptional level by inhibiting mTOR signaling (reducing 4E-BP1 phosphorylation), thereby decreasing PPARγ protein translation rather than altering mRNA levels or PPARγ degradation. Map4k4 depletion requires mTOR and 4E-BP1 to increase PPARγ protein levels.\",\n      \"method\": \"siRNA knockdown, [35S]methionine/cysteine labeling to measure protein synthesis, cycloheximide chase for degradation measurement, adenovirus-mediated MAP4K4 overexpression, mTOR/4E-BP1 pathway analysis\",\n      \"journal\": \"The Journal of biological chemistry\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1-2 / Strong — metabolic labeling, cycloheximide chase, gain- and loss-of-function, multiple orthogonal methods in single study\",\n      \"pmids\": [\"20038583\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2009,\n      \"finding\": \"Map4k4 in macrophages is a mediator of cytokine (TNF-α and IL-1β) expression. Silencing Map4k4 in macrophages in vivo protected mice from LPS-induced lethality by inhibiting TNF-α and IL-1β production.\",\n      \"method\": \"GeRP-mediated siRNA delivery to macrophages in vivo, LPS-induced lethality model, cytokine measurement\",\n      \"journal\": \"Nature\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — in vivo siRNA delivery validated with multiple tissue macrophage populations, functional lethality endpoint, replicated by subsequent studies\",\n      \"pmids\": [\"19407801\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2012,\n      \"finding\": \"Map4k4 is a negative regulator of skeletal muscle differentiation. Its kinase activity is essential for this inhibitory function, as a kinase-inactive mutant enhances myotube formation. Map4k4 acts through upregulation of the myogenic factor Myf5: Map4k4 silencing upregulates Myf5, and Myf5 is required for the enhanced myogenesis seen with Map4k4 depletion. This mechanism is independent of ERK, p38, or JNK signaling.\",\n      \"method\": \"siRNA knockdown and adenovirus-mediated overexpression in C2C12 cells, kinase-inactive mutant expression, Myf5 rescue experiments, myogenic gene expression and myotube morphology analysis\",\n      \"journal\": \"Molecular and cellular biology\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — bidirectional gain/loss of function, kinase-dead mutant, epistasis with Myf5, multiple orthogonal readouts in single study\",\n      \"pmids\": [\"23207904\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2013,\n      \"finding\": \"Map4k4 suppresses adipocyte lipogenesis by inhibiting mTORC1 via AMPK-mediated phosphorylation of raptor (Ser792), leading to suppression of Srebp-1 protein expression and cleavage. This mechanism is independent of JNK signaling: Map4k4 silencing enhances lipogenesis and Srebp-1 upregulation without affecting JNK, and Map4k4 overexpression fails to activate endogenous JNK.\",\n      \"method\": \"siRNA knockdown, full-genome microarray, 14C-glucose and 14C-acetate incorporation into TGs/fatty acids, Srebp-1 rescue experiments, rapamycin treatment, JNK depletion comparison\",\n      \"journal\": \"Journal of lipid research\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1-2 / Strong — metabolic isotope labeling, multiple genetic perturbations, pharmacological inhibitor controls, mechanistic pathway dissection in single rigorous study\",\n      \"pmids\": [\"23924694\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2014,\n      \"finding\": \"MAP4K4 phosphorylates moesin in retracting membranes of migrating endothelial cells at sites of focal adhesion disassembly. Moesin functions downstream of MAP4K4 to inactivate β1-integrin by competing with talin for binding to the β1-integrin intracellular domain, thereby promoting membrane retraction and focal adhesion disassembly. Epistasis analyses established moesin downstream of MAP4K4; α5β1-integrin blockade reversed membrane retraction defects caused by MAP4K4 loss.\",\n      \"method\": \"siRNA and chemical inhibitor screens, phosphorylation analysis, epistasis analysis, α5β1-integrin blocking, in vitro and in vivo angiogenesis assays, inducible Map4k4 knockout mice\",\n      \"journal\": \"Nature\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1-2 / Strong — multiple orthogonal methods (biochemical, genetic epistasis, in vivo KO), substrate phosphorylation demonstrated, competitive binding mechanism established\",\n      \"pmids\": [\"25799996\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2014,\n      \"finding\": \"MAP4K4 associates with microtubule plus-end binding protein EB2 and the Arf6 GEF IQSEC1. MAP4K4 is delivered to focal adhesions via EB2-bound microtubules, where it activates Arf6 through IQSEC1, promoting integrin internalization and focal adhesion disassembly. MAP4K4 knockout stabilizes focal adhesions and impairs cell migration.\",\n      \"method\": \"Quantitative proteomics (MAP4K4 FA-associated complex), knockout, co-immunoprecipitation, Arf6 activation assay, microtubule association studies\",\n      \"journal\": \"Developmental cell\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1-2 / Strong — quantitative proteomics for complex identification, knockout phenotype, biochemical validation of IQSEC1 interaction and Arf6 activation\",\n      \"pmids\": [\"25490267\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2014,\n      \"finding\": \"HGK/MAP4K4 directly phosphorylates TRAF2, promoting its lysosomal degradation and thereby inhibiting IL-6 production in T cells. In T-cell-specific MAP4K4 knockout mice, TRAF2 is stabilized, IL-6 is overproduced, and this drives Th17 differentiation in adipose tissue leading to insulin resistance.\",\n      \"method\": \"T-cell-specific conditional knockout mice, direct phosphorylation assay, IL-6 and IL-17 neutralization, CCL20 neutralization, CCR6 knockout epistasis, leptin receptor knockout epistasis\",\n      \"journal\": \"Nature communications\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — direct phosphorylation demonstrated, multiple genetic epistasis experiments in vivo, T-cell-specific conditional KO with multi-readout phenotype\",\n      \"pmids\": [\"25098764\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2015,\n      \"finding\": \"MAP4K4 directly phosphorylates and coimmunoprecipitates with FARP1 (FERM, RhoGEF, and pleckstrin domain-containing protein 1). MAP4K4 favors a pTL motif in vitro based on kinase specificity profiling. MAP4K4 inhibition in SH-SY5Y cells increases neurite outgrowth, a process known to involve FARP1.\",\n      \"method\": \"Phosphoproteomic analysis of MAP4K4 inhibition, in vitro kinase specificity assay, co-immunoprecipitation, direct phosphorylation assay, neurite outgrowth assay\",\n      \"journal\": \"ACS chemical biology\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 / Moderate — in vitro kinase assay with substrate identification, Co-IP, phosphoproteomics, functional cellular readout, single lab\",\n      \"pmids\": [\"26422651\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2015,\n      \"finding\": \"MAP4K4 is identified as the protein target of militarinone-inspired 4-hydroxy-2-pyridone neuritogenic compounds. The most potent compound is a selective ATP-competitive inhibitor of MAP4K4, confirmed by biochemical analysis and a crystal structure of the inhibitor in complex with MAP4K4.\",\n      \"method\": \"Target identification via activity-based profiling, biochemical kinase assays, X-ray crystallography of inhibitor-MAP4K4 complex\",\n      \"journal\": \"Angewandte Chemie (International ed. in English)\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 / Moderate — crystal structure with bound inhibitor, ATP-competitive mechanism confirmed biochemically\",\n      \"pmids\": [\"25908259\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2015,\n      \"finding\": \"MAP4K4 promotes vascular inflammation and atherosclerosis in endothelial cells by activating NF-κB (reducing its nuclear localization and activity when MAP4K4 is silenced), leading to decreased cell surface adhesion molecule expression and reduced macrophage recruitment.\",\n      \"method\": \"Endothelial-specific MAP4K4 siRNA and gene ablation in Apoe−/− mice, small-molecule MAP4K4 inhibitor in Apoe−/− and Ldlr−/− mice, NF-κB nuclear localization and activity assays in cultured ECs\",\n      \"journal\": \"Nature communications\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — endothelial-specific genetic ablation in vivo, pharmacological inhibitor in two mouse models, NF-κB mechanistic readout in vitro\",\n      \"pmids\": [\"26688060\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2015,\n      \"finding\": \"Inducible whole-body Map4k4 deletion in obese mice improves insulin sensitivity in liver and adipose tissue but not skeletal muscle. Deletion of Map4k4 specifically in Myf5-positive tissues (skeletal muscle) is sufficient to protect from obesity-induced glucose intolerance, and also unexpectedly increases insulin sensitivity in adipose tissue.\",\n      \"method\": \"Tamoxifen-inducible whole-body Cre/ERT2 Map4k4 floxed knockout, tissue-specific conditional knockouts (adiponectin-Cre, albumin-Cre, Myf5-Cre), insulin signaling (AKT phosphorylation) in multiple tissues\",\n      \"journal\": \"Molecular and cellular biology\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — multiple tissue-specific conditional knockout mouse models with metabolic phenotyping, distinguishes cell-autonomous from systemic effects\",\n      \"pmids\": [\"25918248\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2017,\n      \"finding\": \"MAP4K4, MINK1 (MAP4K6), and TNIK (MAP4K7) act redundantly upstream of DLK/JNK signaling in neurons. Targeting all three (but not any one individually) is sufficient to protect neurons from degeneration. MAP4K4/MINK1/TNIK pharmacological inhibition blocks DLK stabilization/phosphorylation within axons and prevents retrograde translocation of the JNK signaling complex to the nucleus.\",\n      \"method\": \"siRNA/pharmacological inhibition in mouse DRG neurons (trophic factor withdrawal model), DLK phosphorylation assays, retrograde signaling analysis, cell viability assays\",\n      \"journal\": \"The Journal of neuroscience\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — genetic and pharmacological epistasis, redundancy dissected with combinatorial knockdown, mechanistic readout of DLK phosphorylation and retrograde transport\",\n      \"pmids\": [\"28993483\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2017,\n      \"finding\": \"Alternative splicing of MAP4K4 exon 16, regulated by SRSF3 in a sequence-dependent manner (with RBM4 as an upstream regulator of SRSF3 splicing), generates MAP4K4 variants with differential effects on JNK1 phosphorylation, which modulates E-cadherin, N-cadherin, and vimentin expression and alters migration/invasion of colorectal cancer cells.\",\n      \"method\": \"Transcriptome analysis of CRC tissues, SRSF3 splicing manipulation, MAP4K4 exon 16 inclusion/exclusion constructs, JNK1 phosphorylation assays, EMT marker expression, migration/invasion assays\",\n      \"journal\": \"Biochimica et biophysica acta. Molecular cell research\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — splice variant functional differences demonstrated with JNK1 phosphorylation readout, RBM4-SRSF3-MAP4K4 cascade established by splicing assays, single lab\",\n      \"pmids\": [\"29138007\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2017,\n      \"finding\": \"MAP4K4 is a novel positive regulator of MAPK/ERK signaling in lung adenocarcinoma that activates ERK through inhibiting protein phosphatase 2 (PP2A) activity.\",\n      \"method\": \"MAP4K4 knockdown in lung adenocarcinoma cells, PP2A activity assay, ERK phosphorylation analysis, xenograft growth and metastasis assays\",\n      \"journal\": \"Molecular oncology\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — PP2A activity directly measured, ERK phosphorylation as mechanistic readout, loss-of-function with multiple cellular phenotypes, single lab\",\n      \"pmids\": [\"28306189\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2019,\n      \"finding\": \"MAP4K4 is activated in failing human hearts and by oxidative stress. MAP4K4 is required for oxidative stress-induced cardiomyocyte death. Pharmacological inhibition with DMX-5804 rescues cell survival, mitochondrial function, and calcium cycling in hiPSC-CMs, and reduces ischemia-reperfusion injury in mice by >50%.\",\n      \"method\": \"MAP4K4 gene silencing in hiPSC-CMs, small-molecule inhibitor (DMX-5804), oxidative stress assays, mitochondrial function, calcium cycling measurement, mouse ischemia-reperfusion model\",\n      \"journal\": \"Cell stem cell\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — genetic and pharmacological inhibition in human stem cell-derived cardiomyocytes and in vivo mouse model, multiple orthogonal functional readouts\",\n      \"pmids\": [\"30853557\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2019,\n      \"finding\": \"MAP4K4 activation regulates motor neuron degeneration in ALS via phosphorylated JNK3 and activation of the canonical c-Jun apoptotic pathway. MAP4K4 suppression promotes survival, prevents neurite degeneration, and decreases mutant SOD1 levels through autophagy (FoxO1-mediated) activation.\",\n      \"method\": \"MAP4K4 knockdown and pharmacological inhibition in ESC/iPSC-derived motor neurons and primary mouse spinal cord MNs, JNK3 phosphorylation assays, c-Jun pathway analysis, autophagy assays, SOD1 aggregate quantification\",\n      \"journal\": \"Cell reports\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — multiple cell models (ESC, iPSC, primary), genetic and pharmacological loss-of-function, mechanistic pathway dissection with JNK3/c-Jun and autophagy readouts\",\n      \"pmids\": [\"30699345\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2020,\n      \"finding\": \"STRIPAK complex member STRN4 is required for SV40 Small T antigen (ST)-PP2A-induced cell transformation. ST recruits STRIPAK (via striatin B''' subunits), which directs PP2A-mediated dephosphorylation of MAP4K4, activating the Hippo pathway effector YAP1 and promoting oncogenic transformation.\",\n      \"method\": \"ST-PP2A interaction proteomics, STRN4 knockdown transformation assays, MAP4K4 phosphorylation analysis, YAP1 activity assays\",\n      \"journal\": \"eLife\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1-2 / Strong — biochemical reconstitution of PP2A-STRIPAK-MAP4K4 axis, direct dephosphorylation demonstrated, genetic epistasis with STRN4 and YAP1\",\n      \"pmids\": [\"31913126\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2020,\n      \"finding\": \"MAP4K4 negatively regulates LFA-1 activation on CD8 T cells through ERM (ezrin, radixin, moesin) proteins, which serve as mediators between MAP4K4 and LFA-1. MAP4K4 deletion or inhibition enhances LFA-1 activation, improves T cell adherence to APCs, and increases CD8 T cell priming, proliferation, cytokine production, and cytotoxic activity. LFA-1 inhibition reverses these phenotypes.\",\n      \"method\": \"Genetic deletion (inducible whole-body Map4k4 KO), pharmacological inhibition, LFA-1 activation assays, T cell-APC conjugation assays, antitumor and antiviral in vivo models, LFA-1 inhibitor epistasis\",\n      \"journal\": \"Science immunology\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — genetic and pharmacological evidence, LFA-1 inhibitor epistasis defining ERM as mediator, multiple functional in vivo readouts\",\n      \"pmids\": [\"32220977\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2021,\n      \"finding\": \"MAP4K4 directly phosphorylates MLK3 on Thr738, increasing MLK3 kinase activity and downstream signaling, thereby promoting pancreatic cancer cell proliferation, migration, and colony formation. MAP4K4 and MLK3 co-immunoprecipitate.\",\n      \"method\": \"Co-immunoprecipitation, direct phosphorylation assay (Thr738 site-specific), cell proliferation and migration assays, pharmacological MAP4K4 inhibitor (GNE-495) in KPC mouse model\",\n      \"journal\": \"Oncogene\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1-2 / Strong — direct phosphorylation with site identification, Co-IP, gain-of-function rescue experiments, in vivo KPC mouse model validation\",\n      \"pmids\": [\"34511598\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2021,\n      \"finding\": \"MAP4K4 is regulated within STRIPAK complexes in cardiomyocytes. FLAG-MAP4K4 associates with all three striatins (STRN1, STRN3, STRN4) and with myosin. Calyculin A (Ser/Thr phosphatase inhibitor) induces MAP4K4 hyperphosphorylation of the activation loop and linker region in a kinase-activity-dependent manner. Surprisingly, FLAG-MAP4K4 inhibits JNK activation by H2O2 in cardiomyocytes and increases myofibrillar organization.\",\n      \"method\": \"5'-RACE and full-length cloning of rat Map4k4, FLAG-MAP4K4 co-immunoprecipitation with striatins and myosin, calyculin A treatment, phosphorylation mapping, kinase-dead mutant, JNK activation assay, myofibrillar staining\",\n      \"journal\": \"The Biochemical journal\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — biochemical Co-IP with all three striatins, activation loop phosphorylation demonstrated, kinase-dead control, unexpected finding (MAP4K4 inhibits JNK in this context) with appropriate caveats, single lab\",\n      \"pmids\": [\"34032269\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2022,\n      \"finding\": \"MAP4K4 interacts with striatin 3 (STRN3) via its CNH domain; STRN3 couples MAP4K4 to PP2A, which inactivates growth-repressing activities of MAP4K4. STRN3 also enables growth factor-induced PKCθ activation and direct phosphorylation of VASP-S157 by MAP4K4, which is required for efficient tumor cell invasion. Loss of STRN3 reactivates Hippo signaling.\",\n      \"method\": \"Co-immunoprecipitation (MAP4K4-STRN3/4), CNH domain deletion constructs, STRN3 and MAP4K4 depletion, VASP-S157 phosphorylation assay, PKCθ activation assay, Hippo pathway (YAP) activity, invasion assays in cerebellar tissue\",\n      \"journal\": \"Communications biology\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1-2 / Strong — direct phosphorylation of VASP-S157 by MAP4K4 demonstrated, CNH domain interaction mapped, PP2A-STRIPAK regulation confirmed, multiple orthogonal functional assays\",\n      \"pmids\": [\"35941177\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2022,\n      \"finding\": \"RhoA counteracts abdominal aortic aneurysm formation by inhibiting MAP4K4 activity. In the absence of RhoA, the PP2A inhibitor Set sequesters PP2A, releasing MAP4K4 from inhibition and activating MAP kinase signaling including MAP4K4. MAP4K4 inhibitor (DMX-5804) decreases AAA formation in RhoA cKO mice.\",\n      \"method\": \"VSMC-specific RhoA conditional knockout mice, MAP4K4 activity measurement, Set-PP2A interaction assays, DMX-5804 pharmacological inhibition, aortic ring contractility, gene expression\",\n      \"journal\": \"Communications biology\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — genetic (RhoA cKO) and pharmacological (DMX-5804) evidence, Set-PP2A-MAP4K4 axis proposed with biochemical support, single lab\",\n      \"pmids\": [\"36207400\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2023,\n      \"finding\": \"MAP4K4 promotes ovarian cancer metastasis by phosphorylating ADAM10 at Ser436, which suppresses ADAM10-mediated cleavage of N-cadherin, leading to N-cadherin stabilization and enhanced cancer cell adhesion, migration, and invasion.\",\n      \"method\": \"MAP4K4 knockdown/overexpression, ADAM10 phosphorylation at Ser436 (site-specific), N-cadherin cleavage assays, cell adhesion/migration/invasion assays, pharmacological MAP4K4 inhibition in peritoneal metastasis model\",\n      \"journal\": \"Oncogene\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1-2 / Strong — direct substrate phosphorylation with site identification, cleavage assay showing functional consequence, in vivo metastasis model\",\n      \"pmids\": [\"36922678\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2023,\n      \"finding\": \"MAP4K4 regulates forces at cell-cell and cell-substrate adhesions to promote collective cell migration. MAP4K4 promotes focal adhesion disassembly through moesin phosphorylation but disassembles adherens junctions through a moesin-independent mechanism. Loss of MAP4K4 increases traction forces and tension at cell-cell adhesions, causing tensional disequilibrium in migrating clusters.\",\n      \"method\": \"MAP4K4 inactivation and overexpression in A431 carcinoma cells, traction force microscopy, intercellular force measurement, moesin phosphorylation assay, focal adhesion and adherens junction dynamics\",\n      \"journal\": \"Life science alliance\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — biophysical force measurements, moesin phosphorylation mechanistic link established, but moesin-independent adherens junction mechanism not fully resolved, single lab\",\n      \"pmids\": [\"37369604\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2024,\n      \"finding\": \"MAP4K4 promotes S-nitrosylation of Drp1 (SNO-Drp1) at Cys644 (human C644/mouse C650) in cardiac microvascular endothelial cells by inhibiting GPX4 expression, which stimulates endothelial ferroptosis and cardiac microvascular dysfunction in diabetes. MAP4K4 inhibition (DMX-5804) reduces SNO-Drp1, alleviates microvascular dysfunction, and improves cardiac function in db/db mice; C650A Drp1 mutation abolishes SNO-Drp1 and its pathological effects.\",\n      \"method\": \"MAP4K4 overexpression/inhibition in CMECs, SNO-Drp1 site-specific analysis (C644/C650A mutation), GPX4 expression assay, ferroptosis assays, DMX-5804 in db/db mice, cardiac functional readouts\",\n      \"journal\": \"Cardiovascular diabetology\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1-2 / Strong — site-specific Cys→Ala mutation abolishing SNO-Drp1, GPX4 mechanistic link, pharmacological and genetic validation in vivo, multiple orthogonal methods\",\n      \"pmids\": [\"38724987\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2018,\n      \"finding\": \"HGK (MAP4K4) activation by the diterpenoid Tanshinone IIA triggers JNK1-dependent c-Jun activation, leading to increased c-Jun recruitment to the AP-1-binding site in the sestrin 2 (SESN2) promoter and upregulation of SESN2 transcription. This SESN2-dependent (but Beclin 1-independent) autophagy mediates the antitumor effects of TIIA in osteosarcoma.\",\n      \"method\": \"ChIP assay (c-Jun at SESN2 promoter), HGK/JNK1/Jun pathway dissection, autophagy assays (SESN2-dependent vs Beclin 1-dependent), in vitro and in vivo (NOD/SCID) osteosarcoma models\",\n      \"journal\": \"Cell death & disease\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — ChIP confirms c-Jun at SESN2 promoter, SESN2 specificity established by depletion, single lab\",\n      \"pmids\": [\"30258193\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2016,\n      \"finding\": \"MAP4K4 promotes NFκB signaling and MAP4K4 silencing in hepatocellular carcinoma cells represses JNK, NFκB, and toll-like receptor signaling pathways. In vitro, MAP4K4 knockdown reduces proliferation, blocks cell cycle at S phase, and increases apoptosis.\",\n      \"method\": \"shRNA knockdown in HCC cell lines, quantitative real-time PCR arrays for signaling pathway analysis, cell cycle analysis, apoptosis assay, xenograft tumor growth\",\n      \"journal\": \"Clinical cancer research\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — shRNA KD with pathway-level PCR array readout, multiple cellular phenotype readouts, in vivo xenograft, but signaling pathway assignment is broad, single lab\",\n      \"pmids\": [\"21196414\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2013,\n      \"finding\": \"MAP4K4 is required for KSHV lytic reactivation from latency and promotes invasiveness of KSHV-infected endothelial cells by regulating expression of COX-2, MMP-7, and MMP-13.\",\n      \"method\": \"Small molecule library screening and siRNA silencing in KSHV-infected endothelial cells, COX-2/MMP expression assays, invasion assays\",\n      \"journal\": \"PLoS pathogens\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — siRNA and small-molecule screening convergent result, multiple downstream target analysis, single lab\",\n      \"pmids\": [\"24244164\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2013,\n      \"finding\": \"MAP4K4 interacts with Pyk2 via the Pyk2 FERM domain, co-immunoprecipitates with Pyk2 (but not FAK), and is a substrate for Pyk2. MAP4K4 knockdown inhibits glioma cell migration and blocks Pyk2 stimulation of cell migration; MAP4K4 overexpression-stimulated migration is blocked by Pyk2 knockdown, indicating mutual dependence.\",\n      \"method\": \"Yeast two-hybrid screen, co-immunoprecipitation, in vitro substrate assay (Pyk2 phosphorylates MAP4K4), siRNA knockdown, cell migration assays\",\n      \"journal\": \"Journal of signal transduction\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — yeast 2-hybrid validated by Co-IP, substrate relationship confirmed in vitro, bidirectional knockdown epistasis, single lab\",\n      \"pmids\": [\"24163766\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2021,\n      \"finding\": \"MAP4K4 directly controls integrin β1 activation and c-Met endocytosis in medulloblastoma cells, promoting receptor recycling that accumulates activated c-Met in cytosolic vesicles and sustains downstream signaling. MAP4K4 depletion restricts HGF-driven matrix invasion in vitro and brain tissue infiltration ex vivo.\",\n      \"method\": \"MAP4K4 siRNA depletion, HGF-stimulated invasion and 3D migration assays, integrin β1 activation assay, c-Met endocytosis and recycling measurement, ex vivo brain slice infiltration\",\n      \"journal\": \"Oncotarget\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — multiple functional assays, endocytic recycling mechanism directly measured, single lab\",\n      \"pmids\": [\"29796184\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2024,\n      \"finding\": \"Dipeptide transport by PEPT1 activates the MAP4K4/G3BP2 signaling pathway in hepatocellular carcinoma cells, leading to phosphorylation of G3BP2 at Thr227 by MAP4K4, which facilitates HCC metastasis.\",\n      \"method\": \"PEPT1 overexpression/knockdown, MAP4K4/G3BP2 pathway analysis, G3BP2-Thr227 phosphorylation assay, migration/invasion assays, lung metastasis mouse model\",\n      \"journal\": \"Advanced science\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — direct G3BP2 phosphorylation at Thr227 demonstrated, in vivo metastasis model, dipeptide-to-kinase activation link established, single lab\",\n      \"pmids\": [\"38639383\"],\n      \"is_preprint\": false\n    }\n  ],\n  \"current_model\": \"MAP4K4 is a Ste20-family serine/threonine kinase that functions as a multi-context stress and inflammatory signaling hub: it activates JNK via a TAK1→MKK4/MKK7 cascade downstream of TNF-α/TNFR1, phosphorylates moesin to promote focal adhesion disassembly and integrin inactivation during cell migration (in a MAP4K4→moesin→talin/β1-integrin pathway), delivers to focal adhesions via EB2-microtubule tracks to activate Arf6 through IQSEC1, directly phosphorylates substrates including TRAF2 (promoting lysosomal degradation), MLK3-Thr738, ADAM10-Ser436, VASP-Ser157, FARP1, and Drp1 at Cys644 (facilitating S-nitrosylation via GPX4 suppression), inhibits mTORC1/AMPK signaling to suppress PPARγ translation and adipocyte lipogenesis, is regulated by STRIPAK complexes (striatins tether PP2A to keep MAP4K4 dephosphorylated/inactive), and acts redundantly with MINK1/TNIK upstream of DLK to drive retrograde JNK stress signaling in neurons.\"\n}\n```","stage2_raw":"{\n  \"mechanistic_narrative\": \"MAP4K4 is a Ste20-family serine/threonine kinase that functions as a stress- and inflammation-responsive signaling hub coupling extracellular cues to JNK-pathway activation, cytoskeletal remodeling, and metabolic control [#0, #6]. Classically, MAP4K4 acts upstream of a TAK1\\u2192MKK4/MKK7\\u2192JNK cascade and is itself activated by TNF-\\u03b1 through TNFR1, with TNF-\\u03b1 also transcriptionally inducing Map4k4 via c-Jun and ATF2 to create a feed-forward inflammatory loop [#0, #4]. In cell migration, MAP4K4 directly phosphorylates moesin at sites of focal adhesion disassembly, where moesin competes with talin to inactivate \\u03b21-integrin and promote membrane retraction; MAP4K4 is delivered to focal adhesions along EB2-bound microtubules to activate Arf6 through IQSEC1, driving integrin internalization and adhesion turnover [#9, #10, #28]. Beyond moesin, MAP4K4 directly phosphorylates a defined set of substrates to control distinct outputs: TRAF2 (promoting its lysosomal degradation to limit IL-6) [#11], MLK3 at Thr738 [#23], ADAM10 at Ser436 (stabilizing N-cadherin) [#27], VASP at Ser157 [#25], FARP1 [#12], and Drp1 (facilitating its S-nitrosylation at Cys644 via GPX4 suppression to drive endothelial ferroptosis) [#29]. Metabolically, MAP4K4 suppresses adipogenesis and lipogenesis by inhibiting mTOR/mTORC1\\u2014reducing PPAR\\u03b3 protein translation and, via AMPK-mediated raptor phosphorylation, suppressing SREBP-1\\u2014and mediates TNF-\\u03b1-induced insulin resistance [#2, #5, #8, #15]. MAP4K4 activity is restrained by STRIPAK: striatins (STRN1/3/4) tether PP2A to keep MAP4K4 dephosphorylated and inactive, linking it to Hippo/YAP and growth control [#21, #25]. Functionally, MAP4K4 promotes neuronal degeneration acting redundantly with MINK1 and TNIK upstream of DLK-driven retrograde JNK stress signaling [#16, #20], drives oxidative-stress cardiomyocyte death and ischemia-reperfusion injury [#19], and restrains CD8 T-cell LFA-1 activation through ERM proteins [#22].\",\n  \"teleology\": [\n    {\n      \"year\": 1999,\n      \"claim\": \"Established MAP4K4's core signaling identity by placing it as a JNK-selective upstream kinase responsive to TNF-\\u03b1, defining the pathway architecture all later work builds on.\",\n      \"evidence\": \"Dominant-negative epistasis with TAK1/MKK4/MKK7 and TNF-\\u03b1 stimulation in 293T cells\",\n      \"pmids\": [\"9890973\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Did not identify direct physiological substrates\", \"Mechanism of TNF-\\u03b1-induced MAP4K4 activation left unresolved\"]\n    },\n    {\n      \"year\": 2006,\n      \"claim\": \"Linked MAP4K4 to physiological outputs\\u2014cancer cell migration via JNK and metabolic regulation of adipogenesis/glucose transport\\u2014revealing it as more than a stress kinase.\",\n      \"evidence\": \"Genome-scale siRNA wound-healing screen across carcinoma lines, and RNAi kinase screen in adipocytes with glucose transport readouts\",\n      \"pmids\": [\"16537454\", \"16461467\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Migration mechanism downstream of JNK undefined\", \"Direct adipocyte substrates not identified\"]\n    },\n    {\n      \"year\": 2007,\n      \"claim\": \"Defined a feed-forward inflammatory circuit and the insulin-resistance mechanism: TNFR1 induces Map4k4 transcription via c-Jun/ATF2, and MAP4K4 mediates TNF-\\u03b1-driven IRS-1 serine phosphorylation.\",\n      \"evidence\": \"Receptor-specific antibodies/siRNA, c-Jun/ATF2 depletion, and epistasis with MAP2K1/MAP2K4 in primary human muscle\",\n      \"pmids\": [\"17500068\", \"17227768\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Whether MAP4K4 directly phosphorylates IRS-1 not established\", \"In vivo relevance to systemic insulin resistance not yet tested\"]\n    },\n    {\n      \"year\": 2009,\n      \"claim\": \"Resolved the mechanism of metabolic suppression: MAP4K4 inhibits mTOR signaling to reduce PPAR\\u03b3 translation and acts in macrophages to drive cytokine production, with systemic protective effects.\",\n      \"evidence\": \"Metabolic labeling/cycloheximide chase with mTOR-4E-BP1 analysis in adipocytes, and in vivo macrophage siRNA in an LPS lethality model\",\n      \"pmids\": [\"20038583\", \"19407801\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"How MAP4K4 inhibits mTOR mechanistically not fully defined\", \"Direct kinase substrate in the mTOR axis unknown\"]\n    },\n    {\n      \"year\": 2012,\n      \"claim\": \"Showed MAP4K4 kinase activity inhibits myogenesis via Myf5, expanding its developmental roles beyond canonical MAPK cascades.\",\n      \"evidence\": \"Bidirectional gain/loss-of-function and kinase-dead mutant with Myf5 rescue in C2C12 cells\",\n      \"pmids\": [\"23207904\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Direct substrate linking MAP4K4 to Myf5 unidentified\", \"ERK/p38/JNK-independent mechanism not molecularly defined\"]\n    },\n    {\n      \"year\": 2013,\n      \"claim\": \"Connected MAP4K4 to adhesion-receptor signaling through Pyk2 interaction and demonstrated roles in viral (KSHV) reactivation and invasion.\",\n      \"evidence\": \"Yeast two-hybrid/Co-IP with Pyk2 FERM domain plus migration epistasis, and siRNA/small-molecule screening in KSHV-infected endothelial cells\",\n      \"pmids\": [\"24163766\", \"24244164\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Pyk2-MAP4K4 phosphorylation directionality and sites incompletely mapped\", \"KSHV regulation is single-lab, mechanism upstream of COX-2/MMPs unclear\"]\n    },\n    {\n      \"year\": 2013,\n      \"claim\": \"Dissected the mTORC1/AMPK arm of metabolic control, showing MAP4K4 suppresses lipogenesis via AMPK-raptor-Ser792 and SREBP-1 independently of JNK.\",\n      \"evidence\": \"Isotope incorporation assays, genetic perturbation, and rapamycin/JNK-depletion controls in adipocytes\",\n      \"pmids\": [\"23924694\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Whether MAP4K4 directly phosphorylates AMPK not shown\", \"Tissue-level lipogenic relevance not yet tested in vivo\"]\n    },\n    {\n      \"year\": 2014,\n      \"claim\": \"Defined the direct cytoskeletal mechanism of migration: MAP4K4 phosphorylates moesin to inactivate \\u03b21-integrin and is trafficked to focal adhesions via EB2 microtubules to activate Arf6 through IQSEC1.\",\n      \"evidence\": \"Substrate phosphorylation, genetic epistasis, integrin blockade, inducible KO mice, and FA-complex proteomics with Arf6 activation assays\",\n      \"pmids\": [\"25799996\", \"25490267\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"How MAP4K4 selects moesin among ERM proteins at adhesions unclear\", \"Coordination of microtubule delivery with kinase activation not resolved\"]\n    },\n    {\n      \"year\": 2014,\n      \"claim\": \"Identified TRAF2 as a direct degradative substrate linking MAP4K4 to immune cytokine control, with in vivo consequences for adipose inflammation and insulin resistance.\",\n      \"evidence\": \"Direct phosphorylation assay and T-cell-specific conditional KO with multiple cytokine/genetic epistasis readouts\",\n      \"pmids\": [\"25098764\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"TRAF2 phosphosite(s) and lysosomal targeting mechanism not fully mapped\"]\n    },\n    {\n      \"year\": 2015,\n      \"claim\": \"Broadened the direct substrate catalog (FARP1) and provided structural/chemical tools, while clarifying tissue-specific metabolic genetics in vivo.\",\n      \"evidence\": \"Phosphoproteomics/in vitro kinase profiling for FARP1, X-ray crystallography of inhibitor-bound MAP4K4, and tissue-specific conditional KO metabolic phenotyping\",\n      \"pmids\": [\"26422651\", \"25908259\", \"25918248\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"FARP1 phosphosite physiological role undefined\", \"Why Myf5-tissue deletion improves adipose insulin sensitivity unexplained\"]\n    },\n    {\n      \"year\": 2015,\n      \"claim\": \"Extended MAP4K4 to vascular inflammation through NF-\\u03baB regulation in endothelium, with atheroprotection upon inhibition.\",\n      \"evidence\": \"Endothelial-specific gene ablation and small-molecule inhibition in Apoe-/- and Ldlr-/- mice with NF-\\u03baB readouts\",\n      \"pmids\": [\"26688060\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Direct MAP4K4 substrate in the NF-\\u03baB pathway not identified\"]\n    },\n    {\n      \"year\": 2016,\n      \"claim\": \"Implicated MAP4K4 in hepatocellular carcinoma proliferation and survival through broad JNK/NF-\\u03baB/TLR pathway engagement.\",\n      \"evidence\": \"shRNA knockdown with PCR-array pathway profiling, cell cycle/apoptosis assays, and xenografts\",\n      \"pmids\": [\"21196414\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Signaling assignment broad and correlative\", \"No direct substrate identified, single-lab\"]\n    },\n    {\n      \"year\": 2017,\n      \"claim\": \"Established neuronal degeneration roles: MAP4K4 acts redundantly with MINK1/TNIK upstream of DLK-driven retrograde JNK stress signaling.\",\n      \"evidence\": \"Combinatorial siRNA/pharmacological inhibition in DRG neurons with DLK phosphorylation and retrograde-transport readouts\",\n      \"pmids\": [\"28993483\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Whether MAP4K4 directly phosphorylates DLK not established\", \"Relative contribution of each MAP4K family member in vivo unresolved\"]\n    },\n    {\n      \"year\": 2017,\n      \"claim\": \"Revealed regulatory complexity: alternative splicing of exon 16 (SRSF3/RBM4) tunes JNK1 signaling and EMT, and MAP4K4 can positively regulate ERK by inhibiting PP2A in lung cancer.\",\n      \"evidence\": \"Splice-variant constructs with JNK1/EMT readouts, and knockdown with PP2A activity/ERK phosphorylation assays plus xenografts\",\n      \"pmids\": [\"29138007\", \"28306189\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Functional difference between splice isoforms mechanistically incomplete\", \"How MAP4K4 inhibits PP2A unclear, both single-lab\"]\n    },\n    {\n      \"year\": 2018,\n      \"claim\": \"Linked MAP4K4-JNK1-c-Jun signaling to SESN2-dependent autophagy as an antitumor mechanism, and showed MAP4K4 controls c-Met recycling/integrin \\u03b21 in medulloblastoma invasion.\",\n      \"evidence\": \"ChIP of c-Jun at the SESN2 promoter with autophagy assays in osteosarcoma, and endocytosis/recycling measurements in medulloblastoma\",\n      \"pmids\": [\"30258193\", \"29796184\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Pharmacological MAP4K4 activation specificity not fully controlled\", \"Direct substrate controlling c-Met endocytosis unidentified\"]\n    },\n    {\n      \"year\": 2019,\n      \"claim\": \"Demonstrated MAP4K4 as a driver of oxidative-stress cardiomyocyte death and motor-neuron degeneration, nominating it as a therapeutic target in heart and ALS contexts.\",\n      \"evidence\": \"Genetic/pharmacological inhibition (DMX-5804) in hiPSC-CMs and mouse ischemia-reperfusion; knockdown/inhibition in ESC/iPSC/primary motor neurons with JNK3/c-Jun and autophagy readouts\",\n      \"pmids\": [\"30853557\", \"30699345\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Direct substrate mediating cardiomyocyte death not identified\", \"Link between MAP4K4 and FoxO1-mediated autophagy mechanistically incomplete\"]\n    },\n    {\n      \"year\": 2020,\n      \"claim\": \"Defined STRIPAK-PP2A as the inhibitory regulator that keeps MAP4K4 dephosphorylated, and showed MAP4K4 negatively regulates CD8 T-cell LFA-1 activation via ERM proteins.\",\n      \"evidence\": \"ST-PP2A proteomics with STRN4 knockdown/dephosphorylation/YAP1 assays, and genetic/pharmacological MAP4K4 perturbation with LFA-1 epistasis in tumor/viral models\",\n      \"pmids\": [\"31913126\", \"32220977\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Which MAP4K4 substrate(s) gate Hippo/YAP downstream of STRIPAK not fully defined\"]\n    },\n    {\n      \"year\": 2021,\n      \"claim\": \"Expanded the direct substrate repertoire (MLK3-Thr738) in pancreatic cancer and mapped CNH-domain-dependent STRN3 binding coupling MAP4K4 to PP2A and VASP-S157 phosphorylation.\",\n      \"evidence\": \"Co-IP, site-specific phosphorylation assays, CNH-deletion constructs, and in vivo KPC model\",\n      \"pmids\": [\"34511598\", \"34032269\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"How STRN3 binding switches between inhibition and substrate selection unresolved\", \"Context-dependent MAP4K4 inhibition of JNK in cardiomyocytes (single-lab) not generalized\"]\n    },\n    {\n      \"year\": 2022,\n      \"claim\": \"Clarified STRN3-PP2A control of MAP4K4 growth-repressing activity and VASP-driven invasion, and positioned RhoA-Set-PP2A as an upstream restraint relevant to aortic aneurysm.\",\n      \"evidence\": \"Co-IP/CNH-domain mapping with VASP-S157 and Hippo assays, and VSMC-specific RhoA cKO with Set-PP2A analysis and DMX-5804\",\n      \"pmids\": [\"35941177\", \"36207400\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Set-PP2A-MAP4K4 axis is proposed with limited reconstitution\", \"RhoA-to-MAP4K4 link is single-lab\"]\n    },\n    {\n      \"year\": 2023,\n      \"claim\": \"Added ADAM10-Ser436 as a direct substrate stabilizing N-cadherin in ovarian cancer, and resolved that MAP4K4 disassembles adherens junctions by a moesin-independent route in collective migration.\",\n      \"evidence\": \"Site-specific ADAM10 phosphorylation/cleavage assays with in vivo metastasis, and traction-force microscopy with moesin phosphorylation in A431 cells\",\n      \"pmids\": [\"36922678\", \"37369604\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Moesin-independent adherens-junction mechanism not molecularly defined\", \"ADAM10 finding single-lab\"]\n    },\n    {\n      \"year\": 2024,\n      \"claim\": \"Identified Drp1-Cys644 S-nitrosylation (via GPX4 suppression) as a MAP4K4-driven ferroptotic mechanism in diabetic cardiac microvasculature, and PEPT1-dipeptide activation of MAP4K4-G3BP2-Thr227 in HCC metastasis.\",\n      \"evidence\": \"Site-specific Cys-to-Ala mutation with GPX4/ferroptosis assays and DMX-5804 in db/db mice; PEPT1 perturbation with G3BP2-Thr227 phosphorylation and lung metastasis model\",\n      \"pmids\": [\"38724987\", \"38639383\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"How MAP4K4 suppresses GPX4 expression mechanistically unclear\", \"G3BP2-Thr227 phosphorylation is single-lab\"]\n    },\n    {\n      \"year\": null,\n      \"claim\": \"It remains unresolved how a single kinase integrates its many context-specific substrates and opposite outputs (e.g., JNK activation vs inhibition; growth promotion vs repression), and what governs substrate selection downstream of STRIPAK-PP2A regulation.\",\n      \"evidence\": \"\",\n      \"pmids\": [],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"No unifying structural/regulatory model for substrate selection\", \"Context-dependent reversal of JNK output unexplained\", \"Endogenous activating signal that overrides STRIPAK inhibition undefined\"]\n    }\n  ],\n  \"mechanism_profile\": {\n    \"molecular_activity\": [\n      {\"term_id\": \"GO:0140096\", \"supporting_discovery_ids\": [9, 11, 12, 23, 25, 27, 29, 35]},\n      {\"term_id\": \"GO:0016740\", \"supporting_discovery_ids\": [0, 9, 23, 27]},\n      {\"term_id\": \"GO:0140110\", \"supporting_discovery_ids\": [2, 5]}\n    ],\n    \"localization\": [\n      {\"term_id\": \"GO:0005856\", \"supporting_discovery_ids\": [9, 10, 28]},\n      {\"term_id\": \"GO:0005829\", \"supporting_discovery_ids\": [10, 34]},\n      {\"term_id\": \"GO:0005815\", \"supporting_discovery_ids\": [10]}\n    ],\n    \"pathway\": [\n      {\"term_id\": \"R-HSA-162582\", \"supporting_discovery_ids\": [0, 16, 21]},\n      {\"term_id\": \"R-HSA-168256\", \"supporting_discovery_ids\": [6, 11, 22]},\n      {\"term_id\": \"R-HSA-1430728\", \"supporting_discovery_ids\": [2, 5, 8, 15]},\n      {\"term_id\": \"R-HSA-1500931\", \"supporting_discovery_ids\": [9, 10, 28]},\n      {\"term_id\": \"R-HSA-5357801\", \"supporting_discovery_ids\": [19, 20, 29]}\n    ],\n    \"complexes\": [\"STRIPAK\"],\n    \"partners\": [\"STRN3\", \"STRN4\", \"STRN1\", \"EB2\", \"IQSEC1\", \"TRAF2\", \"MLK3\", \"Pyk2\"],\n    \"other_free_text\": []\n  }\n}","audit_flag":null,"evaluation":{"pairwise":"tie","faith_supported":7,"faith_total":7,"faith_pct":100.0}}