{"gene":"MED28","run_date":"2026-04-28T18:30:28","timeline":{"discoveries":[{"year":2004,"finding":"MED28 (magicin) was identified as a merlin-specific binding partner that interacts with the NF2 tumor suppressor merlin in vitro and in vivo, colocalizes with merlin beneath the plasma membrane, and associates with the actin cytoskeleton as determined by cofractionation, immunofluorescence, and electron microscopy. MED28 also interacts with the adaptor protein Grb2 via its Grb2-binding motifs, and merlin can form a ternary complex with MED28 and Grb2.","method":"Co-immunoprecipitation, affinity binding, blot overlay, immunofluorescence, electron microscopy, subcellular fractionation","journal":"Oncogene","confidence":"High","confidence_rationale":"Tier 2 — multiple orthogonal methods (Co-IP, pulldown, blot overlay, EM, fractionation) in a single study","pmids":["15467741"],"is_preprint":false},{"year":2006,"finding":"MED28 (magicin) interacts with Fyn tyrosine kinase (identified by yeast two-hybrid), and Fyn phosphorylates magicin in vitro. Src and Lck also interact with magicin. Upon CD3 stimulation in Jurkat cells, magicin is phosphorylated by Lck (the major kinase, as shown by Lck-deficient J.CaM1.6 cells lacking phosphorylation). Site-directed mutagenesis and in vitro kinase assays identified Y64 as the Lck phosphorylation site, creating an SH2-Grb2 binding motif.","method":"Yeast two-hybrid, in vitro kinase assay, site-directed mutagenesis, co-immunoprecipitation, Lck-deficient cell line","journal":"Biochemical and biophysical research communications","confidence":"High","confidence_rationale":"Tier 1–2 — in vitro kinase assay plus mutagenesis plus genetic validation with Lck-deficient cell line","pmids":["16899217"],"is_preprint":false},{"year":2006,"finding":"EG-1 (MED28) overexpression activates c-Src signaling; EG-1 interacts with c-Src (by co-immunoprecipitation) and also binds other Src-family members as well as multiple SH3- and WW-domain-containing signaling molecules, but EG-1 is not a direct Src substrate.","method":"Co-immunoprecipitation, overexpression experiments","journal":"International journal of oncology","confidence":"Medium","confidence_rationale":"Tier 3 — single Co-IP plus overexpression, no mutagenesis or reconstitution","pmids":["16964398"],"is_preprint":false},{"year":2005,"finding":"EG-1 (MED28) overexpression stimulates cellular proliferation in vitro and in vivo (xenograft model), and is associated with activation of ERK1/2, JNK, and p38 MAPK kinases; co-immunoprecipitation showed an association between EG-1 and Src.","method":"Overexpression/siRNA transfection, xenograft assay, co-immunoprecipitation, immunoblot","journal":"Cancer research","confidence":"Medium","confidence_rationale":"Tier 3 — functional proliferation assay plus Co-IP, but no direct mechanistic dissection of pathway","pmids":["16024617"],"is_preprint":false},{"year":2007,"finding":"MED28 functions as a subunit of the mammalian Mediator complex and acts as a repressor of smooth muscle cell (SMC) differentiation. Knockdown of Med28 in NIH3T3 cells induces SMC differentiation genes; overexpression represses them. Med28 functions within the Mediator head module together with Med6, Med8, and Med18, and may act as a scaffolding protein maintaining this submodule's stability.","method":"siRNA knockdown, overexpression, gene expression analysis, genetic epistasis with Med6/Med8/Med18","journal":"The Journal of biological chemistry","confidence":"High","confidence_rationale":"Tier 2 — clean KD and OE with defined cellular phenotype, epistasis with head-module subunits, replicated bidirectionally","pmids":["17848560"],"is_preprint":false},{"year":2011,"finding":"MED28 overexpression enhances EGF-induced cellular migration in MDA-MB-231 breast cancer cells, and this effect is mediated through the EGFR/PI3K signaling pathway; resveratrol suppresses EGF-mediated migration by reducing MED28 and MMP-9 expression.","method":"Overexpression, migration assay, pharmacological inhibition, immunoblot","journal":"Journal of agricultural and food chemistry","confidence":"Medium","confidence_rationale":"Tier 3 — overexpression with functional migration assay, pathway inferred pharmacologically","pmids":["21942447"],"is_preprint":false},{"year":2012,"finding":"MED28 regulates cellular migration and invasion in human breast cancer cells in a MEK1-dependent manner: MED28 knockdown reduces MEK1 and MMP2 expression; MEK1 suppression blocks MED28-induced MMP2 activation and migration; ectopic MEK1 rescues MED28 knockdown-mediated invasion suppression; and exogenous MMP2 rescues invasion upon MED28 or MEK1 knockdown.","method":"siRNA knockdown, dominant-negative construct, MEK1 inhibitors, rescue experiments with MEK1 cDNA and recombinant MMP2, migration/invasion assay","journal":"Journal of cellular physiology","confidence":"High","confidence_rationale":"Tier 2 — epistasis by multiple genetic approaches (siRNA, dominant-negative, rescue) with defined pathway order","pmids":["22495818"],"is_preprint":false},{"year":2015,"finding":"Med28 is essential for mouse peri-implantation development; knockout causes lethality due to loss of inner cell mass pluripotency with reduced Oct4 and Nanog expression. Overexpression of Med28 in mouse embryonic fibroblasts enhances reprogramming efficiency to iPSCs. Cre-mediated inactivation in iPSCs demonstrates Med28 is required for their survival. MED28 has both cytosolic (merlin-interacting) and nuclear (Mediator complex) roles.","method":"Knockout mouse model, Cre-mediated conditional inactivation, reprogramming assay, gene expression analysis","journal":"PloS one","confidence":"High","confidence_rationale":"Tier 2 — clean genetic KO with defined developmental phenotype and molecular markers, plus reprogramming functional assay","pmids":["26445504"],"is_preprint":false},{"year":2015,"finding":"MED28 knockdown in colorectal cancer cells reduces cyclin D1, c-Myc, and nuclear β-catenin expression, while increasing E-cadherin and HBP1 (a negative Wnt/β-catenin regulator); MED28 suppresses HBP1 promoter activity (luciferase reporter assay), and overexpression has the opposite effects, placing MED28 upstream of Wnt/β-catenin signaling via HBP1.","method":"siRNA knockdown, overexpression, luciferase reporter assay, immunoblot","journal":"Journal of cellular physiology","confidence":"Medium","confidence_rationale":"Tier 2–3 — reporter assay plus KD/OE with defined pathway placement, single lab","pmids":["26660958"],"is_preprint":false},{"year":2016,"finding":"MED28 regulates epithelial-mesenchymal transition (EMT) in human breast cancer cells through the NFκB pathway: MED28 suppression attenuates mesenchymal markers and reduces p-NFκB/p65 and Snail; overexpression has the opposite effect. In MCF7 cells, adriamycin-induced EMT correlates with increased MED28 and p-p65, and MED28 knockdown reverses this EMT.","method":"siRNA knockdown, overexpression, immunoblot, morphological analysis, pharmacological EMT induction","journal":"Journal of cellular physiology","confidence":"Medium","confidence_rationale":"Tier 3 — KD/OE with defined phenotype and pathway marker, single lab without reconstitution","pmids":["27662245"],"is_preprint":false},{"year":2017,"finding":"MED28 interacts with ZNF224 in the nucleus (confirmed by co-immunoprecipitation and surface plasmon resonance); the KRAB domain of ZNF224 interacts with the MED domain of MED28. MED28 overexpression stabilizes ZNF224 against camptothecin-induced degradation, resulting in increased colony formation of MCF-7 cells.","method":"Co-immunoprecipitation, surface plasmon resonance, bimolecular fluorescence complementation, overexpression, colony formation assay","journal":"Oncology letters","confidence":"Medium","confidence_rationale":"Tier 2 — SPR plus Co-IP plus BiFC provide orthogonal binding evidence; functional consequence demonstrated","pmids":["29435049"],"is_preprint":false},{"year":2018,"finding":"MED28 interacts with FOXM1 (co-immunoprecipitation); MED28 and FOXM1 mutually affect each other's expression and subcellular localization. Both regulate MMP2-dependent migration and invasion in NSCLC cells; MED28 siRNA-mediated MMP2 suppression is rescued by constitutively active FOXM1, restoring migration and invasion.","method":"Co-immunoprecipitation, siRNA knockdown, doxycycline-inducible overexpression, rescue experiment, migration/invasion assay","journal":"Journal of cellular physiology","confidence":"Medium","confidence_rationale":"Tier 2–3 — Co-IP plus rescue epistasis, single lab","pmids":["30499104"],"is_preprint":false},{"year":2019,"finding":"MED28 expression is regulated by transcription factors E2F1, NRF1, ETS1, and C/EBPβ (identified by luciferase reporter assay). MED28 expression peaks at the G1-S transition and mitosis. Overexpression of MED28 shortens interphase and mitosis duration, whereas knockdown lengthens them. MED28 overexpression increases micronuclei, nuclear budding, and aneuploidy, indicating a role in maintaining genomic stability.","method":"Luciferase reporter assay, cell cycle synchronization, live cell imaging, flow cytometry, fluorescence microscopy, siRNA knockdown, overexpression","journal":"International journal of molecular sciences","confidence":"Medium","confidence_rationale":"Tier 2 — multiple orthogonal methods, single lab","pmids":["30970566"],"is_preprint":false},{"year":2020,"finding":"RCOR1 directly interacts with MED28 (demonstrated in OCSCC cells) and overexpression of RCOR1 abrogates MED28-induced cancer stem cell-like activity (colony/sphere formation and CSC marker expression), placing RCOR1 as a direct negative regulator of MED28 function.","method":"Co-immunoprecipitation (direct binding), overexpression, functional CSC assays","journal":"Journal of oral pathology & medicine","confidence":"Medium","confidence_rationale":"Tier 3 — single Co-IP with functional rescue, single lab","pmids":["32306431"],"is_preprint":false},{"year":2018,"finding":"MDT-28, the C. elegans orthologue of MED28, undergoes lysine acetylation (confirmed by anti-acetyl-lysine immunoprecipitation of GFP::MDT-28). Valproic acid (a lysine deacetylase inhibitor) enhances MDT-28 acetylation and decreases its nuclear localization as detected by FLIM, indicating that the nuclear pool of MED28 is regulated by acetylation.","method":"Immunoprecipitation with anti-acetyl-lysine antibody, fluorescence lifetime imaging microscopy (FLIM), pharmacological treatment with VPA","journal":"Folia biologica","confidence":"Medium","confidence_rationale":"Tier 2 — direct biochemical detection of PTM plus subcellular localization change in a model organism orthologue","pmids":["29871732"],"is_preprint":false},{"year":2024,"finding":"MED28 knockdown in hepatocellular carcinoma cells induces cell cycle arrest and suppresses AKT/mTOR signaling, reduces lipid accumulation, and decreases expression and nuclear localization of SREBP1; MED28 overexpression upregulates AKT/mTOR signaling, placing MED28 upstream of this pathway in liver cancer.","method":"siRNA knockdown, overexpression, immunoblot, subcellular fractionation, cell cycle analysis","journal":"Journal of agricultural and food chemistry","confidence":"Medium","confidence_rationale":"Tier 2–3 — bidirectional KD/OE with defined pathway readouts, single lab","pmids":["38619972"],"is_preprint":false}],"current_model":"MED28 is a dual-compartment protein that functions as a subunit of the mammalian Mediator transcriptional co-activator complex in the nucleus—where it acts as part of the head module (with Med6, Med8, Med18) to repress smooth muscle cell differentiation, support pluripotency, and regulate cell cycle progression—while in the cytoplasm it associates with the actin cytoskeleton as a merlin/Grb2-interacting scaffold (magicin) that is phosphorylated at Y64 by Src-family kinases (principally Lck/Fyn) upon receptor stimulation, and interacts with signaling partners including FOXM1 and ZNF224 to regulate MEK1–MMP2-dependent cell migration, EMT via NFκB, Wnt/β-catenin signaling via HBP1, and AKT/mTOR-dependent proliferation in cancer cells."},"narrative":{"teleology":[{"year":2004,"claim":"Establishing that MED28 is a cytoplasmic scaffolding protein that physically links the NF2 tumor suppressor merlin to the adaptor Grb2 at the cortical actin cytoskeleton resolved the question of how merlin connects to growth factor signaling complexes.","evidence":"Co-IP, affinity binding, blot overlay, EM, and subcellular fractionation in mammalian cells","pmids":["15467741"],"confidence":"High","gaps":["Functional consequence of merlin–MED28–Grb2 ternary complex on downstream signaling not determined","Whether the cytoplasmic pool of MED28 is stoichiometrically distinct from the Mediator-bound nuclear pool was unknown"]},{"year":2005,"claim":"Demonstrating that MED28 overexpression drives proliferation in vitro and in xenografts while activating ERK1/2, JNK, and p38 MAPK established MED28 as a pro-proliferative factor acting upstream of multiple MAPK cascades.","evidence":"Overexpression and siRNA in cancer cell lines, xenograft model, Co-IP with Src, immunoblot","pmids":["16024617"],"confidence":"Medium","gaps":["No direct substrate or kinase–substrate relationship defined for MED28 in MAPK activation","Correlation between Src interaction and MAPK activation not causally dissected"]},{"year":2006,"claim":"Identification of Y64 as the Lck phosphorylation site on MED28—using Lck-deficient cells and site-directed mutagenesis—revealed how TCR stimulation creates an SH2 docking site for Grb2 on MED28, providing a direct biochemical mechanism for signal-dependent scaffold assembly.","evidence":"Yeast two-hybrid, in vitro kinase assay, mutagenesis, Lck-deficient J.CaM1.6 cells","pmids":["16899217"],"confidence":"High","gaps":["Downstream signaling events specifically dependent on pY64-Grb2 recruitment not mapped","Whether Y64 phosphorylation occurs in non-hematopoietic contexts not tested"]},{"year":2007,"claim":"Showing that MED28 is a bona fide subunit of the Mediator head module (with Med6/Med8/Med18) that represses smooth muscle differentiation genes established its nuclear transcriptional role, distinct from its cytoplasmic scaffolding function.","evidence":"siRNA knockdown, overexpression, genetic epistasis with head-module subunits in NIH3T3 cells","pmids":["17848560"],"confidence":"High","gaps":["Structural basis for MED28 integration into the head module not resolved","Mechanism by which head module occupancy at SMC gene promoters leads to repression not determined"]},{"year":2012,"claim":"Epistatic rescue experiments placing MED28 upstream of MEK1 and MMP2 in breast cancer cell migration/invasion defined a linear signaling hierarchy (MED28→MEK1→MMP2) through which MED28 promotes invasiveness.","evidence":"siRNA, dominant-negative constructs, MEK1 cDNA rescue, recombinant MMP2 rescue, migration/invasion assays in breast cancer cells","pmids":["22495818"],"confidence":"High","gaps":["Direct biochemical link between MED28 and MEK1 activation (e.g., Raf-level regulation) not identified","Whether this pathway operates through cytoplasmic or nuclear MED28 not distinguished"]},{"year":2015,"claim":"Genetic knockout in mice and conditional inactivation in iPSCs demonstrated that MED28 is essential for peri-implantation development and pluripotency maintenance via Oct4/Nanog, and that it enhances somatic reprogramming—establishing a non-redundant in vivo requirement.","evidence":"Knockout mouse model, Cre-mediated conditional inactivation, iPSC reprogramming assay, gene expression analysis","pmids":["26445504"],"confidence":"High","gaps":["Whether the developmental requirement is mediated by Mediator function, cytoplasmic signaling, or both is unresolved","Tissue-specific conditional knockouts beyond peri-implantation not reported"]},{"year":2015,"claim":"Demonstrating that MED28 suppresses HBP1 promoter activity to activate Wnt/β-catenin target genes (cyclin D1, c-Myc) in colorectal cancer cells identified a transcriptional mechanism linking MED28 to Wnt pathway output.","evidence":"siRNA knockdown, overexpression, luciferase reporter assay, immunoblot in colorectal cancer cells","pmids":["26660958"],"confidence":"Medium","gaps":["Whether MED28 represses HBP1 directly at its promoter via Mediator or indirectly through an intermediary not resolved","In vivo relevance of this axis not tested"]},{"year":2016,"claim":"Linking MED28 to EMT through NF-κB/p65 phosphorylation and Snail expression in breast cancer cells expanded its oncogenic repertoire beyond proliferation and migration to include phenotypic plasticity.","evidence":"siRNA knockdown, overexpression, adriamycin-induced EMT model, immunoblot in MCF7 and MDA-MB-231 cells","pmids":["27662245"],"confidence":"Medium","gaps":["Direct mechanism by which MED28 activates NF-κB (e.g., IKK regulation) not identified","Single-lab finding without independent replication"]},{"year":2017,"claim":"Identification of ZNF224 as a direct nuclear interaction partner of MED28—with binding mapped to the KRAB domain of ZNF224 and the MED domain of MED28—revealed a mechanism by which MED28 stabilizes a transcriptional repressor to promote colony formation.","evidence":"Co-IP, surface plasmon resonance, bimolecular fluorescence complementation, colony assay in MCF-7 cells","pmids":["29435049"],"confidence":"Medium","gaps":["Genomic targets co-regulated by MED28–ZNF224 not identified","Whether ZNF224 stabilization requires Mediator association is unknown"]},{"year":2018,"claim":"Showing that the C. elegans orthologue MDT-28 undergoes lysine acetylation that reduces its nuclear localization revealed a conserved post-translational mechanism governing the cytoplasmic-to-nuclear partitioning of MED28.","evidence":"Anti-acetyl-lysine immunoprecipitation of GFP::MDT-28, FLIM imaging, VPA treatment in C. elegans","pmids":["29871732"],"confidence":"Medium","gaps":["Specific acetylated lysine residues not mapped","Whether mammalian MED28 is similarly regulated by acetylation not tested","Identity of the acetyltransferase and deacetylase acting on MDT-28/MED28 unknown"]},{"year":2018,"claim":"Demonstrating that MED28 and FOXM1 mutually regulate each other's expression and co-drive MMP2-dependent invasion provided evidence of a feed-forward loop in NSCLC that converges on the same MMP2 effector pathway identified in breast cancer.","evidence":"Co-IP, siRNA, doxycycline-inducible overexpression, FOXM1 rescue of MED28 knockdown, invasion assay in NSCLC cells","pmids":["30499104"],"confidence":"Medium","gaps":["Whether MED28–FOXM1 interaction occurs within the Mediator complex or independently not distinguished","Mechanism of mutual transcriptional regulation not resolved"]},{"year":2019,"claim":"Establishing that MED28 expression is cell-cycle-regulated (peaking at G1–S and mitosis) and that its overexpression shortens both interphase and mitosis while increasing aneuploidy connected MED28 to genomic stability and cell-cycle timing.","evidence":"Cell cycle synchronization, live cell imaging, flow cytometry, fluorescence microscopy, luciferase reporter for promoter analysis","pmids":["30970566"],"confidence":"Medium","gaps":["Molecular targets through which MED28 accelerates cell cycle phases not identified","Whether aneuploidy results from Mediator-dependent transcriptional mis-regulation or cytoskeletal defects is unclear"]},{"year":2024,"claim":"Placing MED28 upstream of AKT/mTOR signaling and SREBP1-dependent lipogenesis in hepatocellular carcinoma cells broadened its oncogenic functions to include metabolic reprogramming.","evidence":"siRNA knockdown, overexpression, immunoblot, subcellular fractionation, cell cycle analysis in HCC cells","pmids":["38619972"],"confidence":"Medium","gaps":["Direct biochemical mechanism linking MED28 to AKT phosphorylation not determined","In vivo validation in liver cancer models not reported"]},{"year":null,"claim":"The mechanism governing MED28 partitioning between its Mediator-associated nuclear pool and its cytoplasmic actin-associated pool, and whether these represent functionally independent or coordinated activities, remains unresolved.","evidence":"","pmids":[],"confidence":"Low","gaps":["No structural data for MED28 within the Mediator head module at sufficient resolution to define contacts","Regulatory signals controlling nuclear–cytoplasmic shuttling in mammalian cells not identified","Whether cytoplasmic signaling functions of MED28 feed back into its Mediator-dependent transcriptional activity is untested"]}],"mechanism_profile":{"molecular_activity":[{"term_id":"GO:0060090","term_label":"molecular adaptor activity","supporting_discovery_ids":[0,1,10,11]},{"term_id":"GO:0140110","term_label":"transcription regulator activity","supporting_discovery_ids":[4,7,8]},{"term_id":"GO:0098772","term_label":"molecular function regulator activity","supporting_discovery_ids":[4,6,9]}],"localization":[{"term_id":"GO:0005856","term_label":"cytoskeleton","supporting_discovery_ids":[0]},{"term_id":"GO:0005634","term_label":"nucleus","supporting_discovery_ids":[4,7,10,14]},{"term_id":"GO:0005829","term_label":"cytosol","supporting_discovery_ids":[0,1]}],"pathway":[{"term_id":"R-HSA-74160","term_label":"Gene expression (Transcription)","supporting_discovery_ids":[4,7,8]},{"term_id":"R-HSA-162582","term_label":"Signal Transduction","supporting_discovery_ids":[1,3,6,9,15]},{"term_id":"R-HSA-1640170","term_label":"Cell Cycle","supporting_discovery_ids":[12]},{"term_id":"R-HSA-1266738","term_label":"Developmental Biology","supporting_discovery_ids":[4,7]}],"complexes":["Mediator complex (head module)"],"partners":["NF2","GRB2","LCK","FYN","MED6","FOXM1","ZNF224","RCOR1"],"other_free_text":[]},"mechanistic_narrative":"MED28 is a dual-compartment scaffolding protein that functions both as a subunit of the Mediator transcriptional co-activator complex and as a cytoplasmic signaling adaptor linking receptor-proximal kinases to downstream proliferative and migratory pathways. Within the nucleus, MED28 operates in the Mediator head module together with Med6, Med8, and Med18 to repress smooth muscle cell differentiation, maintain pluripotency through Oct4/Nanog expression, and regulate cell cycle progression at the G1–S transition and mitosis [PMID:17848560, PMID:26445504, PMID:30970566]. In the cytoplasm, MED28 associates with the actin cytoskeleton and the tumor suppressor merlin (NF2), is phosphorylated at Y64 by Src-family kinases (principally Lck) to create an SH2-binding motif for Grb2, and promotes cell migration and invasion through a MEK1–MMP2 axis and NF-κB–dependent EMT [PMID:15467741, PMID:16899217, PMID:22495818, PMID:27662245]. MED28 also activates Wnt/β-catenin signaling by suppressing the negative regulator HBP1 and promotes AKT/mTOR-dependent proliferation and lipogenesis in cancer cells [PMID:26660958, PMID:38619972]."},"prefetch_data":{"uniprot":{"accession":"Q9H204","full_name":"Mediator of RNA polymerase II transcription subunit 28","aliases":["Endothelial-derived protein 1","Mediator complex subunit 28","Merlin and Grb2-interacting cytoskeletal protein","Magicin","Tumor angiogenesis marker EG-1"],"length_aa":178,"mass_kda":19.5,"function":"Component of the Mediator complex, a coactivator involved in the regulated transcription of nearly all RNA polymerase II-dependent genes. Mediator functions as a bridge to convey information from gene-specific regulatory proteins to the basal RNA polymerase II transcription machinery. Mediator is recruited to promoters by direct interactions with regulatory proteins and serves as a scaffold for the assembly of a functional preinitiation complex with RNA polymerase II and the general transcription factors. May be part of a complex containing NF2/merlin that participates in cellular signaling to the actin cytoskeleton downstream of tyrosine kinase signaling pathways","subcellular_location":"Nucleus; Cytoplasm; Membrane","url":"https://www.uniprot.org/uniprotkb/Q9H204/entry"},"depmap":{"release":"DepMap","has_data":true,"is_common_essential":true,"resolved_as":"","url":"https://depmap.org/portal/gene/MED28","classification":"Common Essential","n_dependent_lines":1167,"n_total_lines":1208,"dependency_fraction":0.9660596026490066},"opencell":{"profiled":true,"resolved_as":"","ensg_id":"ENSG00000118579","cell_line_id":"CID000243","localizations":[{"compartment":"nuclear_punctae","grade":3},{"compartment":"nucleoplasm","grade":3}],"interactors":[{"gene":"MED10","stoichiometry":10.0},{"gene":"MED11","stoichiometry":10.0},{"gene":"MED14","stoichiometry":10.0},{"gene":"MED17","stoichiometry":10.0},{"gene":"MED18","stoichiometry":10.0},{"gene":"MED19","stoichiometry":10.0},{"gene":"MED20","stoichiometry":10.0},{"gene":"MED21","stoichiometry":10.0},{"gene":"MED22","stoichiometry":10.0},{"gene":"MED27","stoichiometry":10.0}],"url":"https://opencell.sf.czbiohub.org/target/CID000243","total_profiled":1310},"omim":[{"mim_id":"620327","title":"NEURODEGENERATION WITH DEVELOPMENTAL DELAY, EARLY RESPIRATORY FAILURE, MYOCLONIC SEIZURES, AND BRAIN ABNORMALITIES; NDDRSB","url":"https://www.omim.org/entry/620327"},{"mim_id":"612383","title":"MEDIATOR COMPLEX SUBUNIT 11; MED11","url":"https://www.omim.org/entry/612383"},{"mim_id":"610311","title":"MEDIATOR COMPLEX SUBUNIT 28; MED28","url":"https://www.omim.org/entry/610311"},{"mim_id":"609423","title":"HUMAN IMMUNODEFICIENCY VIRUS TYPE 1, SUSCEPTIBILITY TO","url":"https://www.omim.org/entry/609423"},{"mim_id":"600069","title":"URIDINE DIPHOSPHATE GLYCOSYLTRANSFERASE 2 FAMILY, MEMBER B15; UGT2B15","url":"https://www.omim.org/entry/600069"}],"hpa":{"profiled":true,"resolved_as":"","reliability":"Supported","locations":[{"location":"Nucleoplasm","reliability":"Supported"}],"tissue_specificity":"Low tissue specificity","tissue_distribution":"Detected in all","driving_tissues":[],"url":"https://www.proteinatlas.org/search/MED28"},"hgnc":{"alias_symbol":["EG1","DKFZP434N185","magicin"],"prev_symbol":[]},"alphafold":{"accession":"Q9H204","domains":[{"cath_id":"1.20.58","chopping":"43-149","consensus_level":"medium","plddt":93.4937,"start":43,"end":149}],"viewer_url":"https://alphafold.ebi.ac.uk/entry/Q9H204","model_url":"https://alphafold.ebi.ac.uk/files/AF-Q9H204-F1-model_v6.cif","pae_url":"https://alphafold.ebi.ac.uk/files/AF-Q9H204-F1-predicted_aligned_error_v6.png","plddt_mean":78.88},"mouse_models":{"mgi_url":"https://www.informatics.jax.org/marker/summary?nomen=MED28","jax_strain_url":"https://www.jax.org/strain/search?query=MED28"},"sequence":{"accession":"Q9H204","fasta_url":"https://rest.uniprot.org/uniprotkb/Q9H204.fasta","uniprot_url":"https://www.uniprot.org/uniprotkb/Q9H204/entry","alphafold_viewer_url":"https://alphafold.ebi.ac.uk/entry/Q9H204"}},"corpus_meta":[{"pmid":"1714386","id":"PMC_1714386","title":"A new human p34 protein kinase, CDK2, identified by complementation of a cdc28 mutation in Saccharomyces cerevisiae, is a homolog of Xenopus Eg1.","date":"1991","source":"The EMBO journal","url":"https://pubmed.ncbi.nlm.nih.gov/1714386","citation_count":278,"is_preprint":false},{"pmid":"1373987","id":"PMC_1373987","title":"Application of monoclonal antibodies against major basic protein (BMK-13) and eosinophil cationic protein (EG1 and EG2) for quantifying eosinophils in bronchial biopsies from atopic asthma.","date":"1992","source":"Clinical and experimental allergy : journal of the British Society for Allergy and Clinical Immunology","url":"https://pubmed.ncbi.nlm.nih.gov/1373987","citation_count":91,"is_preprint":false},{"pmid":"15467741","id":"PMC_15467741","title":"Magicin, a novel cytoskeletal protein associates with the NF2 tumor suppressor merlin and Grb2.","date":"2004","source":"Oncogene","url":"https://pubmed.ncbi.nlm.nih.gov/15467741","citation_count":51,"is_preprint":false},{"pmid":"21942447","id":"PMC_21942447","title":"Resveratrol modulates MED28 (Magicin/EG-1) expression and inhibits epidermal growth factor (EGF)-induced migration in MDA-MB-231 human breast cancer cells.","date":"2011","source":"Journal of agricultural and food chemistry","url":"https://pubmed.ncbi.nlm.nih.gov/21942447","citation_count":38,"is_preprint":false},{"pmid":"19830597","id":"PMC_19830597","title":"Determination of product inhibition of CBH1, CBH2, and EG1 using a novel cellulase activity assay.","date":"2009","source":"Applied biochemistry and biotechnology","url":"https://pubmed.ncbi.nlm.nih.gov/19830597","citation_count":24,"is_preprint":false},{"pmid":"11779215","id":"PMC_11779215","title":"Identification of a novel endothelial-derived gene EG-1.","date":"2002","source":"Biochemical and biophysical research communications","url":"https://pubmed.ncbi.nlm.nih.gov/11779215","citation_count":23,"is_preprint":false},{"pmid":"22495818","id":"PMC_22495818","title":"MED28 regulates MEK1-dependent cellular migration in human breast cancer cells.","date":"2012","source":"Journal of cellular physiology","url":"https://pubmed.ncbi.nlm.nih.gov/22495818","citation_count":23,"is_preprint":false},{"pmid":"17848560","id":"PMC_17848560","title":"Mediator subunit MED28 (Magicin) is a repressor of smooth muscle cell differentiation.","date":"2007","source":"The Journal of biological chemistry","url":"https://pubmed.ncbi.nlm.nih.gov/17848560","citation_count":22,"is_preprint":false},{"pmid":"10496315","id":"PMC_10496315","title":"Reactivity of monoclonal antibodies EG1 and EG2 with eosinophils and their granule proteins.","date":"1999","source":"Journal of leukocyte biology","url":"https://pubmed.ncbi.nlm.nih.gov/10496315","citation_count":22,"is_preprint":false},{"pmid":"15161708","id":"PMC_15161708","title":"Expression pattern of the novel gene EG-1 in cancer.","date":"2004","source":"Clinical cancer research : an official journal of the American Association for Cancer Research","url":"https://pubmed.ncbi.nlm.nih.gov/15161708","citation_count":21,"is_preprint":false},{"pmid":"16024617","id":"PMC_16024617","title":"The novel gene EG-1 stimulates cellular proliferation.","date":"2005","source":"Cancer research","url":"https://pubmed.ncbi.nlm.nih.gov/16024617","citation_count":19,"is_preprint":false},{"pmid":"30499104","id":"PMC_30499104","title":"MED28 and forkhead box M1 (FOXM1) mediate matrix metalloproteinase 2 (MMP2)-dependent cellular migration in human nonsmall cell lung cancer (NSCLC) cells.","date":"2018","source":"Journal of cellular 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one","url":"https://pubmed.ncbi.nlm.nih.gov/26445504","citation_count":14,"is_preprint":false},{"pmid":"27662245","id":"PMC_27662245","title":"MED28 Regulates Epithelial-Mesenchymal Transition Through NFκB in Human Breast Cancer Cells.","date":"2016","source":"Journal of cellular physiology","url":"https://pubmed.ncbi.nlm.nih.gov/27662245","citation_count":13,"is_preprint":false},{"pmid":"16964398","id":"PMC_16964398","title":"EG-1 interacts with c-Src and activates its signaling pathway.","date":"2006","source":"International journal of oncology","url":"https://pubmed.ncbi.nlm.nih.gov/16964398","citation_count":12,"is_preprint":false},{"pmid":"16899217","id":"PMC_16899217","title":"Magicin associates with the Src-family kinases and is phosphorylated upon CD3 stimulation.","date":"2006","source":"Biochemical and biophysical research 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Shortens the Cell Cycle and Induces Genomic Instability.","date":"2019","source":"International journal of molecular sciences","url":"https://pubmed.ncbi.nlm.nih.gov/30970566","citation_count":8,"is_preprint":false},{"pmid":"32306431","id":"PMC_32306431","title":"RCOR1 directly binds to MED28 and weakens its inducing effect on cancer stem cell-like activity of oral cavity squamous cell carcinoma cells.","date":"2020","source":"Journal of oral pathology & medicine : official publication of the International Association of Oral Pathologists and the American Academy of Oral Pathology","url":"https://pubmed.ncbi.nlm.nih.gov/32306431","citation_count":8,"is_preprint":false},{"pmid":"36072467","id":"PMC_36072467","title":"Bioactive Vitamin D Attenuates MED28-Mediated Cell Growth and Epithelial-Mesenchymal Transition in Human Colorectal Cancer Cells.","date":"2022","source":"BioMed research 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Pulp.","date":"2025","source":"Polymers","url":"https://pubmed.ncbi.nlm.nih.gov/40574241","citation_count":1,"is_preprint":false},{"pmid":"11477881","id":"PMC_11477881","title":"[The changes of EG1 and EG2 positive eosinophils and their clinical significance in asthmatics].","date":"1998","source":"Zhonghua jie he he hu xi za zhi = Zhonghua jiehe he huxi zazhi = Chinese journal of tuberculosis and respiratory diseases","url":"https://pubmed.ncbi.nlm.nih.gov/11477881","citation_count":0,"is_preprint":false},{"pmid":"34722257","id":"PMC_34722257","title":"Chemical Synthesis of the PAX Protein Inhibitor EG1 and Its Ability to Slow the Growth of Human Colorectal Carcinoma Cells.","date":"2021","source":"Frontiers in oncology","url":"https://pubmed.ncbi.nlm.nih.gov/34722257","citation_count":0,"is_preprint":false},{"pmid":"29871732","id":"PMC_29871732","title":"Valproic Acid Decreases the Nuclear Localization of MDT-28, the Nematode Orthologue of MED28.","date":"2018","source":"Folia biologica","url":"https://pubmed.ncbi.nlm.nih.gov/29871732","citation_count":0,"is_preprint":false}],"cost":{"stage1":{"model":"claude-sonnet-4-6","input_tokens":16120,"output_tokens":3993,"usd":0.054128},"stage2":{"model":"claude-opus-4-6","input_tokens":7456,"output_tokens":3585,"usd":0.190358},"total_usd":0.244486,"stage1_batch_id":"msgbatch_01Mgbec4vM6c9vXht7JVJzss","stage2_batch_id":"msgbatch_01JkuuH5RB2BdJNsgnhjbC6s","note":"batch pricing = 50% of standard"},"stage1_raw":"```json\n{\n  \"discoveries\": [\n    {\n      \"year\": 2004,\n      \"finding\": \"MED28 (magicin) was identified as a merlin-specific binding partner that interacts with the NF2 tumor suppressor merlin in vitro and in vivo, colocalizes with merlin beneath the plasma membrane, and associates with the actin cytoskeleton as determined by cofractionation, immunofluorescence, and electron microscopy. MED28 also interacts with the adaptor protein Grb2 via its Grb2-binding motifs, and merlin can form a ternary complex with MED28 and Grb2.\",\n      \"method\": \"Co-immunoprecipitation, affinity binding, blot overlay, immunofluorescence, electron microscopy, subcellular fractionation\",\n      \"journal\": \"Oncogene\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 — multiple orthogonal methods (Co-IP, pulldown, blot overlay, EM, fractionation) in a single study\",\n      \"pmids\": [\"15467741\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2006,\n      \"finding\": \"MED28 (magicin) interacts with Fyn tyrosine kinase (identified by yeast two-hybrid), and Fyn phosphorylates magicin in vitro. Src and Lck also interact with magicin. Upon CD3 stimulation in Jurkat cells, magicin is phosphorylated by Lck (the major kinase, as shown by Lck-deficient J.CaM1.6 cells lacking phosphorylation). Site-directed mutagenesis and in vitro kinase assays identified Y64 as the Lck phosphorylation site, creating an SH2-Grb2 binding motif.\",\n      \"method\": \"Yeast two-hybrid, in vitro kinase assay, site-directed mutagenesis, co-immunoprecipitation, Lck-deficient cell line\",\n      \"journal\": \"Biochemical and biophysical research communications\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1–2 — in vitro kinase assay plus mutagenesis plus genetic validation with Lck-deficient cell line\",\n      \"pmids\": [\"16899217\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2006,\n      \"finding\": \"EG-1 (MED28) overexpression activates c-Src signaling; EG-1 interacts with c-Src (by co-immunoprecipitation) and also binds other Src-family members as well as multiple SH3- and WW-domain-containing signaling molecules, but EG-1 is not a direct Src substrate.\",\n      \"method\": \"Co-immunoprecipitation, overexpression experiments\",\n      \"journal\": \"International journal of oncology\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 3 — single Co-IP plus overexpression, no mutagenesis or reconstitution\",\n      \"pmids\": [\"16964398\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2005,\n      \"finding\": \"EG-1 (MED28) overexpression stimulates cellular proliferation in vitro and in vivo (xenograft model), and is associated with activation of ERK1/2, JNK, and p38 MAPK kinases; co-immunoprecipitation showed an association between EG-1 and Src.\",\n      \"method\": \"Overexpression/siRNA transfection, xenograft assay, co-immunoprecipitation, immunoblot\",\n      \"journal\": \"Cancer research\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 3 — functional proliferation assay plus Co-IP, but no direct mechanistic dissection of pathway\",\n      \"pmids\": [\"16024617\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2007,\n      \"finding\": \"MED28 functions as a subunit of the mammalian Mediator complex and acts as a repressor of smooth muscle cell (SMC) differentiation. Knockdown of Med28 in NIH3T3 cells induces SMC differentiation genes; overexpression represses them. Med28 functions within the Mediator head module together with Med6, Med8, and Med18, and may act as a scaffolding protein maintaining this submodule's stability.\",\n      \"method\": \"siRNA knockdown, overexpression, gene expression analysis, genetic epistasis with Med6/Med8/Med18\",\n      \"journal\": \"The Journal of biological chemistry\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 — clean KD and OE with defined cellular phenotype, epistasis with head-module subunits, replicated bidirectionally\",\n      \"pmids\": [\"17848560\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2011,\n      \"finding\": \"MED28 overexpression enhances EGF-induced cellular migration in MDA-MB-231 breast cancer cells, and this effect is mediated through the EGFR/PI3K signaling pathway; resveratrol suppresses EGF-mediated migration by reducing MED28 and MMP-9 expression.\",\n      \"method\": \"Overexpression, migration assay, pharmacological inhibition, immunoblot\",\n      \"journal\": \"Journal of agricultural and food chemistry\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 3 — overexpression with functional migration assay, pathway inferred pharmacologically\",\n      \"pmids\": [\"21942447\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2012,\n      \"finding\": \"MED28 regulates cellular migration and invasion in human breast cancer cells in a MEK1-dependent manner: MED28 knockdown reduces MEK1 and MMP2 expression; MEK1 suppression blocks MED28-induced MMP2 activation and migration; ectopic MEK1 rescues MED28 knockdown-mediated invasion suppression; and exogenous MMP2 rescues invasion upon MED28 or MEK1 knockdown.\",\n      \"method\": \"siRNA knockdown, dominant-negative construct, MEK1 inhibitors, rescue experiments with MEK1 cDNA and recombinant MMP2, migration/invasion assay\",\n      \"journal\": \"Journal of cellular physiology\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 — epistasis by multiple genetic approaches (siRNA, dominant-negative, rescue) with defined pathway order\",\n      \"pmids\": [\"22495818\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2015,\n      \"finding\": \"Med28 is essential for mouse peri-implantation development; knockout causes lethality due to loss of inner cell mass pluripotency with reduced Oct4 and Nanog expression. Overexpression of Med28 in mouse embryonic fibroblasts enhances reprogramming efficiency to iPSCs. Cre-mediated inactivation in iPSCs demonstrates Med28 is required for their survival. MED28 has both cytosolic (merlin-interacting) and nuclear (Mediator complex) roles.\",\n      \"method\": \"Knockout mouse model, Cre-mediated conditional inactivation, reprogramming assay, gene expression analysis\",\n      \"journal\": \"PloS one\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 — clean genetic KO with defined developmental phenotype and molecular markers, plus reprogramming functional assay\",\n      \"pmids\": [\"26445504\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2015,\n      \"finding\": \"MED28 knockdown in colorectal cancer cells reduces cyclin D1, c-Myc, and nuclear β-catenin expression, while increasing E-cadherin and HBP1 (a negative Wnt/β-catenin regulator); MED28 suppresses HBP1 promoter activity (luciferase reporter assay), and overexpression has the opposite effects, placing MED28 upstream of Wnt/β-catenin signaling via HBP1.\",\n      \"method\": \"siRNA knockdown, overexpression, luciferase reporter assay, immunoblot\",\n      \"journal\": \"Journal of cellular physiology\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2–3 — reporter assay plus KD/OE with defined pathway placement, single lab\",\n      \"pmids\": [\"26660958\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2016,\n      \"finding\": \"MED28 regulates epithelial-mesenchymal transition (EMT) in human breast cancer cells through the NFκB pathway: MED28 suppression attenuates mesenchymal markers and reduces p-NFκB/p65 and Snail; overexpression has the opposite effect. In MCF7 cells, adriamycin-induced EMT correlates with increased MED28 and p-p65, and MED28 knockdown reverses this EMT.\",\n      \"method\": \"siRNA knockdown, overexpression, immunoblot, morphological analysis, pharmacological EMT induction\",\n      \"journal\": \"Journal of cellular physiology\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 3 — KD/OE with defined phenotype and pathway marker, single lab without reconstitution\",\n      \"pmids\": [\"27662245\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2017,\n      \"finding\": \"MED28 interacts with ZNF224 in the nucleus (confirmed by co-immunoprecipitation and surface plasmon resonance); the KRAB domain of ZNF224 interacts with the MED domain of MED28. MED28 overexpression stabilizes ZNF224 against camptothecin-induced degradation, resulting in increased colony formation of MCF-7 cells.\",\n      \"method\": \"Co-immunoprecipitation, surface plasmon resonance, bimolecular fluorescence complementation, overexpression, colony formation assay\",\n      \"journal\": \"Oncology letters\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 — SPR plus Co-IP plus BiFC provide orthogonal binding evidence; functional consequence demonstrated\",\n      \"pmids\": [\"29435049\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2018,\n      \"finding\": \"MED28 interacts with FOXM1 (co-immunoprecipitation); MED28 and FOXM1 mutually affect each other's expression and subcellular localization. Both regulate MMP2-dependent migration and invasion in NSCLC cells; MED28 siRNA-mediated MMP2 suppression is rescued by constitutively active FOXM1, restoring migration and invasion.\",\n      \"method\": \"Co-immunoprecipitation, siRNA knockdown, doxycycline-inducible overexpression, rescue experiment, migration/invasion assay\",\n      \"journal\": \"Journal of cellular physiology\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2–3 — Co-IP plus rescue epistasis, single lab\",\n      \"pmids\": [\"30499104\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2019,\n      \"finding\": \"MED28 expression is regulated by transcription factors E2F1, NRF1, ETS1, and C/EBPβ (identified by luciferase reporter assay). MED28 expression peaks at the G1-S transition and mitosis. Overexpression of MED28 shortens interphase and mitosis duration, whereas knockdown lengthens them. MED28 overexpression increases micronuclei, nuclear budding, and aneuploidy, indicating a role in maintaining genomic stability.\",\n      \"method\": \"Luciferase reporter assay, cell cycle synchronization, live cell imaging, flow cytometry, fluorescence microscopy, siRNA knockdown, overexpression\",\n      \"journal\": \"International journal of molecular sciences\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 — multiple orthogonal methods, single lab\",\n      \"pmids\": [\"30970566\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2020,\n      \"finding\": \"RCOR1 directly interacts with MED28 (demonstrated in OCSCC cells) and overexpression of RCOR1 abrogates MED28-induced cancer stem cell-like activity (colony/sphere formation and CSC marker expression), placing RCOR1 as a direct negative regulator of MED28 function.\",\n      \"method\": \"Co-immunoprecipitation (direct binding), overexpression, functional CSC assays\",\n      \"journal\": \"Journal of oral pathology & medicine\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 3 — single Co-IP with functional rescue, single lab\",\n      \"pmids\": [\"32306431\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2018,\n      \"finding\": \"MDT-28, the C. elegans orthologue of MED28, undergoes lysine acetylation (confirmed by anti-acetyl-lysine immunoprecipitation of GFP::MDT-28). Valproic acid (a lysine deacetylase inhibitor) enhances MDT-28 acetylation and decreases its nuclear localization as detected by FLIM, indicating that the nuclear pool of MED28 is regulated by acetylation.\",\n      \"method\": \"Immunoprecipitation with anti-acetyl-lysine antibody, fluorescence lifetime imaging microscopy (FLIM), pharmacological treatment with VPA\",\n      \"journal\": \"Folia biologica\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 — direct biochemical detection of PTM plus subcellular localization change in a model organism orthologue\",\n      \"pmids\": [\"29871732\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2024,\n      \"finding\": \"MED28 knockdown in hepatocellular carcinoma cells induces cell cycle arrest and suppresses AKT/mTOR signaling, reduces lipid accumulation, and decreases expression and nuclear localization of SREBP1; MED28 overexpression upregulates AKT/mTOR signaling, placing MED28 upstream of this pathway in liver cancer.\",\n      \"method\": \"siRNA knockdown, overexpression, immunoblot, subcellular fractionation, cell cycle analysis\",\n      \"journal\": \"Journal of agricultural and food chemistry\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2–3 — bidirectional KD/OE with defined pathway readouts, single lab\",\n      \"pmids\": [\"38619972\"],\n      \"is_preprint\": false\n    }\n  ],\n  \"current_model\": \"MED28 is a dual-compartment protein that functions as a subunit of the mammalian Mediator transcriptional co-activator complex in the nucleus—where it acts as part of the head module (with Med6, Med8, Med18) to repress smooth muscle cell differentiation, support pluripotency, and regulate cell cycle progression—while in the cytoplasm it associates with the actin cytoskeleton as a merlin/Grb2-interacting scaffold (magicin) that is phosphorylated at Y64 by Src-family kinases (principally Lck/Fyn) upon receptor stimulation, and interacts with signaling partners including FOXM1 and ZNF224 to regulate MEK1–MMP2-dependent cell migration, EMT via NFκB, Wnt/β-catenin signaling via HBP1, and AKT/mTOR-dependent proliferation in cancer cells.\"\n}\n```","stage2_raw":"```json\n{\n  \"mechanistic_narrative\": \"MED28 is a dual-compartment scaffolding protein that functions both as a subunit of the Mediator transcriptional co-activator complex and as a cytoplasmic signaling adaptor linking receptor-proximal kinases to downstream proliferative and migratory pathways. Within the nucleus, MED28 operates in the Mediator head module together with Med6, Med8, and Med18 to repress smooth muscle cell differentiation, maintain pluripotency through Oct4/Nanog expression, and regulate cell cycle progression at the G1–S transition and mitosis [PMID:17848560, PMID:26445504, PMID:30970566]. In the cytoplasm, MED28 associates with the actin cytoskeleton and the tumor suppressor merlin (NF2), is phosphorylated at Y64 by Src-family kinases (principally Lck) to create an SH2-binding motif for Grb2, and promotes cell migration and invasion through a MEK1–MMP2 axis and NF-κB–dependent EMT [PMID:15467741, PMID:16899217, PMID:22495818, PMID:27662245]. MED28 also activates Wnt/β-catenin signaling by suppressing the negative regulator HBP1 and promotes AKT/mTOR-dependent proliferation and lipogenesis in cancer cells [PMID:26660958, PMID:38619972].\",\n  \"teleology\": [\n    {\n      \"year\": 2004,\n      \"claim\": \"Establishing that MED28 is a cytoplasmic scaffolding protein that physically links the NF2 tumor suppressor merlin to the adaptor Grb2 at the cortical actin cytoskeleton resolved the question of how merlin connects to growth factor signaling complexes.\",\n      \"evidence\": \"Co-IP, affinity binding, blot overlay, EM, and subcellular fractionation in mammalian cells\",\n      \"pmids\": [\"15467741\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\n        \"Functional consequence of merlin–MED28–Grb2 ternary complex on downstream signaling not determined\",\n        \"Whether the cytoplasmic pool of MED28 is stoichiometrically distinct from the Mediator-bound nuclear pool was unknown\"\n      ]\n    },\n    {\n      \"year\": 2005,\n      \"claim\": \"Demonstrating that MED28 overexpression drives proliferation in vitro and in xenografts while activating ERK1/2, JNK, and p38 MAPK established MED28 as a pro-proliferative factor acting upstream of multiple MAPK cascades.\",\n      \"evidence\": \"Overexpression and siRNA in cancer cell lines, xenograft model, Co-IP with Src, immunoblot\",\n      \"pmids\": [\"16024617\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\n        \"No direct substrate or kinase–substrate relationship defined for MED28 in MAPK activation\",\n        \"Correlation between Src interaction and MAPK activation not causally dissected\"\n      ]\n    },\n    {\n      \"year\": 2006,\n      \"claim\": \"Identification of Y64 as the Lck phosphorylation site on MED28—using Lck-deficient cells and site-directed mutagenesis—revealed how TCR stimulation creates an SH2 docking site for Grb2 on MED28, providing a direct biochemical mechanism for signal-dependent scaffold assembly.\",\n      \"evidence\": \"Yeast two-hybrid, in vitro kinase assay, mutagenesis, Lck-deficient J.CaM1.6 cells\",\n      \"pmids\": [\"16899217\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\n        \"Downstream signaling events specifically dependent on pY64-Grb2 recruitment not mapped\",\n        \"Whether Y64 phosphorylation occurs in non-hematopoietic contexts not tested\"\n      ]\n    },\n    {\n      \"year\": 2007,\n      \"claim\": \"Showing that MED28 is a bona fide subunit of the Mediator head module (with Med6/Med8/Med18) that represses smooth muscle differentiation genes established its nuclear transcriptional role, distinct from its cytoplasmic scaffolding function.\",\n      \"evidence\": \"siRNA knockdown, overexpression, genetic epistasis with head-module subunits in NIH3T3 cells\",\n      \"pmids\": [\"17848560\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\n        \"Structural basis for MED28 integration into the head module not resolved\",\n        \"Mechanism by which head module occupancy at SMC gene promoters leads to repression not determined\"\n      ]\n    },\n    {\n      \"year\": 2012,\n      \"claim\": \"Epistatic rescue experiments placing MED28 upstream of MEK1 and MMP2 in breast cancer cell migration/invasion defined a linear signaling hierarchy (MED28→MEK1→MMP2) through which MED28 promotes invasiveness.\",\n      \"evidence\": \"siRNA, dominant-negative constructs, MEK1 cDNA rescue, recombinant MMP2 rescue, migration/invasion assays in breast cancer cells\",\n      \"pmids\": [\"22495818\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\n        \"Direct biochemical link between MED28 and MEK1 activation (e.g., Raf-level regulation) not identified\",\n        \"Whether this pathway operates through cytoplasmic or nuclear MED28 not distinguished\"\n      ]\n    },\n    {\n      \"year\": 2015,\n      \"claim\": \"Genetic knockout in mice and conditional inactivation in iPSCs demonstrated that MED28 is essential for peri-implantation development and pluripotency maintenance via Oct4/Nanog, and that it enhances somatic reprogramming—establishing a non-redundant in vivo requirement.\",\n      \"evidence\": \"Knockout mouse model, Cre-mediated conditional inactivation, iPSC reprogramming assay, gene expression analysis\",\n      \"pmids\": [\"26445504\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\n        \"Whether the developmental requirement is mediated by Mediator function, cytoplasmic signaling, or both is unresolved\",\n        \"Tissue-specific conditional knockouts beyond peri-implantation not reported\"\n      ]\n    },\n    {\n      \"year\": 2015,\n      \"claim\": \"Demonstrating that MED28 suppresses HBP1 promoter activity to activate Wnt/β-catenin target genes (cyclin D1, c-Myc) in colorectal cancer cells identified a transcriptional mechanism linking MED28 to Wnt pathway output.\",\n      \"evidence\": \"siRNA knockdown, overexpression, luciferase reporter assay, immunoblot in colorectal cancer cells\",\n      \"pmids\": [\"26660958\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\n        \"Whether MED28 represses HBP1 directly at its promoter via Mediator or indirectly through an intermediary not resolved\",\n        \"In vivo relevance of this axis not tested\"\n      ]\n    },\n    {\n      \"year\": 2016,\n      \"claim\": \"Linking MED28 to EMT through NF-κB/p65 phosphorylation and Snail expression in breast cancer cells expanded its oncogenic repertoire beyond proliferation and migration to include phenotypic plasticity.\",\n      \"evidence\": \"siRNA knockdown, overexpression, adriamycin-induced EMT model, immunoblot in MCF7 and MDA-MB-231 cells\",\n      \"pmids\": [\"27662245\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\n        \"Direct mechanism by which MED28 activates NF-κB (e.g., IKK regulation) not identified\",\n        \"Single-lab finding without independent replication\"\n      ]\n    },\n    {\n      \"year\": 2017,\n      \"claim\": \"Identification of ZNF224 as a direct nuclear interaction partner of MED28—with binding mapped to the KRAB domain of ZNF224 and the MED domain of MED28—revealed a mechanism by which MED28 stabilizes a transcriptional repressor to promote colony formation.\",\n      \"evidence\": \"Co-IP, surface plasmon resonance, bimolecular fluorescence complementation, colony assay in MCF-7 cells\",\n      \"pmids\": [\"29435049\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\n        \"Genomic targets co-regulated by MED28–ZNF224 not identified\",\n        \"Whether ZNF224 stabilization requires Mediator association is unknown\"\n      ]\n    },\n    {\n      \"year\": 2018,\n      \"claim\": \"Showing that the C. elegans orthologue MDT-28 undergoes lysine acetylation that reduces its nuclear localization revealed a conserved post-translational mechanism governing the cytoplasmic-to-nuclear partitioning of MED28.\",\n      \"evidence\": \"Anti-acetyl-lysine immunoprecipitation of GFP::MDT-28, FLIM imaging, VPA treatment in C. elegans\",\n      \"pmids\": [\"29871732\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\n        \"Specific acetylated lysine residues not mapped\",\n        \"Whether mammalian MED28 is similarly regulated by acetylation not tested\",\n        \"Identity of the acetyltransferase and deacetylase acting on MDT-28/MED28 unknown\"\n      ]\n    },\n    {\n      \"year\": 2018,\n      \"claim\": \"Demonstrating that MED28 and FOXM1 mutually regulate each other's expression and co-drive MMP2-dependent invasion provided evidence of a feed-forward loop in NSCLC that converges on the same MMP2 effector pathway identified in breast cancer.\",\n      \"evidence\": \"Co-IP, siRNA, doxycycline-inducible overexpression, FOXM1 rescue of MED28 knockdown, invasion assay in NSCLC cells\",\n      \"pmids\": [\"30499104\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\n        \"Whether MED28–FOXM1 interaction occurs within the Mediator complex or independently not distinguished\",\n        \"Mechanism of mutual transcriptional regulation not resolved\"\n      ]\n    },\n    {\n      \"year\": 2019,\n      \"claim\": \"Establishing that MED28 expression is cell-cycle-regulated (peaking at G1–S and mitosis) and that its overexpression shortens both interphase and mitosis while increasing aneuploidy connected MED28 to genomic stability and cell-cycle timing.\",\n      \"evidence\": \"Cell cycle synchronization, live cell imaging, flow cytometry, fluorescence microscopy, luciferase reporter for promoter analysis\",\n      \"pmids\": [\"30970566\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\n        \"Molecular targets through which MED28 accelerates cell cycle phases not identified\",\n        \"Whether aneuploidy results from Mediator-dependent transcriptional mis-regulation or cytoskeletal defects is unclear\"\n      ]\n    },\n    {\n      \"year\": 2024,\n      \"claim\": \"Placing MED28 upstream of AKT/mTOR signaling and SREBP1-dependent lipogenesis in hepatocellular carcinoma cells broadened its oncogenic functions to include metabolic reprogramming.\",\n      \"evidence\": \"siRNA knockdown, overexpression, immunoblot, subcellular fractionation, cell cycle analysis in HCC cells\",\n      \"pmids\": [\"38619972\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\n        \"Direct biochemical mechanism linking MED28 to AKT phosphorylation not determined\",\n        \"In vivo validation in liver cancer models not reported\"\n      ]\n    },\n    {\n      \"year\": null,\n      \"claim\": \"The mechanism governing MED28 partitioning between its Mediator-associated nuclear pool and its cytoplasmic actin-associated pool, and whether these represent functionally independent or coordinated activities, remains unresolved.\",\n      \"evidence\": \"\",\n      \"pmids\": [],\n      \"confidence\": \"Low\",\n      \"gaps\": [\n        \"No structural data for MED28 within the Mediator head module at sufficient resolution to define contacts\",\n        \"Regulatory signals controlling nuclear–cytoplasmic shuttling in mammalian cells not identified\",\n        \"Whether cytoplasmic signaling functions of MED28 feed back into its Mediator-dependent transcriptional activity is untested\"\n      ]\n    }\n  ],\n  \"mechanism_profile\": {\n    \"molecular_activity\": [\n      {\"term_id\": \"GO:0060090\", \"supporting_discovery_ids\": [0, 1, 10, 11]},\n      {\"term_id\": \"GO:0140110\", \"supporting_discovery_ids\": [4, 7, 8]},\n      {\"term_id\": \"GO:0098772\", \"supporting_discovery_ids\": [4, 6, 9]}\n    ],\n    \"localization\": [\n      {\"term_id\": \"GO:0005856\", \"supporting_discovery_ids\": [0]},\n      {\"term_id\": \"GO:0005634\", \"supporting_discovery_ids\": [4, 7, 10, 14]},\n      {\"term_id\": \"GO:0005829\", \"supporting_discovery_ids\": [0, 1]}\n    ],\n    \"pathway\": [\n      {\"term_id\": \"R-HSA-74160\", \"supporting_discovery_ids\": [4, 7, 8]},\n      {\"term_id\": \"R-HSA-162582\", \"supporting_discovery_ids\": [1, 3, 6, 9, 15]},\n      {\"term_id\": \"R-HSA-1640170\", \"supporting_discovery_ids\": [12]},\n      {\"term_id\": \"R-HSA-1266738\", \"supporting_discovery_ids\": [4, 7]}\n    ],\n    \"complexes\": [\n      \"Mediator complex (head module)\"\n    ],\n    \"partners\": [\n      \"NF2\",\n      \"GRB2\",\n      \"LCK\",\n      \"FYN\",\n      \"MED6\",\n      \"FOXM1\",\n      \"ZNF224\",\n      \"RCOR1\"\n    ],\n    \"other_free_text\": []\n  }\n}\n```"}