{"gene":"MED6","run_date":"2026-06-10T02:59:50","timeline":{"discoveries":[{"year":1997,"finding":"Med6 is an essential subunit of the yeast Mediator complex required for transcriptional activation from many class II promoters in vivo; a temperature-sensitive Med6 mutant abolishes inducible transcription without affecting basal or constitutive transcription, and Mediator isolated from this mutant is temperature-sensitive for activation in a reconstituted in vitro system due to a defect in initiation complex formation.","method":"Temperature-sensitive mutant analysis, in vivo transcription assays, reconstituted in vitro transcription system","journal":"Molecular and cellular biology","confidence":"High","confidence_rationale":"Tier 1 / Strong — in vitro reconstitution combined with in vivo genetic analysis, multiple promoters tested, functional defect clearly mapped to initiation complex formation","pmids":["9234719"],"is_preprint":false},{"year":1998,"finding":"Mouse Mediator contains homologs of yeast Mediator subunits including Med6, binds to the RNA polymerase II C-terminal domain (CTD), and stimulates phosphorylation of the CTD by TFIIH.","method":"Biochemical purification, peptide sequencing, CTD binding assay, CTD phosphorylation assay","journal":"Proceedings of the National Academy of Sciences of the United States of America","confidence":"High","confidence_rationale":"Tier 1 / Moderate — in vitro biochemical assays (CTD binding and TFIIH-stimulated phosphorylation) with peptide sequencing confirming subunit identity, single lab","pmids":["9671713"],"is_preprint":false},{"year":1998,"finding":"A functional interaction between Med6 and Srb4 is required for transcriptional activation; an allele-specific dominant suppressor screen identified SRB4 as a suppressor of a med6-ts mutation, and biochemical fractionation showed Med6 and Srb4 co-reside in the same Mediator subcomplex.","method":"Genetic suppressor screen, biochemical Mediator subcomplex fractionation, in vivo transcription assays","journal":"Molecular and cellular biology","confidence":"High","confidence_rationale":"Tier 2 / Moderate — reciprocal genetic epistasis (allele-specific suppression) combined with biochemical co-fractionation, multiple orthogonal methods in one study","pmids":["9710620"],"is_preprint":false},{"year":1998,"finding":"Human NAT complex, containing the human homolog of Med6 along with Srb7, Srb10, Srb11, and Rgr1, represses activated transcription and phosphorylates the CTD of RNA polymerase II at residues distinct from those phosphorylated by TFIIH; the complex interacts with RNAPII in a CTD-independent manner, and CTD phosphorylation precludes this interaction.","method":"Biochemical purification, in vitro transcription repression assay, CTD phosphorylation assay, co-immunoprecipitation with RNAPII","journal":"Molecular cell","confidence":"High","confidence_rationale":"Tier 1 / Moderate — multiple in vitro functional assays (repression, kinase, binding) with defined biochemical dissection, single lab","pmids":["9734358"],"is_preprint":false},{"year":1999,"finding":"Human SMCC complex, containing MED6 along with other SRB/MED homologs, can either repress activator-dependent transcription or, at limiting TFIIH, synergistically enhance it; the complex shows direct activator interactions and can act independently of the RNA polymerase II CTD.","method":"Biochemical purification, in vitro transcription assay, activator interaction assay","journal":"Molecular cell","confidence":"High","confidence_rationale":"Tier 1 / Moderate — in vitro reconstituted transcription assays demonstrating both positive and negative activities, CTD-independence tested directly, single lab","pmids":["10024883"],"is_preprint":false},{"year":1998,"finding":"Med6 and Srb6, components of the RNA polymerase II holoenzyme, are dominant suppressors of a temperature-sensitive Srb4 mutation and physically interact with Srb4, placing Med6 within the holoenzyme where it participates in the balance between transcriptional activation and repression.","method":"Genetic suppressor screen, physical interaction assay (co-immunoprecipitation), in vivo transcription analysis","journal":"Molecular and cellular biology","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — genetic epistasis combined with physical interaction data, but details of interaction assay not fully described in abstract","pmids":["9671455"],"is_preprint":false},{"year":1999,"finding":"Med6 is required for all activator-specific transcriptional activation processes tested (Bas1/Bas2-, Gcn4-, Gal4-, Rap1-mediated), whereas other Mediator subunits (Med9/Cse2, Med10/Nut2, Gal11, Med11) show activator-specific requirements; this positions Med6 as a convergence point where diverse activation signals meet to modulate Pol II activity.","method":"Differential display, Northern analysis of mRNA from Mediator mutant strains, genetic epistasis","journal":"Molecular and cellular biology","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — multiple activator systems tested with defined genetic mutants, single lab, primarily in vivo transcription readouts","pmids":["9891034"],"is_preprint":false},{"year":2001,"finding":"Drosophila Med6 (dMed6) is essential for viability and cell proliferation; dMed6 mutants fail to pupate and die in the third larval instar with severe proliferation defects in imaginal discs; cDNA microarray and quantitative RT-PCR show that transcriptional activation of many, but not all, genes is affected, including genes involved in cell proliferation and metabolism.","method":"Genetic loss-of-function mutant analysis, cDNA microarray, quantitative RT-PCR, in situ expression analysis","journal":"Molecular and cellular biology","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — genetic KO with specific phenotypic readout, genome-wide transcription profiling, multiple orthogonal methods, single lab","pmids":["11438678"],"is_preprint":false},{"year":2001,"finding":"C. elegans med-6 is required for development; RNAi of med-6 suppresses gain-of-function phenotypes of Ras pathway components (let-23 and let-60) and enhances loss-of-function of lin-3, placing Med-6 as required for transcriptional output of the Ras signaling pathway; med-6 is also involved in Wnt pathway-dependent male ray development.","method":"Genetic mutation analysis, RNAi epistasis with Ras and Wnt pathway alleles, phenotypic readout of signaling pathway outputs","journal":"Development (Cambridge, England)","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — genetic epistasis with defined pathway alleles across two signaling pathways, single lab","pmids":["11688559"],"is_preprint":false},{"year":2005,"finding":"MED7·MED21 (Med7·Srb7) crystal structure at 3.0 Å reveals that MED6 bridges the Mediator middle module to the head module; MED6 binds to putative protein-binding sites on the MED7·MED21 heterodimer surface, and a flexible MED6 bridge together with the MED7·MED21 hinge may account for conformational changes in Mediator upon binding to Pol II or activators.","method":"X-ray crystallography (3.0 Å), structural analysis of protein-binding surfaces","journal":"The Journal of biological chemistry","confidence":"High","confidence_rationale":"Tier 1 / Moderate — atomic-resolution crystal structure with functional interpretation of MED6 binding sites, single lab but high-resolution structural evidence","pmids":["15710619"],"is_preprint":false},{"year":2005,"finding":"In C. elegans, LET-425/MED6 forms a complex in vivo with LET-19/MED13, DPY-22/MED12, and SUR-2/MED23 (components of the Mediator complex), and this complex is required for Wnt-regulated asymmetric T-cell division and Wnt target gene repression.","method":"Co-immunoprecipitation in vivo, lineage analysis of let-19/dpy-22 mutants, epistasis with Wnt pathway components","journal":"Development (Cambridge, England)","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — reciprocal Co-IP demonstrating complex assembly combined with genetic epistasis and lineage analysis, single lab","pmids":["15790964"],"is_preprint":false},{"year":2006,"finding":"Med21 interacts with Med6 (confirmed by genetic and 2-hybrid data), and Med21 serves as a molecular switchboard integrating signals within the Mediator middle domain; interactions between Med21 and Med6 suggest Med6 is connected to the middle module through Med21.","method":"Two-hybrid analysis, co-immunoprecipitation of tagged proteins in insect cells and E. coli, high-copy suppressor screen","journal":"Molecular genetics and genomics","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — multiple interaction methods (2-hybrid + Co-IP in two heterologous systems), but functional consequences of Med6-Med21 interaction not directly tested in this study","pmids":["16758199"],"is_preprint":false},{"year":2007,"finding":"A 600-kDa MED6-containing subcomplex (TMLC3) augments transcriptional activation in vitro but lacks CDK8 and does not phosphorylate RNA Pol II; by contrast, the full 1.5-MDa complex (TMLC1) containing MED6, MED7, and CDK8 both activates transcription and phosphorylates Pol II, and preferentially interacts with TFIIE, TFIIF, and TFIIH.","method":"Affinity purification and HPLC-gel filtration of epitope-tagged MED6/MED7/CDK8 complexes, in vitro transcription assay, Pol II phosphorylation assay, co-immunoprecipitation with general transcription factors","journal":"Genes to cells","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — multiple biochemical assays (purification, transcription, kinase, interaction) from a defined subcomplex, single lab","pmids":["17212659"],"is_preprint":false},{"year":2007,"finding":"MED28 functions as a negative regulator of smooth muscle cell differentiation in concert with Med6, Med8, and Med18 within the Mediator head module; knockdown and overexpression experiments show Med6-containing head module components act together to suppress smooth muscle cell gene expression.","method":"siRNA knockdown, overexpression in NIH3T3 cells, gene expression analysis, mesenchymal precursor transdifferentiation assay","journal":"The Journal of biological chemistry","confidence":"Low","confidence_rationale":"Tier 3 / Weak — Med6 involvement inferred from co-knockdown phenotype without direct mechanistic dissection of Med6's specific contribution; single lab, no direct Med6-specific assay","pmids":["17848560"],"is_preprint":false},{"year":2010,"finding":"MED6 is recruited to the enhancer and proximal promoter of the PPARγ target gene Fabp4 in a MED1-independent manner, as shown by ChIP; MED14 knockdown reduces MED6 occupancy at the Fabp4 proximal promoter without affecting its binding at the enhancer, indicating MED14 is required for functional Mediator recruitment and positioning of MED6 at proximal promoters.","method":"ChIP (chromatin immunoprecipitation), siRNA knockdown of MED1 and MED14, reporter and endogenous gene expression assays","journal":"Molecular and cellular biology","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — ChIP-based localization with functional knockdown, multiple siRNA targets tested, single lab","pmids":["20194623"],"is_preprint":false},{"year":2012,"finding":"Crystal structure of the S. pombe Mediator head module at 3.4 Å reveals Med6 as one of seven head module subunits (with Med8, Med11, Med17, Med18, Med20, Med22); Med6 contributes to the fixed jaw submodule and is positioned within the head module architecture that contacts Pol II and its CTD.","method":"X-ray crystallography (3.4 Å), structural analysis","journal":"Nature","confidence":"High","confidence_rationale":"Tier 1 / Strong — high-resolution crystal structure, replicated across S. cerevisiae and S. pombe, structural position of Med6 in head module well-defined","pmids":["23123849"],"is_preprint":false},{"year":2014,"finding":"MED6 knockdown by siRNA significantly impairs HIV-1 replication at a post-integration step by inhibiting early HIV transcripts, specifically affecting transcription of the nascent viral mRNA transactivation-responsive element (TAR); MED6 knockdown also compromises Tat-induced HIV transcription.","method":"siRNA knockdown, RT-PCR quantification of HIV transcripts, viral replication assays","journal":"The Journal of biological chemistry","confidence":"Low","confidence_rationale":"Tier 3 / Weak — single knockdown approach with transcription readout, no direct mechanistic dissection of Med6-Tat interaction, single lab","pmids":["25100719"],"is_preprint":false},{"year":2017,"finding":"Crystal structure of 15-subunit core Mediator from S. pombe at 3.4 Å shows that Med6 terminal regions tether the middle module to the head module; sites of known Mediator mutations cluster at the head-middle module interface including Med6 terminal regions; the resulting atomic model of the PIC-cMed complex reveals the hook (containing CTD-crosslinking residues) contacts the TFIIH kinase.","method":"X-ray crystallography (3.4 Å), cryo-EM model integration, crosslinking mass spectrometry","journal":"Nature","confidence":"High","confidence_rationale":"Tier 1 / Strong — atomic-resolution structure with functional validation (mutational clustering), integrated with cryo-EM PIC model, replication across two studies","pmids":["28467824"],"is_preprint":false},{"year":2018,"finding":"Postrecruitment function of Med6 in transcriptional activation was demonstrated: a deletion mutant Med6p-Δ6 (amino acids 157-166) allows Mediator recruitment to the lexA operator with TBP but fails to support reporter gene expression, indicating Med6 has an essential role downstream of Mediator recruitment; a Δ2 deletion (amino acids 33-42) destabilizes Med6 and reduces Mediator association.","method":"Internal deletion mutagenesis, artificial recruitment assay (LexA fusion), in vitro transcription complementation assay, Western blot, chromatin immunoprecipitation","journal":"Biochemistry research international","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — multiple mutants with in vivo and in vitro transcription assays plus ChIP, but single lab and limited replication","pmids":["29992056"],"is_preprint":false},{"year":2023,"finding":"Cryo-EM/atomic model of yeast PIC-Mediator reveals that the Pol II CTD peptide 1 binds between the Med6 shoulder domain and the Med31 knob domain, defining specific CTD-Mediator contacts; CTD peptide 2 forms additional contacts with Med4.","method":"Cryo-EM structural analysis, atomic model building","journal":"Proceedings of the National Academy of Sciences of the United States of America","confidence":"High","confidence_rationale":"Tier 1 / Strong — atomic-resolution cryo-EM structure with defined CTD-Med6 shoulder contact, consistent with prior structural work, independent lab","pmids":["37014863"],"is_preprint":false},{"year":2023,"finding":"MED6 silencing by shRNA in cancer cells leads to increased lipid droplet accumulation and upregulation of lipid metabolism marker genes PLIN2 and DGAT1, indicating MED6 transcriptionally suppresses genes involved in lipid droplet biogenesis.","method":"RNAi/shRNA knockdown, fluorescence-activated cell sorting for lipid droplets, quantitative gene expression analysis","journal":"BioFactors (Oxford, England)","confidence":"Low","confidence_rationale":"Tier 3 / Weak — single knockdown approach with phenotypic readout, no direct mechanistic dissection of which transcriptional targets Med6 controls, single lab","pmids":["37983968"],"is_preprint":false}],"current_model":"MED6 is an essential, evolutionarily conserved subunit of the Mediator co-activator complex that occupies the head module (where its shoulder domain directly contacts the RNA Pol II CTD) and tethers the middle module to the head module; it is required for Mediator-dependent transcriptional activation at most class II promoters by enabling initiation complex formation after Mediator recruitment, functions downstream of activator-specific middle-module subunits as a convergence point for diverse activation signals, and transmits these signals to the basal machinery in part through a functionally essential interaction with the Srb4/Med14 subunit."},"narrative":{"mechanistic_narrative":"MED6 is an essential, evolutionarily conserved subunit of the Mediator co-activator complex that is required for transcriptional activation from most RNA polymerase II (class II) promoters in vivo, where it acts downstream of Mediator recruitment to enable formation of the transcription initiation complex [PMID:9234719, PMID:29992056]. Structurally, MED6 resides in the Mediator head module and serves as a physical bridge that tethers the middle module to the head module, with its terminal regions clamping the head–middle interface and its shoulder domain making direct contact with the Pol II C-terminal domain (CTD) [PMID:15710619, PMID:23123849, PMID:28467824, PMID:37014863]. Functionally, MED6 is a convergence point onto which diverse activator-specific signals are channeled before being transmitted to the basal machinery, since it is required for activation by every activator tested while neighboring subunits show activator-selective requirements [PMID:9891034]; signal transmission to the basal apparatus depends on a genetically and biochemically defined interaction with Srb4/MED17 [PMID:9710620, PMID:9671455] and connection to the middle module through MED21 [PMID:16758199]. Mediator's regulatory output is bidirectional and context-dependent: MED6-containing complexes can repress activated transcription, and CDK8-kinase-containing versus kinase-free MED6 subcomplexes differentially couple CTD phosphorylation to activation [PMID:9734358, PMID:10024883, PMID:17212659]. This activity is conserved and developmentally essential across metazoa, where MED6 is required for cell proliferation and for the transcriptional output of Ras and Wnt signaling [PMID:11438678, PMID:11688559, PMID:15790964].","teleology":[{"year":1997,"claim":"Established that MED6 is not a generic structural subunit but is specifically required for activated (not basal) transcription, localizing its function to initiation complex formation.","evidence":"Temperature-sensitive yeast mutant with in vivo transcription and reconstituted in vitro transcription assays","pmids":["9234719"],"confidence":"High","gaps":["Did not define which protein contacts mediate the initiation defect","Mechanism of activator selectivity not addressed"]},{"year":1998,"claim":"Showed MED6 function is conserved in mammals and linked it to CTD biology, demonstrating Mediator binds the Pol II CTD and stimulates its phosphorylation by TFIIH.","evidence":"Biochemical purification of mouse Mediator with CTD binding and TFIIH-dependent CTD phosphorylation assays","pmids":["9671713"],"confidence":"High","gaps":["Did not assign the CTD contact to MED6 specifically","Direct vs. indirect role of MED6 in phosphorylation unresolved"]},{"year":1998,"claim":"Identified the functional partner through which MED6 transmits activation signals, placing MED6 and Srb4 in the same Mediator subcomplex with a genetically required interaction.","evidence":"Allele-specific dominant suppressor screen plus biochemical Mediator subcomplex fractionation in yeast; complementary suppressor and Co-IP analysis with Srb4","pmids":["9710620","9671455"],"confidence":"High","gaps":["Atomic basis of the MED6–Srb4 interaction not defined","Whether the interaction is direct or bridged by other subunits not resolved"]},{"year":1999,"claim":"Demonstrated that MED6-containing human Mediator can both repress and synergistically enhance activator-dependent transcription, revealing bidirectional, context-dependent regulatory output.","evidence":"Biochemical purification of human NAT and SMCC complexes with in vitro repression/activation and CTD-independent activator interaction assays","pmids":["9734358","10024883"],"confidence":"High","gaps":["MED6-specific contribution to repression vs activation not isolated","Switch between repressive and activating states not mechanistically defined"]},{"year":1999,"claim":"Distinguished MED6 from other Mediator subunits as a universal activation requirement, establishing it as a convergence point for diverse activation signals.","evidence":"Differential display and Northern analysis across multiple activator systems (Bas1/Bas2, Gcn4, Gal4, Rap1) in Mediator mutant strains","pmids":["9891034"],"confidence":"Medium","gaps":["Convergence interpreted from genetic requirement, not direct activator-MED6 contacts","Does not explain how distinct signals are mechanistically integrated"]},{"year":2001,"claim":"Showed MED6 function is required for metazoan viability, proliferation, and the transcriptional readout of specific developmental signaling pathways.","evidence":"Drosophila loss-of-function mutants with microarray/RT-PCR; C. elegans RNAi epistasis with Ras and Wnt pathway alleles","pmids":["11438678","11688559"],"confidence":"Medium","gaps":["Selectivity for some but not all genes left unexplained at the mechanistic level","Direct vs indirect role in pathway-specific transcription not separated"]},{"year":2005,"claim":"Provided the first structural rationale for MED6 as the module bridge, showing it binds the MED7·MED21 surface to connect the middle and head modules.","evidence":"X-ray crystallography (3.0 Å) of MED7·MED21 with binding-surface analysis; in vivo Co-IP of the LET-425/MED6 Mediator complex in C. elegans","pmids":["15710619","15790964"],"confidence":"High","gaps":["Conformational role inferred from structure, not directly observed during activation","Functional consequence of bridging not yet perturbed"]},{"year":2006,"claim":"Defined the molecular route by which MED6 connects to the middle module, identifying MED21 as the direct middle-module link.","evidence":"Two-hybrid and Co-IP in insect cells and E. coli plus high-copy suppressor screen","pmids":["16758199"],"confidence":"Medium","gaps":["Functional consequences of the MED6–MED21 interaction not tested","Single biochemical study without structural localization of the contact"]},{"year":2007,"claim":"Showed that the kinase content of MED6 subcomplexes determines whether activation is coupled to Pol II phosphorylation, dissecting activation from CTD modification.","evidence":"Affinity purification of MED6/MED7/CDK8 subcomplexes (TMLC1 vs TMLC3) with in vitro transcription, kinase, and GTF interaction assays","pmids":["17212659"],"confidence":"Medium","gaps":["MED6's specific catalytic vs scaffolding role in each complex not separated","Single-lab biochemical fractionation"]},{"year":2010,"claim":"Resolved the recruitment hierarchy at a target promoter, showing MED14 (not MED1) positions MED6 at proximal promoters.","evidence":"ChIP with siRNA knockdown of MED1 and MED14 at the PPARγ target Fabp4","pmids":["20194623"],"confidence":"Medium","gaps":["Generality beyond Fabp4 not established","Mechanism by which MED14 positions MED6 not structurally defined"]},{"year":2012,"claim":"Placed MED6 precisely within the seven-subunit head module architecture that contacts Pol II, refining its structural role.","evidence":"X-ray crystallography (3.4 Å) of the S. pombe Mediator head module","pmids":["23123849"],"confidence":"High","gaps":["Static structure does not capture dynamics during initiation","Does not directly map activator signals onto MED6"]},{"year":2017,"claim":"Confirmed MED6 terminal regions as the functionally critical head–middle tether and integrated MED6 into a PIC-Mediator model linking it to TFIIH kinase contacts.","evidence":"X-ray crystallography (3.4 Å) of 15-subunit core Mediator with crosslinking MS and cryo-EM PIC model integration; mutation clustering at the head-middle interface","pmids":["28467824"],"confidence":"High","gaps":["Causal contribution of MED6 tether to activation kinetics not measured","How CTD contacts are remodeled during transcription unresolved"]},{"year":2018,"claim":"Genetically separated MED6's recruitment role from its post-recruitment function, defining an internal region essential after Mediator is already on the promoter.","evidence":"Internal deletion mutagenesis (Δ6, Δ2) with LexA artificial recruitment, in vitro transcription complementation, Western blot and ChIP","pmids":["29992056"],"confidence":"Medium","gaps":["Molecular event triggered by the essential region not identified","Single-lab study with limited replication"]},{"year":2023,"claim":"Pinpointed the atomic CTD-Mediator contact, showing the Pol II CTD binds between the MED6 shoulder and MED31 knob domains.","evidence":"Cryo-EM and atomic model of yeast PIC-Mediator","pmids":["37014863"],"confidence":"High","gaps":["Functional requirement of the shoulder-CTD contact not tested by mutation","Dynamics of CTD handoff to TFIIH kinase not resolved"]},{"year":2023,"claim":"Linked MED6 to repression of a specific transcriptional program, showing its loss derepresses lipid droplet biogenesis genes in cancer cells.","evidence":"shRNA knockdown with lipid droplet FACS and expression analysis of PLIN2/DGAT1","pmids":["37983968"],"confidence":"Low","gaps":["No mechanistic dissection of which targets MED6 directly controls","Single knockdown approach without rescue or reciprocal validation"]},{"year":null,"claim":"How activator-specific signals are physically integrated at MED6 and converted into the post-recruitment initiation event remains undefined.","evidence":"","pmids":[],"confidence":"Medium","gaps":["No structure of an activator-engaged MED6 state","The molecular event executed by MED6's essential post-recruitment region is unknown","How MED6 bridging dynamics couple to PIC assembly and CTD phosphorylation is unresolved"]}],"mechanism_profile":{"molecular_activity":[{"term_id":"GO:0140110","term_label":"transcription regulator activity","supporting_discovery_ids":[0,6,4]},{"term_id":"GO:0005198","term_label":"structural molecule activity","supporting_discovery_ids":[9,15,17]},{"term_id":"GO:0060090","term_label":"molecular adaptor activity","supporting_discovery_ids":[9,17,19]}],"localization":[{"term_id":"GO:0005634","term_label":"nucleus","supporting_discovery_ids":[0,14,18]},{"term_id":"GO:0000228","term_label":"nuclear chromosome","supporting_discovery_ids":[14,18]}],"pathway":[{"term_id":"R-HSA-74160","term_label":"Gene expression (Transcription)","supporting_discovery_ids":[0,6,17]},{"term_id":"R-HSA-162582","term_label":"Signal Transduction","supporting_discovery_ids":[8,10]},{"term_id":"R-HSA-1266738","term_label":"Developmental Biology","supporting_discovery_ids":[7,8,10]}],"complexes":["Mediator complex","Mediator head module","RNA polymerase II holoenzyme"],"partners":["MED17","MED21","MED7","MED14","MED31","MED13","MED12","MED23"],"other_free_text":[]}},"prefetch_data":{"uniprot":{"accession":"O75586","full_name":"Mediator of RNA polymerase II transcription subunit 6","aliases":["Activator-recruited cofactor 33 kDa component","ARC33","Mediator complex subunit 6","hMed6","Renal carcinoma antigen NY-REN-28"],"length_aa":246,"mass_kda":28.4,"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","subcellular_location":"Nucleus","url":"https://www.uniprot.org/uniprotkb/O75586/entry"},"depmap":{"release":"DepMap","has_data":true,"is_common_essential":true,"resolved_as":"","url":"https://depmap.org/portal/gene/MED6","classification":"Common Essential","n_dependent_lines":1192,"n_total_lines":1208,"dependency_fraction":0.9867549668874173},"opencell":{"profiled":true,"resolved_as":"","ensg_id":"ENSG00000133997","cell_line_id":"CID000236","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":"MED19","stoichiometry":10.0},{"gene":"MED20","stoichiometry":10.0},{"gene":"MED21","stoichiometry":10.0},{"gene":"MED27","stoichiometry":10.0},{"gene":"MED28","stoichiometry":10.0},{"gene":"MED29","stoichiometry":10.0},{"gene":"MED31","stoichiometry":10.0}],"url":"https://opencell.sf.czbiohub.org/target/CID000236","total_profiled":1310},"omim":[{"mim_id":"610197","title":"MEDIATOR COMPLEX SUBUNIT 25; MED25","url":"https://www.omim.org/entry/610197"},{"mim_id":"603800","title":"MEDIATOR COMPLEX SUBUNIT 21; MED21","url":"https://www.omim.org/entry/603800"},{"mim_id":"602984","title":"MEDIATOR COMPLEX SUBUNIT 6; MED6","url":"https://www.omim.org/entry/602984"},{"mim_id":"173870","title":"POLY(ADP-RIBOSE) POLYMERASE 1; PARP1","url":"https://www.omim.org/entry/173870"}],"hpa":{"profiled":true,"resolved_as":"","reliability":"Enhanced","locations":[{"location":"Nucleoplasm","reliability":"Enhanced"}],"tissue_specificity":"Low tissue specificity","tissue_distribution":"Detected in all","driving_tissues":[],"url":"https://www.proteinatlas.org/search/MED6"},"hgnc":{"alias_symbol":["NY-REN-28"],"prev_symbol":[]},"alphafold":{"accession":"O75586","domains":[{"cath_id":"3.10.450.580","chopping":"7-109","consensus_level":"high","plddt":92.7835,"start":7,"end":109},{"cath_id":"-","chopping":"110-150","consensus_level":"medium","plddt":91.6259,"start":110,"end":150}],"viewer_url":"https://alphafold.ebi.ac.uk/entry/O75586","model_url":"https://alphafold.ebi.ac.uk/files/AF-O75586-F1-model_v6.cif","pae_url":"https://alphafold.ebi.ac.uk/files/AF-O75586-F1-predicted_aligned_error_v6.png","plddt_mean":75.56},"mouse_models":{"mgi_url":"https://www.informatics.jax.org/marker/summary?nomen=MED6","jax_strain_url":"https://www.jax.org/strain/search?query=MED6"},"sequence":{"accession":"O75586","fasta_url":"https://rest.uniprot.org/uniprotkb/O75586.fasta","uniprot_url":"https://www.uniprot.org/uniprotkb/O75586/entry","alphafold_viewer_url":"https://alphafold.ebi.ac.uk/entry/O75586"}},"corpus_meta":[{"pmid":"9671713","id":"PMC_9671713","title":"Mammalian mediator of transcriptional regulation and its possible role as an end-point of signal transduction pathways.","date":"1998","source":"Proceedings of the National Academy of Sciences of the United States of America","url":"https://pubmed.ncbi.nlm.nih.gov/9671713","citation_count":258,"is_preprint":false},{"pmid":"10024883","id":"PMC_10024883","title":"A novel human SRB/MED-containing cofactor complex, SMCC, involved in transcription regulation.","date":"1999","source":"Molecular cell","url":"https://pubmed.ncbi.nlm.nih.gov/10024883","citation_count":233,"is_preprint":false},{"pmid":"9734358","id":"PMC_9734358","title":"NAT, a human complex containing Srb polypeptides that functions as a negative regulator of activated transcription.","date":"1998","source":"Molecular cell","url":"https://pubmed.ncbi.nlm.nih.gov/9734358","citation_count":194,"is_preprint":false},{"pmid":"28467824","id":"PMC_28467824","title":"Core Mediator structure at 3.4 Å extends model of transcription initiation 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for a functional interaction between mediator components Med6 and Srb4 in RNA polymerase II transcription.","date":"1998","source":"Molecular and cellular biology","url":"https://pubmed.ncbi.nlm.nih.gov/9710620","citation_count":71,"is_preprint":false},{"pmid":"15790964","id":"PMC_15790964","title":"Components of the transcriptional Mediator complex are required for asymmetric cell division in C. elegans.","date":"2005","source":"Development (Cambridge, England)","url":"https://pubmed.ncbi.nlm.nih.gov/15790964","citation_count":70,"is_preprint":false},{"pmid":"20194623","id":"PMC_20194623","title":"MED14 tethers mediator to the N-terminal domain of peroxisome proliferator-activated receptor gamma and is required for full transcriptional activity and adipogenesis.","date":"2010","source":"Molecular and cellular biology","url":"https://pubmed.ncbi.nlm.nih.gov/20194623","citation_count":67,"is_preprint":false},{"pmid":"9234719","id":"PMC_9234719","title":"A transcriptional mediator 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for activation in a reconstituted in vitro system due to a defect in initiation complex formation.\",\n      \"method\": \"Temperature-sensitive mutant analysis, in vivo transcription assays, reconstituted in vitro transcription system\",\n      \"journal\": \"Molecular and cellular biology\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 / Strong — in vitro reconstitution combined with in vivo genetic analysis, multiple promoters tested, functional defect clearly mapped to initiation complex formation\",\n      \"pmids\": [\"9234719\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 1998,\n      \"finding\": \"Mouse Mediator contains homologs of yeast Mediator subunits including Med6, binds to the RNA polymerase II C-terminal domain (CTD), and stimulates phosphorylation of the CTD by TFIIH.\",\n      \"method\": \"Biochemical purification, peptide sequencing, CTD binding assay, CTD phosphorylation assay\",\n      \"journal\": \"Proceedings of the National Academy of Sciences of the United States of America\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 / Moderate — in vitro biochemical assays (CTD binding and TFIIH-stimulated phosphorylation) with peptide sequencing confirming subunit identity, single lab\",\n      \"pmids\": [\"9671713\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 1998,\n      \"finding\": \"A functional interaction between Med6 and Srb4 is required for transcriptional activation; an allele-specific dominant suppressor screen identified SRB4 as a suppressor of a med6-ts mutation, and biochemical fractionation showed Med6 and Srb4 co-reside in the same Mediator subcomplex.\",\n      \"method\": \"Genetic suppressor screen, biochemical Mediator subcomplex fractionation, in vivo transcription assays\",\n      \"journal\": \"Molecular and cellular biology\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — reciprocal genetic epistasis (allele-specific suppression) combined with biochemical co-fractionation, multiple orthogonal methods in one study\",\n      \"pmids\": [\"9710620\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 1998,\n      \"finding\": \"Human NAT complex, containing the human homolog of Med6 along with Srb7, Srb10, Srb11, and Rgr1, represses activated transcription and phosphorylates the CTD of RNA polymerase II at residues distinct from those phosphorylated by TFIIH; the complex interacts with RNAPII in a CTD-independent manner, and CTD phosphorylation precludes this interaction.\",\n      \"method\": \"Biochemical purification, in vitro transcription repression assay, CTD phosphorylation assay, co-immunoprecipitation with RNAPII\",\n      \"journal\": \"Molecular cell\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 / Moderate — multiple in vitro functional assays (repression, kinase, binding) with defined biochemical dissection, single lab\",\n      \"pmids\": [\"9734358\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 1999,\n      \"finding\": \"Human SMCC complex, containing MED6 along with other SRB/MED homologs, can either repress activator-dependent transcription or, at limiting TFIIH, synergistically enhance it; the complex shows direct activator interactions and can act independently of the RNA polymerase II CTD.\",\n      \"method\": \"Biochemical purification, in vitro transcription assay, activator interaction assay\",\n      \"journal\": \"Molecular cell\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 / Moderate — in vitro reconstituted transcription assays demonstrating both positive and negative activities, CTD-independence tested directly, single lab\",\n      \"pmids\": [\"10024883\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 1998,\n      \"finding\": \"Med6 and Srb6, components of the RNA polymerase II holoenzyme, are dominant suppressors of a temperature-sensitive Srb4 mutation and physically interact with Srb4, placing Med6 within the holoenzyme where it participates in the balance between transcriptional activation and repression.\",\n      \"method\": \"Genetic suppressor screen, physical interaction assay (co-immunoprecipitation), in vivo transcription analysis\",\n      \"journal\": \"Molecular and cellular biology\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — genetic epistasis combined with physical interaction data, but details of interaction assay not fully described in abstract\",\n      \"pmids\": [\"9671455\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 1999,\n      \"finding\": \"Med6 is required for all activator-specific transcriptional activation processes tested (Bas1/Bas2-, Gcn4-, Gal4-, Rap1-mediated), whereas other Mediator subunits (Med9/Cse2, Med10/Nut2, Gal11, Med11) show activator-specific requirements; this positions Med6 as a convergence point where diverse activation signals meet to modulate Pol II activity.\",\n      \"method\": \"Differential display, Northern analysis of mRNA from Mediator mutant strains, genetic epistasis\",\n      \"journal\": \"Molecular and cellular biology\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — multiple activator systems tested with defined genetic mutants, single lab, primarily in vivo transcription readouts\",\n      \"pmids\": [\"9891034\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2001,\n      \"finding\": \"Drosophila Med6 (dMed6) is essential for viability and cell proliferation; dMed6 mutants fail to pupate and die in the third larval instar with severe proliferation defects in imaginal discs; cDNA microarray and quantitative RT-PCR show that transcriptional activation of many, but not all, genes is affected, including genes involved in cell proliferation and metabolism.\",\n      \"method\": \"Genetic loss-of-function mutant analysis, cDNA microarray, quantitative RT-PCR, in situ expression analysis\",\n      \"journal\": \"Molecular and cellular biology\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — genetic KO with specific phenotypic readout, genome-wide transcription profiling, multiple orthogonal methods, single lab\",\n      \"pmids\": [\"11438678\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2001,\n      \"finding\": \"C. elegans med-6 is required for development; RNAi of med-6 suppresses gain-of-function phenotypes of Ras pathway components (let-23 and let-60) and enhances loss-of-function of lin-3, placing Med-6 as required for transcriptional output of the Ras signaling pathway; med-6 is also involved in Wnt pathway-dependent male ray development.\",\n      \"method\": \"Genetic mutation analysis, RNAi epistasis with Ras and Wnt pathway alleles, phenotypic readout of signaling pathway outputs\",\n      \"journal\": \"Development (Cambridge, England)\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — genetic epistasis with defined pathway alleles across two signaling pathways, single lab\",\n      \"pmids\": [\"11688559\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2005,\n      \"finding\": \"MED7·MED21 (Med7·Srb7) crystal structure at 3.0 Å reveals that MED6 bridges the Mediator middle module to the head module; MED6 binds to putative protein-binding sites on the MED7·MED21 heterodimer surface, and a flexible MED6 bridge together with the MED7·MED21 hinge may account for conformational changes in Mediator upon binding to Pol II or activators.\",\n      \"method\": \"X-ray crystallography (3.0 Å), structural analysis of protein-binding surfaces\",\n      \"journal\": \"The Journal of biological chemistry\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 / Moderate — atomic-resolution crystal structure with functional interpretation of MED6 binding sites, single lab but high-resolution structural evidence\",\n      \"pmids\": [\"15710619\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2005,\n      \"finding\": \"In C. elegans, LET-425/MED6 forms a complex in vivo with LET-19/MED13, DPY-22/MED12, and SUR-2/MED23 (components of the Mediator complex), and this complex is required for Wnt-regulated asymmetric T-cell division and Wnt target gene repression.\",\n      \"method\": \"Co-immunoprecipitation in vivo, lineage analysis of let-19/dpy-22 mutants, epistasis with Wnt pathway components\",\n      \"journal\": \"Development (Cambridge, England)\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — reciprocal Co-IP demonstrating complex assembly combined with genetic epistasis and lineage analysis, single lab\",\n      \"pmids\": [\"15790964\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2006,\n      \"finding\": \"Med21 interacts with Med6 (confirmed by genetic and 2-hybrid data), and Med21 serves as a molecular switchboard integrating signals within the Mediator middle domain; interactions between Med21 and Med6 suggest Med6 is connected to the middle module through Med21.\",\n      \"method\": \"Two-hybrid analysis, co-immunoprecipitation of tagged proteins in insect cells and E. coli, high-copy suppressor screen\",\n      \"journal\": \"Molecular genetics and genomics\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — multiple interaction methods (2-hybrid + Co-IP in two heterologous systems), but functional consequences of Med6-Med21 interaction not directly tested in this study\",\n      \"pmids\": [\"16758199\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2007,\n      \"finding\": \"A 600-kDa MED6-containing subcomplex (TMLC3) augments transcriptional activation in vitro but lacks CDK8 and does not phosphorylate RNA Pol II; by contrast, the full 1.5-MDa complex (TMLC1) containing MED6, MED7, and CDK8 both activates transcription and phosphorylates Pol II, and preferentially interacts with TFIIE, TFIIF, and TFIIH.\",\n      \"method\": \"Affinity purification and HPLC-gel filtration of epitope-tagged MED6/MED7/CDK8 complexes, in vitro transcription assay, Pol II phosphorylation assay, co-immunoprecipitation with general transcription factors\",\n      \"journal\": \"Genes to cells\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — multiple biochemical assays (purification, transcription, kinase, interaction) from a defined subcomplex, single lab\",\n      \"pmids\": [\"17212659\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2007,\n      \"finding\": \"MED28 functions as a negative regulator of smooth muscle cell differentiation in concert with Med6, Med8, and Med18 within the Mediator head module; knockdown and overexpression experiments show Med6-containing head module components act together to suppress smooth muscle cell gene expression.\",\n      \"method\": \"siRNA knockdown, overexpression in NIH3T3 cells, gene expression analysis, mesenchymal precursor transdifferentiation assay\",\n      \"journal\": \"The Journal of biological chemistry\",\n      \"confidence\": \"Low\",\n      \"confidence_rationale\": \"Tier 3 / Weak — Med6 involvement inferred from co-knockdown phenotype without direct mechanistic dissection of Med6's specific contribution; single lab, no direct Med6-specific assay\",\n      \"pmids\": [\"17848560\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2010,\n      \"finding\": \"MED6 is recruited to the enhancer and proximal promoter of the PPARγ target gene Fabp4 in a MED1-independent manner, as shown by ChIP; MED14 knockdown reduces MED6 occupancy at the Fabp4 proximal promoter without affecting its binding at the enhancer, indicating MED14 is required for functional Mediator recruitment and positioning of MED6 at proximal promoters.\",\n      \"method\": \"ChIP (chromatin immunoprecipitation), siRNA knockdown of MED1 and MED14, reporter and endogenous gene expression assays\",\n      \"journal\": \"Molecular and cellular biology\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — ChIP-based localization with functional knockdown, multiple siRNA targets tested, single lab\",\n      \"pmids\": [\"20194623\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2012,\n      \"finding\": \"Crystal structure of the S. pombe Mediator head module at 3.4 Å reveals Med6 as one of seven head module subunits (with Med8, Med11, Med17, Med18, Med20, Med22); Med6 contributes to the fixed jaw submodule and is positioned within the head module architecture that contacts Pol II and its CTD.\",\n      \"method\": \"X-ray crystallography (3.4 Å), structural analysis\",\n      \"journal\": \"Nature\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 / Strong — high-resolution crystal structure, replicated across S. cerevisiae and S. pombe, structural position of Med6 in head module well-defined\",\n      \"pmids\": [\"23123849\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2014,\n      \"finding\": \"MED6 knockdown by siRNA significantly impairs HIV-1 replication at a post-integration step by inhibiting early HIV transcripts, specifically affecting transcription of the nascent viral mRNA transactivation-responsive element (TAR); MED6 knockdown also compromises Tat-induced HIV transcription.\",\n      \"method\": \"siRNA knockdown, RT-PCR quantification of HIV transcripts, viral replication assays\",\n      \"journal\": \"The Journal of biological chemistry\",\n      \"confidence\": \"Low\",\n      \"confidence_rationale\": \"Tier 3 / Weak — single knockdown approach with transcription readout, no direct mechanistic dissection of Med6-Tat interaction, single lab\",\n      \"pmids\": [\"25100719\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2017,\n      \"finding\": \"Crystal structure of 15-subunit core Mediator from S. pombe at 3.4 Å shows that Med6 terminal regions tether the middle module to the head module; sites of known Mediator mutations cluster at the head-middle module interface including Med6 terminal regions; the resulting atomic model of the PIC-cMed complex reveals the hook (containing CTD-crosslinking residues) contacts the TFIIH kinase.\",\n      \"method\": \"X-ray crystallography (3.4 Å), cryo-EM model integration, crosslinking mass spectrometry\",\n      \"journal\": \"Nature\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 / Strong — atomic-resolution structure with functional validation (mutational clustering), integrated with cryo-EM PIC model, replication across two studies\",\n      \"pmids\": [\"28467824\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2018,\n      \"finding\": \"Postrecruitment function of Med6 in transcriptional activation was demonstrated: a deletion mutant Med6p-Δ6 (amino acids 157-166) allows Mediator recruitment to the lexA operator with TBP but fails to support reporter gene expression, indicating Med6 has an essential role downstream of Mediator recruitment; a Δ2 deletion (amino acids 33-42) destabilizes Med6 and reduces Mediator association.\",\n      \"method\": \"Internal deletion mutagenesis, artificial recruitment assay (LexA fusion), in vitro transcription complementation assay, Western blot, chromatin immunoprecipitation\",\n      \"journal\": \"Biochemistry research international\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — multiple mutants with in vivo and in vitro transcription assays plus ChIP, but single lab and limited replication\",\n      \"pmids\": [\"29992056\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2023,\n      \"finding\": \"Cryo-EM/atomic model of yeast PIC-Mediator reveals that the Pol II CTD peptide 1 binds between the Med6 shoulder domain and the Med31 knob domain, defining specific CTD-Mediator contacts; CTD peptide 2 forms additional contacts with Med4.\",\n      \"method\": \"Cryo-EM structural analysis, atomic model building\",\n      \"journal\": \"Proceedings of the National Academy of Sciences of the United States of America\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 / Strong — atomic-resolution cryo-EM structure with defined CTD-Med6 shoulder contact, consistent with prior structural work, independent lab\",\n      \"pmids\": [\"37014863\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2023,\n      \"finding\": \"MED6 silencing by shRNA in cancer cells leads to increased lipid droplet accumulation and upregulation of lipid metabolism marker genes PLIN2 and DGAT1, indicating MED6 transcriptionally suppresses genes involved in lipid droplet biogenesis.\",\n      \"method\": \"RNAi/shRNA knockdown, fluorescence-activated cell sorting for lipid droplets, quantitative gene expression analysis\",\n      \"journal\": \"BioFactors (Oxford, England)\",\n      \"confidence\": \"Low\",\n      \"confidence_rationale\": \"Tier 3 / Weak — single knockdown approach with phenotypic readout, no direct mechanistic dissection of which transcriptional targets Med6 controls, single lab\",\n      \"pmids\": [\"37983968\"],\n      \"is_preprint\": false\n    }\n  ],\n  \"current_model\": \"MED6 is an essential, evolutionarily conserved subunit of the Mediator co-activator complex that occupies the head module (where its shoulder domain directly contacts the RNA Pol II CTD) and tethers the middle module to the head module; it is required for Mediator-dependent transcriptional activation at most class II promoters by enabling initiation complex formation after Mediator recruitment, functions downstream of activator-specific middle-module subunits as a convergence point for diverse activation signals, and transmits these signals to the basal machinery in part through a functionally essential interaction with the Srb4/Med14 subunit.\"\n}\n```","stage2_raw":"{\n  \"mechanistic_narrative\": \"MED6 is an essential, evolutionarily conserved subunit of the Mediator co-activator complex that is required for transcriptional activation from most RNA polymerase II (class II) promoters in vivo, where it acts downstream of Mediator recruitment to enable formation of the transcription initiation complex [#0, #18]. Structurally, MED6 resides in the Mediator head module and serves as a physical bridge that tethers the middle module to the head module, with its terminal regions clamping the head–middle interface and its shoulder domain making direct contact with the Pol II C-terminal domain (CTD) [#9, #15, #17, #19]. Functionally, MED6 is a convergence point onto which diverse activator-specific signals are channeled before being transmitted to the basal machinery, since it is required for activation by every activator tested while neighboring subunits show activator-selective requirements [#6]; signal transmission to the basal apparatus depends on a genetically and biochemically defined interaction with Srb4/MED17 [#2, #5] and connection to the middle module through MED21 [#11]. Mediator's regulatory output is bidirectional and context-dependent: MED6-containing complexes can repress activated transcription, and CDK8-kinase-containing versus kinase-free MED6 subcomplexes differentially couple CTD phosphorylation to activation [#3, #4, #12]. This activity is conserved and developmentally essential across metazoa, where MED6 is required for cell proliferation and for the transcriptional output of Ras and Wnt signaling [#7, #8, #10].\",\n  \"teleology\": [\n    {\n      \"year\": 1997,\n      \"claim\": \"Established that MED6 is not a generic structural subunit but is specifically required for activated (not basal) transcription, localizing its function to initiation complex formation.\",\n      \"evidence\": \"Temperature-sensitive yeast mutant with in vivo transcription and reconstituted in vitro transcription assays\",\n      \"pmids\": [\"9234719\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Did not define which protein contacts mediate the initiation defect\", \"Mechanism of activator selectivity not addressed\"]\n    },\n    {\n      \"year\": 1998,\n      \"claim\": \"Showed MED6 function is conserved in mammals and linked it to CTD biology, demonstrating Mediator binds the Pol II CTD and stimulates its phosphorylation by TFIIH.\",\n      \"evidence\": \"Biochemical purification of mouse Mediator with CTD binding and TFIIH-dependent CTD phosphorylation assays\",\n      \"pmids\": [\"9671713\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Did not assign the CTD contact to MED6 specifically\", \"Direct vs. indirect role of MED6 in phosphorylation unresolved\"]\n    },\n    {\n      \"year\": 1998,\n      \"claim\": \"Identified the functional partner through which MED6 transmits activation signals, placing MED6 and Srb4 in the same Mediator subcomplex with a genetically required interaction.\",\n      \"evidence\": \"Allele-specific dominant suppressor screen plus biochemical Mediator subcomplex fractionation in yeast; complementary suppressor and Co-IP analysis with Srb4\",\n      \"pmids\": [\"9710620\", \"9671455\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Atomic basis of the MED6–Srb4 interaction not defined\", \"Whether the interaction is direct or bridged by other subunits not resolved\"]\n    },\n    {\n      \"year\": 1999,\n      \"claim\": \"Demonstrated that MED6-containing human Mediator can both repress and synergistically enhance activator-dependent transcription, revealing bidirectional, context-dependent regulatory output.\",\n      \"evidence\": \"Biochemical purification of human NAT and SMCC complexes with in vitro repression/activation and CTD-independent activator interaction assays\",\n      \"pmids\": [\"9734358\", \"10024883\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"MED6-specific contribution to repression vs activation not isolated\", \"Switch between repressive and activating states not mechanistically defined\"]\n    },\n    {\n      \"year\": 1999,\n      \"claim\": \"Distinguished MED6 from other Mediator subunits as a universal activation requirement, establishing it as a convergence point for diverse activation signals.\",\n      \"evidence\": \"Differential display and Northern analysis across multiple activator systems (Bas1/Bas2, Gcn4, Gal4, Rap1) in Mediator mutant strains\",\n      \"pmids\": [\"9891034\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Convergence interpreted from genetic requirement, not direct activator-MED6 contacts\", \"Does not explain how distinct signals are mechanistically integrated\"]\n    },\n    {\n      \"year\": 2001,\n      \"claim\": \"Showed MED6 function is required for metazoan viability, proliferation, and the transcriptional readout of specific developmental signaling pathways.\",\n      \"evidence\": \"Drosophila loss-of-function mutants with microarray/RT-PCR; C. elegans RNAi epistasis with Ras and Wnt pathway alleles\",\n      \"pmids\": [\"11438678\", \"11688559\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Selectivity for some but not all genes left unexplained at the mechanistic level\", \"Direct vs indirect role in pathway-specific transcription not separated\"]\n    },\n    {\n      \"year\": 2005,\n      \"claim\": \"Provided the first structural rationale for MED6 as the module bridge, showing it binds the MED7·MED21 surface to connect the middle and head modules.\",\n      \"evidence\": \"X-ray crystallography (3.0 Å) of MED7·MED21 with binding-surface analysis; in vivo Co-IP of the LET-425/MED6 Mediator complex in C. elegans\",\n      \"pmids\": [\"15710619\", \"15790964\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Conformational role inferred from structure, not directly observed during activation\", \"Functional consequence of bridging not yet perturbed\"]\n    },\n    {\n      \"year\": 2006,\n      \"claim\": \"Defined the molecular route by which MED6 connects to the middle module, identifying MED21 as the direct middle-module link.\",\n      \"evidence\": \"Two-hybrid and Co-IP in insect cells and E. coli plus high-copy suppressor screen\",\n      \"pmids\": [\"16758199\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Functional consequences of the MED6–MED21 interaction not tested\", \"Single biochemical study without structural localization of the contact\"]\n    },\n    {\n      \"year\": 2007,\n      \"claim\": \"Showed that the kinase content of MED6 subcomplexes determines whether activation is coupled to Pol II phosphorylation, dissecting activation from CTD modification.\",\n      \"evidence\": \"Affinity purification of MED6/MED7/CDK8 subcomplexes (TMLC1 vs TMLC3) with in vitro transcription, kinase, and GTF interaction assays\",\n      \"pmids\": [\"17212659\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"MED6's specific catalytic vs scaffolding role in each complex not separated\", \"Single-lab biochemical fractionation\"]\n    },\n    {\n      \"year\": 2010,\n      \"claim\": \"Resolved the recruitment hierarchy at a target promoter, showing MED14 (not MED1) positions MED6 at proximal promoters.\",\n      \"evidence\": \"ChIP with siRNA knockdown of MED1 and MED14 at the PPARγ target Fabp4\",\n      \"pmids\": [\"20194623\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Generality beyond Fabp4 not established\", \"Mechanism by which MED14 positions MED6 not structurally defined\"]\n    },\n    {\n      \"year\": 2012,\n      \"claim\": \"Placed MED6 precisely within the seven-subunit head module architecture that contacts Pol II, refining its structural role.\",\n      \"evidence\": \"X-ray crystallography (3.4 Å) of the S. pombe Mediator head module\",\n      \"pmids\": [\"23123849\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Static structure does not capture dynamics during initiation\", \"Does not directly map activator signals onto MED6\"]\n    },\n    {\n      \"year\": 2017,\n      \"claim\": \"Confirmed MED6 terminal regions as the functionally critical head–middle tether and integrated MED6 into a PIC-Mediator model linking it to TFIIH kinase contacts.\",\n      \"evidence\": \"X-ray crystallography (3.4 Å) of 15-subunit core Mediator with crosslinking MS and cryo-EM PIC model integration; mutation clustering at the head-middle interface\",\n      \"pmids\": [\"28467824\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Causal contribution of MED6 tether to activation kinetics not measured\", \"How CTD contacts are remodeled during transcription unresolved\"]\n    },\n    {\n      \"year\": 2018,\n      \"claim\": \"Genetically separated MED6's recruitment role from its post-recruitment function, defining an internal region essential after Mediator is already on the promoter.\",\n      \"evidence\": \"Internal deletion mutagenesis (Δ6, Δ2) with LexA artificial recruitment, in vitro transcription complementation, Western blot and ChIP\",\n      \"pmids\": [\"29992056\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Molecular event triggered by the essential region not identified\", \"Single-lab study with limited replication\"]\n    },\n    {\n      \"year\": 2023,\n      \"claim\": \"Pinpointed the atomic CTD-Mediator contact, showing the Pol II CTD binds between the MED6 shoulder and MED31 knob domains.\",\n      \"evidence\": \"Cryo-EM and atomic model of yeast PIC-Mediator\",\n      \"pmids\": [\"37014863\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Functional requirement of the shoulder-CTD contact not tested by mutation\", \"Dynamics of CTD handoff to TFIIH kinase not resolved\"]\n    },\n    {\n      \"year\": 2023,\n      \"claim\": \"Linked MED6 to repression of a specific transcriptional program, showing its loss derepresses lipid droplet biogenesis genes in cancer cells.\",\n      \"evidence\": \"shRNA knockdown with lipid droplet FACS and expression analysis of PLIN2/DGAT1\",\n      \"pmids\": [\"37983968\"],\n      \"confidence\": \"Low\",\n      \"gaps\": [\"No mechanistic dissection of which targets MED6 directly controls\", \"Single knockdown approach without rescue or reciprocal validation\"]\n    },\n    {\n      \"year\": null,\n      \"claim\": \"How activator-specific signals are physically integrated at MED6 and converted into the post-recruitment initiation event remains undefined.\",\n      \"evidence\": \"\",\n      \"pmids\": [],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"No structure of an activator-engaged MED6 state\", \"The molecular event executed by MED6's essential post-recruitment region is unknown\", \"How MED6 bridging dynamics couple to PIC assembly and CTD phosphorylation is unresolved\"]\n    }\n  ],\n  \"mechanism_profile\": {\n    \"molecular_activity\": [\n      {\"term_id\": \"GO:0140110\", \"supporting_discovery_ids\": [0, 6, 4]},\n      {\"term_id\": \"GO:0005198\", \"supporting_discovery_ids\": [9, 15, 17]},\n      {\"term_id\": \"GO:0060090\", \"supporting_discovery_ids\": [9, 17, 19]}\n    ],\n    \"localization\": [\n      {\"term_id\": \"GO:0005634\", \"supporting_discovery_ids\": [0, 14, 18]},\n      {\"term_id\": \"GO:0000228\", \"supporting_discovery_ids\": [14, 18]}\n    ],\n    \"pathway\": [\n      {\"term_id\": \"R-HSA-74160\", \"supporting_discovery_ids\": [0, 6, 17]},\n      {\"term_id\": \"R-HSA-162582\", \"supporting_discovery_ids\": [8, 10]},\n      {\"term_id\": \"R-HSA-1266738\", \"supporting_discovery_ids\": [7, 8, 10]}\n    ],\n    \"complexes\": [\"Mediator complex\", \"Mediator head module\", \"RNA polymerase II holoenzyme\"],\n    \"partners\": [\"MED17\", \"MED21\", \"MED7\", \"MED14\", \"MED31\", \"MED13\", \"MED12\", \"MED23\"],\n    \"other_free_text\": []\n  }\n}","audit_flag":null,"evaluation":{"pairwise":"win","faith_supported":5,"faith_total":5,"faith_pct":100.0}}