{"gene":"MEST","run_date":"2026-06-10T02:59:50","timeline":{"discoveries":[{"year":1995,"finding":"PEG1/MEST was identified as a paternally expressed imprinted gene expressed only from the paternal allele; it belongs to the alpha/beta hydrolase fold family, placing it among enzymes with hydrolase activity.","method":"cDNA subtraction hybridization between normal and parthenogenetic embryos; sequence analysis","journal":"Nature genetics","confidence":"High","confidence_rationale":"Tier 2 / Strong — foundational identification by two independent labs, replicated across multiple subsequent studies; family assignment based on sequence analysis","pmids":["7550314"],"is_preprint":false},{"year":1997,"finding":"The human PEG1/MEST CpG island is methylated in a parent-of-origin-specific manner: the active paternal allele is unmethylated, while the silenced maternal allele is fully methylated at CpG sites.","method":"Restriction enzyme analysis (MspI/HpaII), bisulfite sequencing, RT-PCR with intragenic polymorphism for allele-specific expression analysis","journal":"Genomics","confidence":"High","confidence_rationale":"Tier 2 / Strong — replicated across multiple labs with orthogonal methods; differential methylation mechanism confirmed in human fetal tissues","pmids":["9192843","9302270"],"is_preprint":false},{"year":1997,"finding":"The expressed paternal Peg1 allele is unmethylated and the silenced maternal allele is fully methylated at the CpG island spanning exon 1; gametes carry the epigenetic information necessary to establish this allele-specific methylation pattern.","method":"Targeted mutation allele tracking in mouse embryos; methylation analysis of sperm and parthenogenetic embryos","journal":"Human molecular genetics","confidence":"High","confidence_rationale":"Tier 2 / Strong — direct allele-specific methylation mapped using knock-in allele; orthogonal confirmation with sperm and parthenogenote analysis","pmids":["9302270"],"is_preprint":false},{"year":1998,"finding":"Paternal transmission of a targeted Mest null allele causes embryonic growth retardation, reduced postnatal survival, and abnormal maternal behaviour (including impaired placentophagia) in mice; the mutation is reversibly silenced by passage through the female germ line, demonstrating the imprint is functional.","method":"Gene targeting in ES cells; paternal vs. maternal transmission of the null allele; behavioral phenotyping","journal":"Nature genetics","confidence":"High","confidence_rationale":"Tier 2 / Strong — clean genetic knockout with defined phenotypic readouts; imprinting mechanism confirmed by reversible silencing through female germline","pmids":["9771709"],"is_preprint":false},{"year":1999,"finding":"DNA methylation of the Mest promoter region suppresses its transcription; demethylation of a hypermethylated silent subclone reactivated Mest expression, and a methylated reporter construct failed to drive luciferase activity, whereas a second methylation-independent mechanism operates in adult tissues to silence Mest despite an unmethylated paternal allele.","method":"Reporter (luciferase) assay with methylated vs. unmethylated constructs; 5-azacytidine demethylation in cell lines; subclone analysis","journal":"Gene","confidence":"Medium","confidence_rationale":"Tier 1 / Moderate — direct reporter assay with methylated construct plus pharmacological demethylation; single lab, two orthogonal approaches","pmids":["9931489"],"is_preprint":false},{"year":2000,"finding":"MEST/Mest is expressed in endothelial cells of maternal decidua and hemangioblast/endothelial precursor cells in extraembryonic mesoderm; in decidual endothelium only the paternally derived allele is active. Expression in trophoblast-derived cells is absent in mice but present in human. The expression pattern in placental vasculature suggests a role in oncofetal angiogenesis.","method":"RT-PCR, restriction fragment length variant (RFLV) analysis for allele-specific expression; immunohistochemistry; in situ hybridization in mouse and human placenta","journal":"Developmental dynamics","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — allele-specific expression confirmed by RFLV analysis; localization by multiple histological methods in two species; single lab","pmids":["10679925"],"is_preprint":false},{"year":2000,"finding":"The paternal-specific methylation imprint at MEST/PEG1 is established during spermatogenesis: MEST remains unmethylated at all stages of male germ cell differentiation, including mature spermatozoa, whereas H19 methylation appears first in a subset of adult spermatogonia. Both genes are unmethylated in fetal spermatogonia, indicating erasure of pre-existing imprints at an early fetal stage.","method":"Bisulfite sequencing on microdissected individual cells at defined stages of human spermatogenesis","journal":"Human molecular genetics","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — single-cell bisulfite sequencing across developmental stages; single lab, single method","pmids":["10958657"],"is_preprint":false},{"year":2002,"finding":"An antisense transcript (PEG1-AS/MESTIT1) is expressed exclusively from the paternal allele and shares a common bidirectional promoter with the PEG1 sense isoform 2; CpG methylation of this shared promoter region abolishes its activity, linking methylation to transcriptional silencing of both sense and antisense transcripts.","method":"Northern blot; RT-PCR; bisulfite sequencing; luciferase reporter assay with CpG methylase treatment; allele-specific expression in somatic cell hybrids","journal":"The Journal of biological chemistry","confidence":"Medium","confidence_rationale":"Tier 1 / Moderate — direct reporter assay with methylase treatment plus allele-specific expression; single lab, multiple methods","pmids":["11821432"],"is_preprint":false},{"year":2004,"finding":"Ectopic expression of Mest/Peg1 in 3T3-L1 cells increased expression of adipogenic marker genes (PPARγ, C/EBPα, aP2), and transgenic overexpression in mouse adipose tissue caused marked enlargement of adipocytes, establishing Mest as a regulator of adipocyte size.","method":"Ectopic overexpression in 3T3-L1 cells; adipose-specific transgenic mouse; gene expression analysis","journal":"American journal of physiology. Endocrinology and metabolism","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — in vitro overexpression plus in vivo transgenic model; single lab, two complementary approaches","pmids":["15353408"],"is_preprint":false},{"year":2005,"finding":"The CpG methylation pattern of the Peg1/Mest CpG island is heterogeneous in freshly ovulated oocytes and changes dynamically during in vitro aging and preimplantation development; non-CpG methylation occurs in a stage-specific manner on fully CpG-methylated alleles and is reduced in two-cell stage embryos and blastocysts, indicating that imprint establishment at this locus is more dynamic than previously thought.","method":"Bisulfite sequencing of CpG and non-CpG sites in oocytes and preimplantation embryos","journal":"The Journal of biological chemistry","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — bisulfite sequencing at single-molecule resolution across developmental stages; single lab","pmids":["15778220"],"is_preprint":false},{"year":2008,"finding":"TIF1β (KAP1/TRIM28), through its interaction with HP1, maintains a heterochromatin-like structure at the MEST promoter characterized by H3K9 trimethylation, H4K20 trimethylation, hypoacetylation, DNA hypermethylation, and HP1 enrichment that represses MEST transcription. Disruption of the TIF1β–HP1 interaction releases TIF1β from the promoter, switches marks from H3K9me3/DNA hypermethylation to H3K27me3/DNA hypomethylation, and rapidly reactivates MEST expression from the non-imprinted allele, while the imprinted (maternal) allele's DNA methylation is insensitive to TIF1β loss of function.","method":"ChIP for histone marks and HP1; immunofluorescence; siRNA knockdown of TIF1β; allele-specific methylation analysis; RNAi of TIF1β–HP1 interaction domain","journal":"Molecular biology of the cell","confidence":"High","confidence_rationale":"Tier 2 / Strong — ChIP for multiple histone marks combined with allele-specific methylation and knockdown; multiple orthogonal methods distinguishing imprinted vs. non-imprinted allele regulation","pmids":["18923144"],"is_preprint":false},{"year":2011,"finding":"Mest/Peg1 inhibits Wnt/β-catenin signaling by blocking the glycosylation and plasma membrane localization of the Wnt co-receptor LRP6, thereby enhancing ubiquitination and degradation of β-catenin. Knockdown of Mest/Peg1 blocked adipogenic differentiation of 3T3-L1 cells, linking its Wnt-inhibitory activity to adipogenesis.","method":"Reporter assay (Wnt-responsive luciferase); co-immunoprecipitation; Western blot for β-catenin ubiquitination; LRP6 glycosylation/maturation assay; siRNA knockdown; adipogenesis assay","journal":"The Biochemical journal","confidence":"High","confidence_rationale":"Tier 2 / Strong — multiple orthogonal methods (reporter assay, Co-IP, glycosylation analysis, knockdown) establishing mechanism; single lab but mechanistically rigorous","pmids":["21375506"],"is_preprint":false},{"year":2011,"finding":"Alternative polyadenylation at the Mest locus generates long Mest transcripts (MestXL) that extend >10 kb into the antisense gene Copg2, exclusively in the developing CNS. MestXL formation causes preferential expression of Copg2 from the maternal allele in MestXL-expressing tissues via transcriptional interference; truncation of Mest mRNA eliminates MestXL and abolishes the allelic bias at Copg2.","method":"Northern blot; RT-PCR; allele-specific expression analysis; targeted Mest truncation allele in mice; tissue-specific poly-A site analysis","journal":"Nucleic acids research","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — genetic truncation allele directly tested for loss of MestXL and Copg2 allelic bias; single lab, multiple supporting methods","pmids":["22053079"],"is_preprint":false},{"year":2015,"finding":"In human adipocytes, MEST knockdown enhanced adipocyte differentiation by promoting PPARγ signaling, glycolysis and fatty acid biosynthesis pathways, and increased phosphorylation of pro-adipogenic transcription factors CREB and ATF1; conversely, MEST overexpression impaired adipogenesis. MEST silencing fully substituted for IBMX as an inducer of differentiation. This establishes MEST as an inhibitor of human adipogenesis, in contrast to its adipogenesis-promoting role described in mouse.","method":"siRNA knockdown; overexpression; adipogenic differentiation assay in human adipose-derived stem cells; transcriptome profiling; Western blot for CREB/ATF1 phosphorylation","journal":"International journal of obesity","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — reciprocal gain- and loss-of-function experiments with pathway-level readouts; single lab, multiple orthogonal assays","pmids":["26119994"],"is_preprint":false},{"year":2017,"finding":"Mest knockdown in mouse embryonic neocortex by in utero electroporation significantly reduced neuronal migration to the cortical plate and disrupted the bipolar-to-multipolar transition of neurons in the sub-ventricular zone; Mest-depleted neurons lost attachment to radial glia and adopted tangential migration. The differentiation and migration properties were mediated via Wnt-Akt signaling, and miR-335 (encoded in the Mest intron) was identified as blocking default tangential migration.","method":"In utero electroporation with shRNA; live imaging; radial glia attachment assay; pathway analysis (Wnt-Akt); co-expression of miR-335","journal":"Neuroscience","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — direct in vivo knockdown with specific neuronal phenotype and pathway placement; single lab","pmids":["28501506"],"is_preprint":false},{"year":2017,"finding":"Mest knockout mice show progressive loss of dopaminergic neurons in the substantia nigra (SNc) during adulthood, reflected by ~50% decrease in TH protein and dopamine release in the striatum, and reduced climbing behavior. Analysis of Lrp6 KO embryos showed a subtle opposing phenotype, suggesting Mest loss of function affects WNT signaling in the mdDA neuronal context.","method":"Mest KO mice; immunohistochemistry for TH; HPLC for striatal dopamine; behavioral testing (climbing assay); comparison with Lrp6 KO and Pitx3 mutant phenotypes","journal":"Frontiers in molecular neuroscience","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — clean KO with defined neuroanatomical and behavioral phenotype; epistasis comparison with Lrp6 KO suggests WNT-pathway link; single lab","pmids":["28133444"],"is_preprint":false},{"year":2017,"finding":"Mice with paternal allele-specific inactivation of Mest (MestpKO) fed high-fat diet show reduced adipose tissue expansion and adipocyte hypertrophy, improved glucose tolerance, and reduced WAT expression of hypoxia and inflammation genes without changes in caloric intake or energy expenditure. Ear-derived mesenchymal stem cells from Mest global KO showed reduced adipogenic capacity when Gpat4 was silenced, suggesting MEST facilitates lipid accumulation in adipocytes.","method":"Global and adipose-tissue-specific Mest KO mice; high-fat diet feeding; adipose tissue morphometry; glucose tolerance test; WAT transcriptome profiling; EMSC adipogenic assay with Gpat4 knockdown","journal":"PloS one","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — clean tissue-specific KO with multiple metabolic readouts; single lab, multiple orthogonal approaches","pmids":["28640866"],"is_preprint":false},{"year":2019,"finding":"MEST activates the IL-6/JAK/STAT3/Twist-1 signaling pathway to induce EMT and tumor metastasis in breast cancer; the C-terminal region of MEST is essential for STAT3 activation via induction of JAK2/STAT3 complex formation, and MEST promotes STAT3 nuclear translocation leading to Twist-1 induction.","method":"Overexpression and knockdown experiments; co-immunoprecipitation (JAK2/STAT3 complex); immunofluorescence (STAT3 nuclear translocation); reporter assays; in vivo metastasis model; C-terminal deletion constructs","journal":"Cell death and differentiation","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — Co-IP for complex formation, domain mapping, nuclear translocation imaging, and in vivo metastasis model; single lab, multiple methods","pmids":["30903102"],"is_preprint":false},{"year":2021,"finding":"MEST promotes lung cancer invasion and metastasis by interacting directly with VCP (valosin-containing protein); this interaction increases VCP–IκBα association, accelerating IκBα degradation and activating NF-κB signaling. VCP silencing abrogated MEST-driven NF-κB activation, placing MEST upstream of the VCP/IκBα/NF-κB pathway.","method":"Co-immunoprecipitation; mass spectrometry-based interactome; immunofluorescence colocalization; VCP siRNA epistasis; in vitro and in vivo metastasis assays","journal":"Journal of experimental & clinical cancer research","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — reciprocal Co-IP plus MS identification of VCP, epistasis by VCP silencing; single lab, multiple orthogonal methods","pmids":["34560900"],"is_preprint":false},{"year":2021,"finding":"Hypermethylation of the Mest promoter in Alzheimer's disease brain reduces Mest mRNA levels and activates Wnt signaling. CRISPR/Cas9 knockout of Mest in mouse embryonic stem cells and P19 cells causes neuronal differentiation arrest. shRNA-mediated Mest depletion in primary hippocampal neurons causes neurodegeneration, and depletion in primary cortical neurons induces tau phosphorylation at S199 and T231.","method":"Promoter methylation analysis in AD patient brains; CRISPR/Cas9 KO in mouse ES/P19 cells; lentiviral shMest in primary hippocampal and cortical neurons; phospho-tau Western blot; neurodegeneration assessment","journal":"Scientific reports","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — multiple loss-of-function models (CRISPR KO, shRNA), defined cellular phenotypes (differentiation arrest, tau phosphorylation); single lab","pmids":["34625606"],"is_preprint":false},{"year":2023,"finding":"MEST physically interacts with PURA and activates the SRCIN1/RASAL1-ERK-Snail signaling cascade to promote cancer invasion and metastasis in esophageal squamous cell carcinoma. Blockade of the MEST–PURA interaction with small molecule G699-0288 (identified by computational docking) significantly inhibits cancer metastasis.","method":"Genome-wide CRISPR/Cas9 screen; protein interactome (Co-IP/MS); RNA-seq; whole-genome methylation sequencing; surface plasmon resonance; modified ELISA; homology modeling; small-molecule functional assays in vitro and in vivo","journal":"EBioMedicine","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — CRISPR screen plus Co-IP/MS identification of PURA, SPR binding confirmation, and in vivo rescue; single lab, multiple orthogonal methods","pmids":["37149929"],"is_preprint":false},{"year":2016,"finding":"MEST protein is highly expressed in invasive extravillous trophoblast (EVT) cells. siRNA knockdown of MEST in HTR-8/SVneo cells significantly reduced cell invasion and migration as well as extravillous explant outgrowth, associated with downregulation of Twist, N-cadherin, and Vimentin. Decreased MEST expression correlated with isoform 2 promoter hypermethylation in placentas of missed abortions.","method":"Western blot; immunofluorescence; immunohistochemistry; siRNA knockdown; Matrigel invasion assay; Transwell migration; xCELLigence; bisulfite sequencing PCR for promoter methylation","journal":"Placenta","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — siRNA knockdown with multiple invasion/migration readouts; methylation-expression correlation in clinical samples; single lab","pmids":["27697227"],"is_preprint":false},{"year":2023,"finding":"Lewis Y (LeY) α1,3-fucosylation at Asn163 of MEST by FUT4 promotes trophoblast cell migration and invasion. Decreased LeY modification on MEST impairs its binding to translation initiation factor eIF4E2, thereby inhibiting translation initiation of implantation-related genes and causing trophoblast dysfunction associated with pregnancy failure.","method":"Lectin array; FUT4 siRNA knockdown; proteomics and translatomics; Co-IP of MEST with eIF4E2; site-directed glycosylation site identification; functional invasion/migration assays","journal":"Cell death & disease","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — identification of glycosylation site plus Co-IP of modified MEST with eIF4E2; functional assays; single lab, multiple methods","pmids":["37798282"],"is_preprint":false},{"year":2002,"finding":"In breast cancer, the switch from monoallelic to biallelic PEG1/MEST expression is caused by a promoter switch mechanism: isoform 1 remains imprinted (monoallelic) in both normal and tumor tissue, whereas upregulation of the normally minor isoform 2 (which is biallelically expressed) accounts for the apparent 'loss of imprinting' detected in invasive carcinomas.","method":"RT-PCR with isoform-specific primers; allele-specific expression analysis; real-time RT-PCR quantification of isoform ratio","journal":"Human molecular genetics","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — isoform-resolved allele-specific expression in matched normal/tumor pairs; single lab, mechanistically defines promoter-switch as the operative mechanism","pmids":["12023987"],"is_preprint":false},{"year":2002,"finding":"Peg1 (Mest) is expressed in myocardial trabeculae of the developing mouse heart, and mice lacking Peg1 show altered trabeculation pattern — increased compact myocardium thickness and reduced trabecular density — resembling human ventricular noncompaction cardiomyopathy.","method":"In situ hybridization; analysis of cardiac morphology in Peg1 KO mice by histology","journal":"Developmental dynamics","confidence":"Low","confidence_rationale":"Tier 3 / Weak — morphological phenotype in KO with no molecular mechanism identified; single lab, single histological method","pmids":["12242721"],"is_preprint":false},{"year":2015,"finding":"Mest (but not miR-335) is required for normal muscle regeneration: Mest+/- mice show retardation of body growth and decreased muscle growth during cardiotoxin-induced regeneration. Mest loss also affects expression of maternally expressed imprinted genes H19 and Igf2r in tibialis anterior muscle, suggesting Mest mediates muscle regeneration through regulation of imprinted gene networks.","method":"Mest+/- and miR-335-deficient mice; cardiotoxin-induced muscle regeneration; body weight measurement; gene expression analysis of imprinted gene network","journal":"PloS one","confidence":"Low","confidence_rationale":"Tier 3 / Weak — KO phenotype with pathway-level gene expression readouts but no direct molecular mechanism; single lab","pmids":["26098312"],"is_preprint":false},{"year":2018,"finding":"Peg1/Mest is expressed in mammary epithelial cells during gestation and knockdown in mammary epithelial cells suppresses alveoli formation and proliferation; overexpression in HC11 cells impairs lactogenic differentiation (β-casein induction), establishing a role for Mest in mammary gland maturation.","method":"Immunohistochemistry; immunofluorescence in 3D culture; siRNA knockdown; overexpression in HC11 cells; qRT-PCR for β-casein","journal":"Journal of cellular physiology","confidence":"Low","confidence_rationale":"Tier 3 / Weak — KD/OE with cellular phenotype (alveoli formation, differentiation) but no identified binding partner or molecular pathway; single lab","pmids":["30144363"],"is_preprint":false},{"year":2020,"finding":"MEST is expressed in the cytoplasm of periodontal ligament stem cells (PDLSCs); knockdown reduces stem cell markers (CD105, CD146, p75NTR, N-cadherin, NANOG), proliferative potential, and multilineage differentiation capacity, while overexpression in low-potency cells enhances stemness markers and differentiation capacity, establishing MEST as a regulator of PDLSC stemness.","method":"siRNA knockdown; lentiviral overexpression; flow cytometry for stem cell markers; multilineage differentiation assays (osteoblast, adipocyte, chondrocyte); immunofluorescence for subcellular localization","journal":"Stem cells international","confidence":"Low","confidence_rationale":"Tier 3 / Weak — reciprocal KD/OE with defined stem cell phenotype, but no molecular mechanism identified beyond marker expression; single lab","pmids":["32724317"],"is_preprint":false},{"year":2023,"finding":"NFIL3 transcription factor binds directly to the MEST DNA promoter to increase MEST transcription under low-shear-stress conditions; hydrogen sulfide inhibits this by sulfhydrylating NFIL3, reducing its binding to the MEST promoter and thereby suppressing MEST-driven endothelial-mesenchymal transition (EndMT) and atherosclerosis.","method":"ChIP-qPCR; luciferase reporter assay; NFIL3 knockdown; MEST overexpression/knockdown in vivo (AAV); sulfhydrylation assay; EndMT markers","journal":"Nitric oxide : biology and chemistry","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — direct NFIL3-MEST promoter binding confirmed by ChIP and reporter assay; in vivo AAV model; single lab, multiple methods","pmids":["38049061"],"is_preprint":false}],"current_model":"MEST/PEG1 is a paternally expressed imprinted gene belonging to the α/β-hydrolase fold family whose maternal allele is silenced by promoter CpG methylation maintained through a TIF1β–HP1–H3K9me3 heterochromatin mechanism; the active MEST protein inhibits Wnt/β-catenin signaling by blocking LRP6 glycosylation and membrane localization, promotes or inhibits adipogenesis in a species-dependent manner, supports neuronal migration via Wnt-Akt signaling and dopaminergic neuron survival, drives cancer invasion and metastasis through JAK2/STAT3/Twist-1 EMT induction and through interaction with VCP to activate NF-κB or with PURA to activate SRCIN1/RASAL1-ERK-Snail cascades, and undergoes α1,3-fucosylation at Asn163 that enables it to bind eIF4E2 and promote trophoblast invasion."},"narrative":{"mechanistic_narrative":"MEST/PEG1 is a paternally expressed imprinted gene of the α/β-hydrolase fold family that functions as a developmental and signaling regulator whose silenced maternal allele is controlled by parent-of-origin-specific CpG methylation [PMID:7550314, PMID:9192843, PMID:9302270, PMID:9771709]. The imprint is established during spermatogenesis and maintained at the promoter through a TIF1β(KAP1)–HP1 heterochromatin mechanism that enforces H3K9me3, H4K20me3, hypoacetylation, and DNA hypermethylation; disrupting the TIF1β–HP1 interaction reactivates the non-imprinted allele while leaving the imprinted allele's methylation intact [PMID:10958657, PMID:18923144]. Paternal loss of Mest in mice causes embryonic growth retardation, reduced survival, and abnormal maternal behavior, confirming the imprint is functional [PMID:9771709]. At the molecular level MEST acts as a node in Wnt/β-catenin signaling, inhibiting the pathway by blocking glycosylation and plasma-membrane localization of the co-receptor LRP6 and thereby promoting β-catenin ubiquitination and degradation [PMID:21375506]; through this and related signaling it supports neuronal migration and dopaminergic neuron survival via Wnt-Akt signaling [PMID:28501506, PMID:28133444] and regulates adipogenesis, promoting adipocyte size and lipid accumulation in mouse yet inhibiting differentiation in human cells [PMID:15353408, PMID:26119994, PMID:28640866]. In cancer MEST drives EMT, invasion, and metastasis through multiple effector axes: IL-6/JAK2/STAT3/Twist-1 activation requiring its C-terminus [PMID:30903102], direct interaction with VCP to accelerate IκBα degradation and activate NF-κB [PMID:34560900], and interaction with PURA to engage the SRCIN1/RASAL1-ERK-Snail cascade [PMID:37149929]. In trophoblast biology, Lewis-Y α1,3-fucosylation of MEST at Asn163 by FUT4 enables binding to the translation initiation factor eIF4E2 to promote invasion, and reduced MEST expression accompanies promoter hypermethylation in pregnancy failure [PMID:27697227, PMID:37798282].","teleology":[{"year":1995,"claim":"Establishing that PEG1/MEST is a paternally expressed imprinted gene of the α/β-hydrolase family defined the locus and predicted a hydrolase-type biochemical activity.","evidence":"cDNA subtraction between normal and parthenogenetic embryos with sequence analysis","pmids":["7550314"],"confidence":"High","gaps":["No enzymatic substrate identified for the predicted hydrolase activity","Family assignment is sequence-based, not biochemically demonstrated"]},{"year":1997,"claim":"Mapping parent-of-origin CpG methylation showed the silent maternal allele is methylated and the active paternal allele unmethylated, defining the epigenetic basis of imprinting at this locus.","evidence":"Bisulfite sequencing, restriction analysis, and allele-specific RT-PCR in human tissues and mouse knock-in embryos","pmids":["9192843","9302270"],"confidence":"High","gaps":["Did not identify the factors that read or maintain the methylation mark"]},{"year":1998,"claim":"Paternal transmission of a Mest null allele produced growth retardation and abnormal maternal behavior reversibly silenced through the female germline, proving the imprint is biologically functional.","evidence":"Gene targeting in mice with reciprocal allele transmission and behavioral phenotyping","pmids":["9771709"],"confidence":"High","gaps":["Molecular effector linking Mest loss to growth and behavior not defined at this stage"]},{"year":2000,"claim":"Single-cell methylation analysis across spermatogenesis showed the MEST imprint is established during male germ-cell development, locating the timing of imprint setting.","evidence":"Bisulfite sequencing of microdissected human germ cells at defined stages","pmids":["10958657"],"confidence":"Medium","gaps":["Enzymes establishing the male-germline imprint not identified"]},{"year":2002,"claim":"Discovery that biallelic MEST in breast cancer arises from a promoter switch upregulating a biallelic isoform 2 rather than true loss of imprinting refined how the locus is dysregulated in disease.","evidence":"Isoform-specific allele-resolved RT-PCR in matched normal/tumor pairs","pmids":["12023987"],"confidence":"Medium","gaps":["Trigger for isoform-2 promoter activation in tumors unknown"]},{"year":2008,"claim":"Identifying the TIF1β–HP1 heterochromatin complex as the maintainer of the repressive promoter state explained how the non-imprinted MEST allele is kept silent independently of the imprinted allele's methylation.","evidence":"ChIP for histone marks and HP1, siRNA of TIF1β, and allele-specific methylation analysis","pmids":["18923144"],"confidence":"High","gaps":["Does not explain how the imprinted maternal-allele methylation is maintained, which is TIF1β-insensitive"]},{"year":2011,"claim":"Demonstrating that MEST blocks LRP6 glycosylation and membrane localization to inhibit Wnt/β-catenin signaling provided the first defined molecular mechanism for the protein.","evidence":"Wnt reporter assay, Co-IP, LRP6 glycosylation assay, β-catenin ubiquitination Western, and adipogenesis assay","pmids":["21375506"],"confidence":"High","gaps":["Whether MEST acts catalytically (as the α/β-hydrolase prediction implies) or as a scaffold in blocking LRP6 is unresolved"]},{"year":2017,"claim":"In vivo knockdown and knockout placed MEST in neuronal migration and dopaminergic neuron survival via Wnt-Akt signaling, extending its Wnt regulatory role to the nervous system.","evidence":"In utero electroporation shRNA with live imaging, and Mest KO mice with TH immunohistochemistry, striatal dopamine HPLC, and behavioral testing","pmids":["28501506","28133444"],"confidence":"Medium","gaps":["Direct molecular link between MEST and Wnt-Akt in neurons not biochemically resolved","Mechanism of intronic miR-335 contribution incompletely separated from MEST protein effects"]},{"year":2017,"claim":"Reciprocal mouse and human studies revealed a species-divergent role in adipogenesis, with MEST facilitating lipid accumulation and adipocyte expansion in mouse but inhibiting differentiation in human cells.","evidence":"3T3-L1 and human ASC overexpression/knockdown, transgenic and tissue-specific KO mice with metabolic profiling","pmids":["15353408","26119994","28640866"],"confidence":"Medium","gaps":["Basis for the opposite mouse-vs-human direction not mechanistically explained"]},{"year":2021,"claim":"Identification of MEST–VCP and the upstream JAK2/STAT3/Twist-1 axis established distinct effector pathways through which MEST drives cancer EMT and metastasis.","evidence":"Co-IP/MS interactome, domain mapping, STAT3 nuclear translocation imaging, VCP epistasis, and in vivo metastasis models","pmids":["30903102","34560900"],"confidence":"Medium","gaps":["How a single protein engages multiple metastatic cascades (STAT3, NF-κB) is not unified","Direct binding interfaces not structurally resolved"]},{"year":2023,"claim":"Identification of MEST–PURA driving the SRCIN1/RASAL1-ERK-Snail cascade, and a small molecule blocking the interaction, validated MEST as a metastasis-promoting hub and a druggable target.","evidence":"Genome-wide CRISPR screen, Co-IP/MS, SPR binding, and small-molecule rescue in vitro and in vivo","pmids":["37149929"],"confidence":"Medium","gaps":["Structural basis of the MEST–PURA interaction inferred by homology modeling only"]},{"year":2023,"claim":"Discovery of FUT4-mediated Lewis-Y fucosylation at Asn163 enabling MEST binding to eIF4E2 linked a post-translational modification to translational control of trophoblast invasion.","evidence":"Lectin array, FUT4 knockdown, glycosite mapping, Co-IP of modified MEST with eIF4E2, and invasion assays","pmids":["27697227","37798282"],"confidence":"Medium","gaps":["Which implantation-related mRNAs are translationally controlled by the MEST–eIF4E2 complex not fully enumerated"]},{"year":null,"claim":"Whether MEST possesses genuine catalytic hydrolase activity, and how its scaffolding interactions (LRP6, VCP, PURA, eIF4E2) relate to its predicted α/β-hydrolase fold, remains unresolved.","evidence":"","pmids":[],"confidence":"Low","gaps":["No enzymatic substrate identified","No structural model of the catalytic pocket","Unclear whether one protein activity underlies its many context-specific roles"]}],"mechanism_profile":{"molecular_activity":[{"term_id":"GO:0016787","term_label":"hydrolase activity","supporting_discovery_ids":[0]},{"term_id":"GO:0060089","term_label":"molecular transducer activity","supporting_discovery_ids":[11,17,18,20]}],"localization":[{"term_id":"GO:0005829","term_label":"cytosol","supporting_discovery_ids":[27]},{"term_id":"GO:0005886","term_label":"plasma membrane","supporting_discovery_ids":[11]}],"pathway":[{"term_id":"R-HSA-162582","term_label":"Signal Transduction","supporting_discovery_ids":[11,17,18,20]},{"term_id":"R-HSA-1643685","term_label":"Disease","supporting_discovery_ids":[17,18,20,23]},{"term_id":"R-HSA-4839726","term_label":"Chromatin organization","supporting_discovery_ids":[10]}],"complexes":[],"partners":["LRP6","VCP","PURA","STAT3","JAK2","EIF4E2","TRIM28","NFIL3"],"other_free_text":[]}},"prefetch_data":{"uniprot":{"accession":"Q5EB52","full_name":"Mesoderm-specific transcript homolog protein","aliases":["Paternally-expressed gene 1 protein"],"length_aa":335,"mass_kda":38.8,"function":"Plays an important role in enabling neurons to transition from bipolar to multipolar shapes, a process essential for their correct migration toward the cortical plate during brain development. This function is likely achieved through the regulation of N-cadherin-dependent cell adhesion and the modulation of Akt and canonical Wnt signaling pathways. Essential for the development and maintenance of meso-diencephalic dopaminergic (mdDA) neurons during development. 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microbiology","url":"https://pubmed.ncbi.nlm.nih.gov/28492351","citation_count":13,"is_preprint":false},{"pmid":"10955473","id":"PMC_10955473","title":"Maternal chromosome 7 hetero/isodisomy in Silver-Russell syndrome and PEG1 biallelic expression.","date":"2000","source":"Clinical dysmorphology","url":"https://pubmed.ncbi.nlm.nih.gov/10955473","citation_count":12,"is_preprint":false},{"pmid":"34625606","id":"PMC_34625606","title":"Hypermethylation of Mest promoter causes aberrant Wnt signaling in patients with Alzheimer's disease.","date":"2021","source":"Scientific reports","url":"https://pubmed.ncbi.nlm.nih.gov/34625606","citation_count":12,"is_preprint":false},{"pmid":"38049061","id":"PMC_38049061","title":"Hydrogen sulfide attenuates atherosclerosis induced by low shear stress by sulfhydrylating endothelium NFIL3 to restrain MEST mediated endothelial mesenchymal transformation.","date":"2023","source":"Nitric oxide : biology and chemistry","url":"https://pubmed.ncbi.nlm.nih.gov/38049061","citation_count":12,"is_preprint":false},{"pmid":"38344730","id":"PMC_38344730","title":"Systematic Review of the Link Between Oxford MEST-C Classification and Complement Activation in IgA Nephropathy.","date":"2023","source":"Kidney international reports","url":"https://pubmed.ncbi.nlm.nih.gov/38344730","citation_count":11,"is_preprint":false},{"pmid":"35117228","id":"PMC_35117228","title":"MEST promotes bladder cancer cell proliferation, migration and invasion via STAT3/Twist-1-mediated EMT.","date":"2020","source":"Translational cancer research","url":"https://pubmed.ncbi.nlm.nih.gov/35117228","citation_count":11,"is_preprint":false},{"pmid":"12034494","id":"PMC_12034494","title":"The gene TSGA14, adjacent to the imprinted gene MEST, escapes genomic imprinting.","date":"2002","source":"Gene","url":"https://pubmed.ncbi.nlm.nih.gov/12034494","citation_count":11,"is_preprint":false},{"pmid":"9526615","id":"PMC_9526615","title":"PEG1 expression in maternal uniparental disomy 7.","date":"1997","source":"Annales de genetique","url":"https://pubmed.ncbi.nlm.nih.gov/9526615","citation_count":11,"is_preprint":false},{"pmid":"32724317","id":"PMC_32724317","title":"MEST Regulates the Stemness of Human Periodontal Ligament Stem Cells.","date":"2020","source":"Stem cells international","url":"https://pubmed.ncbi.nlm.nih.gov/32724317","citation_count":11,"is_preprint":false},{"pmid":"11032317","id":"PMC_11032317","title":"Molecular cloning and characterization of the Fugu rubripes MEST/COPG2 imprinting cluster and chromosomal localization in Fugu and Tetraodon nigroviridis.","date":"2000","source":"Chromosome research : an international journal on the molecular, supramolecular and evolutionary aspects of chromosome biology","url":"https://pubmed.ncbi.nlm.nih.gov/11032317","citation_count":11,"is_preprint":false},{"pmid":"37928396","id":"PMC_37928396","title":"Impact of DNA methylation of the human mesoderm-specific transcript (MEST) on male infertility.","date":"2023","source":"Heliyon","url":"https://pubmed.ncbi.nlm.nih.gov/37928396","citation_count":10,"is_preprint":false},{"pmid":"28854270","id":"PMC_28854270","title":"Hypermethylation of the non-imprinted maternal MEG3 and paternal MEST alleles is highly variable among normal individuals.","date":"2017","source":"PloS one","url":"https://pubmed.ncbi.nlm.nih.gov/28854270","citation_count":10,"is_preprint":false},{"pmid":"17687499","id":"PMC_17687499","title":"Imprinting analysis of the porcine MEST gene in 75 and 90 day placentas and prenatal tissues.","date":"2007","source":"Acta biochimica et biophysica Sinica","url":"https://pubmed.ncbi.nlm.nih.gov/17687499","citation_count":10,"is_preprint":false},{"pmid":"29157033","id":"PMC_29157033","title":"Alteration in methylation level at differential methylated regions of MEST and DLK1 in fetus of preeclampsia.","date":"2017","source":"Hypertension in pregnancy","url":"https://pubmed.ncbi.nlm.nih.gov/29157033","citation_count":10,"is_preprint":false},{"pmid":"11527825","id":"PMC_11527825","title":"Reactivity of MEST-1 (antigalactofuranose) with Trypanosoma cruzi glycosylinositol phosphorylceramides (GIPCs): immunolocalization of GIPCs in acidic vesicles of epimastigotes.","date":"2001","source":"Clinical and diagnostic laboratory immunology","url":"https://pubmed.ncbi.nlm.nih.gov/11527825","citation_count":10,"is_preprint":false},{"pmid":"22522229","id":"PMC_22522229","title":"Somatic reactivation of expression of the silent maternal Mest allele and acquisition of normal reproductive behaviour in a colony of Peg1/Mest mutant mice.","date":"2012","source":"The Journal of reproduction and development","url":"https://pubmed.ncbi.nlm.nih.gov/22522229","citation_count":9,"is_preprint":false},{"pmid":"25598283","id":"PMC_25598283","title":"Isoform-specific imprinting of the MEST gene in porcine parthenogenetic fetuses.","date":"2015","source":"Gene","url":"https://pubmed.ncbi.nlm.nih.gov/25598283","citation_count":9,"is_preprint":false},{"pmid":"36269039","id":"PMC_36269039","title":"MiR-29c-3p represses gastric cancer development via modulating MEST.","date":"2022","source":"Histology and histopathology","url":"https://pubmed.ncbi.nlm.nih.gov/36269039","citation_count":9,"is_preprint":false},{"pmid":"6257589","id":"PMC_6257589","title":"[Restriction and electron microscopic analyses of deletion derivatives thermosensitive with respect to maintaining plasmid pEG1].","date":"1980","source":"Genetika","url":"https://pubmed.ncbi.nlm.nih.gov/6257589","citation_count":9,"is_preprint":false},{"pmid":"35419530","id":"PMC_35419530","title":"Rationale and Trial Design of MesEnchymal Stem Cell Trial in Preventing Venous Stenosis of Hemodialysis Vascular Access Arteriovenous Fistula (MEST AVF Trial).","date":"2021","source":"Kidney360","url":"https://pubmed.ncbi.nlm.nih.gov/35419530","citation_count":9,"is_preprint":false},{"pmid":"32241207","id":"PMC_32241207","title":"The effect of folic acid deficiency on Mest/Peg1 in neural tube defects.","date":"2020","source":"The International journal of neuroscience","url":"https://pubmed.ncbi.nlm.nih.gov/32241207","citation_count":8,"is_preprint":false},{"pmid":"30144363","id":"PMC_30144363","title":"Peg1/Mest, an imprinted gene, is involved in mammary gland maturation.","date":"2018","source":"Journal of cellular physiology","url":"https://pubmed.ncbi.nlm.nih.gov/30144363","citation_count":7,"is_preprint":false},{"pmid":"33996346","id":"PMC_33996346","title":"Supercharged Mechanical Stromal-cell Transfer (MEST).","date":"2021","source":"Plastic and reconstructive surgery. Global open","url":"https://pubmed.ncbi.nlm.nih.gov/33996346","citation_count":7,"is_preprint":false},{"pmid":"40195296","id":"PMC_40195296","title":"A HOTAIR-associated super-enhancer orchestrates glioblastoma malignancy via MEST.","date":"2025","source":"Oncogenesis","url":"https://pubmed.ncbi.nlm.nih.gov/40195296","citation_count":7,"is_preprint":false},{"pmid":"35867696","id":"PMC_35867696","title":"Social and maternal behavior in mesoderm specific transcript (Mest)-deficient mice.","date":"2022","source":"PloS one","url":"https://pubmed.ncbi.nlm.nih.gov/35867696","citation_count":7,"is_preprint":false},{"pmid":"37798282","id":"PMC_37798282","title":"α1,3-fucosylation of MEST promotes invasion potential of cytotrophoblast cells by activating translation initiation.","date":"2023","source":"Cell death & disease","url":"https://pubmed.ncbi.nlm.nih.gov/37798282","citation_count":7,"is_preprint":false}],"cost":{"stage1":{"model":"claude-sonnet-4-6","input_tokens":49480,"output_tokens":7495,"usd":0.130433,"stage1_stop_reason":"end_turn"},"stage2":{"model":"claude-opus-4-8","input_tokens":17090,"output_tokens":3933,"usd":0.091887,"stage2_stop_reason":"end_turn"},"total_usd":0.22232,"stage1_batch_id":"msgbatch_01BbTCMyrRPyhtEq6b1MdwCN","stage2_batch_id":"msgbatch_011dEed5bbxYvTAaQWKJevxu","note":"batch pricing = 50% of standard"},"stage1_raw":"```json\n{\n  \"discoveries\": [\n    {\n      \"year\": 1995,\n      \"finding\": \"PEG1/MEST was identified as a paternally expressed imprinted gene expressed only from the paternal allele; it belongs to the alpha/beta hydrolase fold family, placing it among enzymes with hydrolase activity.\",\n      \"method\": \"cDNA subtraction hybridization between normal and parthenogenetic embryos; sequence analysis\",\n      \"journal\": \"Nature genetics\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — foundational identification by two independent labs, replicated across multiple subsequent studies; family assignment based on sequence analysis\",\n      \"pmids\": [\"7550314\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 1997,\n      \"finding\": \"The human PEG1/MEST CpG island is methylated in a parent-of-origin-specific manner: the active paternal allele is unmethylated, while the silenced maternal allele is fully methylated at CpG sites.\",\n      \"method\": \"Restriction enzyme analysis (MspI/HpaII), bisulfite sequencing, RT-PCR with intragenic polymorphism for allele-specific expression analysis\",\n      \"journal\": \"Genomics\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — replicated across multiple labs with orthogonal methods; differential methylation mechanism confirmed in human fetal tissues\",\n      \"pmids\": [\"9192843\", \"9302270\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 1997,\n      \"finding\": \"The expressed paternal Peg1 allele is unmethylated and the silenced maternal allele is fully methylated at the CpG island spanning exon 1; gametes carry the epigenetic information necessary to establish this allele-specific methylation pattern.\",\n      \"method\": \"Targeted mutation allele tracking in mouse embryos; methylation analysis of sperm and parthenogenetic embryos\",\n      \"journal\": \"Human molecular genetics\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — direct allele-specific methylation mapped using knock-in allele; orthogonal confirmation with sperm and parthenogenote analysis\",\n      \"pmids\": [\"9302270\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 1998,\n      \"finding\": \"Paternal transmission of a targeted Mest null allele causes embryonic growth retardation, reduced postnatal survival, and abnormal maternal behaviour (including impaired placentophagia) in mice; the mutation is reversibly silenced by passage through the female germ line, demonstrating the imprint is functional.\",\n      \"method\": \"Gene targeting in ES cells; paternal vs. maternal transmission of the null allele; behavioral phenotyping\",\n      \"journal\": \"Nature genetics\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — clean genetic knockout with defined phenotypic readouts; imprinting mechanism confirmed by reversible silencing through female germline\",\n      \"pmids\": [\"9771709\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 1999,\n      \"finding\": \"DNA methylation of the Mest promoter region suppresses its transcription; demethylation of a hypermethylated silent subclone reactivated Mest expression, and a methylated reporter construct failed to drive luciferase activity, whereas a second methylation-independent mechanism operates in adult tissues to silence Mest despite an unmethylated paternal allele.\",\n      \"method\": \"Reporter (luciferase) assay with methylated vs. unmethylated constructs; 5-azacytidine demethylation in cell lines; subclone analysis\",\n      \"journal\": \"Gene\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 1 / Moderate — direct reporter assay with methylated construct plus pharmacological demethylation; single lab, two orthogonal approaches\",\n      \"pmids\": [\"9931489\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2000,\n      \"finding\": \"MEST/Mest is expressed in endothelial cells of maternal decidua and hemangioblast/endothelial precursor cells in extraembryonic mesoderm; in decidual endothelium only the paternally derived allele is active. Expression in trophoblast-derived cells is absent in mice but present in human. The expression pattern in placental vasculature suggests a role in oncofetal angiogenesis.\",\n      \"method\": \"RT-PCR, restriction fragment length variant (RFLV) analysis for allele-specific expression; immunohistochemistry; in situ hybridization in mouse and human placenta\",\n      \"journal\": \"Developmental dynamics\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — allele-specific expression confirmed by RFLV analysis; localization by multiple histological methods in two species; single lab\",\n      \"pmids\": [\"10679925\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2000,\n      \"finding\": \"The paternal-specific methylation imprint at MEST/PEG1 is established during spermatogenesis: MEST remains unmethylated at all stages of male germ cell differentiation, including mature spermatozoa, whereas H19 methylation appears first in a subset of adult spermatogonia. Both genes are unmethylated in fetal spermatogonia, indicating erasure of pre-existing imprints at an early fetal stage.\",\n      \"method\": \"Bisulfite sequencing on microdissected individual cells at defined stages of human spermatogenesis\",\n      \"journal\": \"Human molecular genetics\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — single-cell bisulfite sequencing across developmental stages; single lab, single method\",\n      \"pmids\": [\"10958657\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2002,\n      \"finding\": \"An antisense transcript (PEG1-AS/MESTIT1) is expressed exclusively from the paternal allele and shares a common bidirectional promoter with the PEG1 sense isoform 2; CpG methylation of this shared promoter region abolishes its activity, linking methylation to transcriptional silencing of both sense and antisense transcripts.\",\n      \"method\": \"Northern blot; RT-PCR; bisulfite sequencing; luciferase reporter assay with CpG methylase treatment; allele-specific expression in somatic cell hybrids\",\n      \"journal\": \"The Journal of biological chemistry\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 1 / Moderate — direct reporter assay with methylase treatment plus allele-specific expression; single lab, multiple methods\",\n      \"pmids\": [\"11821432\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2004,\n      \"finding\": \"Ectopic expression of Mest/Peg1 in 3T3-L1 cells increased expression of adipogenic marker genes (PPARγ, C/EBPα, aP2), and transgenic overexpression in mouse adipose tissue caused marked enlargement of adipocytes, establishing Mest as a regulator of adipocyte size.\",\n      \"method\": \"Ectopic overexpression in 3T3-L1 cells; adipose-specific transgenic mouse; gene expression analysis\",\n      \"journal\": \"American journal of physiology. Endocrinology and metabolism\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — in vitro overexpression plus in vivo transgenic model; single lab, two complementary approaches\",\n      \"pmids\": [\"15353408\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2005,\n      \"finding\": \"The CpG methylation pattern of the Peg1/Mest CpG island is heterogeneous in freshly ovulated oocytes and changes dynamically during in vitro aging and preimplantation development; non-CpG methylation occurs in a stage-specific manner on fully CpG-methylated alleles and is reduced in two-cell stage embryos and blastocysts, indicating that imprint establishment at this locus is more dynamic than previously thought.\",\n      \"method\": \"Bisulfite sequencing of CpG and non-CpG sites in oocytes and preimplantation embryos\",\n      \"journal\": \"The Journal of biological chemistry\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — bisulfite sequencing at single-molecule resolution across developmental stages; single lab\",\n      \"pmids\": [\"15778220\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2008,\n      \"finding\": \"TIF1β (KAP1/TRIM28), through its interaction with HP1, maintains a heterochromatin-like structure at the MEST promoter characterized by H3K9 trimethylation, H4K20 trimethylation, hypoacetylation, DNA hypermethylation, and HP1 enrichment that represses MEST transcription. Disruption of the TIF1β–HP1 interaction releases TIF1β from the promoter, switches marks from H3K9me3/DNA hypermethylation to H3K27me3/DNA hypomethylation, and rapidly reactivates MEST expression from the non-imprinted allele, while the imprinted (maternal) allele's DNA methylation is insensitive to TIF1β loss of function.\",\n      \"method\": \"ChIP for histone marks and HP1; immunofluorescence; siRNA knockdown of TIF1β; allele-specific methylation analysis; RNAi of TIF1β–HP1 interaction domain\",\n      \"journal\": \"Molecular biology of the cell\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — ChIP for multiple histone marks combined with allele-specific methylation and knockdown; multiple orthogonal methods distinguishing imprinted vs. non-imprinted allele regulation\",\n      \"pmids\": [\"18923144\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2011,\n      \"finding\": \"Mest/Peg1 inhibits Wnt/β-catenin signaling by blocking the glycosylation and plasma membrane localization of the Wnt co-receptor LRP6, thereby enhancing ubiquitination and degradation of β-catenin. Knockdown of Mest/Peg1 blocked adipogenic differentiation of 3T3-L1 cells, linking its Wnt-inhibitory activity to adipogenesis.\",\n      \"method\": \"Reporter assay (Wnt-responsive luciferase); co-immunoprecipitation; Western blot for β-catenin ubiquitination; LRP6 glycosylation/maturation assay; siRNA knockdown; adipogenesis assay\",\n      \"journal\": \"The Biochemical journal\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — multiple orthogonal methods (reporter assay, Co-IP, glycosylation analysis, knockdown) establishing mechanism; single lab but mechanistically rigorous\",\n      \"pmids\": [\"21375506\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2011,\n      \"finding\": \"Alternative polyadenylation at the Mest locus generates long Mest transcripts (MestXL) that extend >10 kb into the antisense gene Copg2, exclusively in the developing CNS. MestXL formation causes preferential expression of Copg2 from the maternal allele in MestXL-expressing tissues via transcriptional interference; truncation of Mest mRNA eliminates MestXL and abolishes the allelic bias at Copg2.\",\n      \"method\": \"Northern blot; RT-PCR; allele-specific expression analysis; targeted Mest truncation allele in mice; tissue-specific poly-A site analysis\",\n      \"journal\": \"Nucleic acids research\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — genetic truncation allele directly tested for loss of MestXL and Copg2 allelic bias; single lab, multiple supporting methods\",\n      \"pmids\": [\"22053079\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2015,\n      \"finding\": \"In human adipocytes, MEST knockdown enhanced adipocyte differentiation by promoting PPARγ signaling, glycolysis and fatty acid biosynthesis pathways, and increased phosphorylation of pro-adipogenic transcription factors CREB and ATF1; conversely, MEST overexpression impaired adipogenesis. MEST silencing fully substituted for IBMX as an inducer of differentiation. This establishes MEST as an inhibitor of human adipogenesis, in contrast to its adipogenesis-promoting role described in mouse.\",\n      \"method\": \"siRNA knockdown; overexpression; adipogenic differentiation assay in human adipose-derived stem cells; transcriptome profiling; Western blot for CREB/ATF1 phosphorylation\",\n      \"journal\": \"International journal of obesity\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — reciprocal gain- and loss-of-function experiments with pathway-level readouts; single lab, multiple orthogonal assays\",\n      \"pmids\": [\"26119994\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2017,\n      \"finding\": \"Mest knockdown in mouse embryonic neocortex by in utero electroporation significantly reduced neuronal migration to the cortical plate and disrupted the bipolar-to-multipolar transition of neurons in the sub-ventricular zone; Mest-depleted neurons lost attachment to radial glia and adopted tangential migration. The differentiation and migration properties were mediated via Wnt-Akt signaling, and miR-335 (encoded in the Mest intron) was identified as blocking default tangential migration.\",\n      \"method\": \"In utero electroporation with shRNA; live imaging; radial glia attachment assay; pathway analysis (Wnt-Akt); co-expression of miR-335\",\n      \"journal\": \"Neuroscience\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — direct in vivo knockdown with specific neuronal phenotype and pathway placement; single lab\",\n      \"pmids\": [\"28501506\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2017,\n      \"finding\": \"Mest knockout mice show progressive loss of dopaminergic neurons in the substantia nigra (SNc) during adulthood, reflected by ~50% decrease in TH protein and dopamine release in the striatum, and reduced climbing behavior. Analysis of Lrp6 KO embryos showed a subtle opposing phenotype, suggesting Mest loss of function affects WNT signaling in the mdDA neuronal context.\",\n      \"method\": \"Mest KO mice; immunohistochemistry for TH; HPLC for striatal dopamine; behavioral testing (climbing assay); comparison with Lrp6 KO and Pitx3 mutant phenotypes\",\n      \"journal\": \"Frontiers in molecular neuroscience\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — clean KO with defined neuroanatomical and behavioral phenotype; epistasis comparison with Lrp6 KO suggests WNT-pathway link; single lab\",\n      \"pmids\": [\"28133444\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2017,\n      \"finding\": \"Mice with paternal allele-specific inactivation of Mest (MestpKO) fed high-fat diet show reduced adipose tissue expansion and adipocyte hypertrophy, improved glucose tolerance, and reduced WAT expression of hypoxia and inflammation genes without changes in caloric intake or energy expenditure. Ear-derived mesenchymal stem cells from Mest global KO showed reduced adipogenic capacity when Gpat4 was silenced, suggesting MEST facilitates lipid accumulation in adipocytes.\",\n      \"method\": \"Global and adipose-tissue-specific Mest KO mice; high-fat diet feeding; adipose tissue morphometry; glucose tolerance test; WAT transcriptome profiling; EMSC adipogenic assay with Gpat4 knockdown\",\n      \"journal\": \"PloS one\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — clean tissue-specific KO with multiple metabolic readouts; single lab, multiple orthogonal approaches\",\n      \"pmids\": [\"28640866\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2019,\n      \"finding\": \"MEST activates the IL-6/JAK/STAT3/Twist-1 signaling pathway to induce EMT and tumor metastasis in breast cancer; the C-terminal region of MEST is essential for STAT3 activation via induction of JAK2/STAT3 complex formation, and MEST promotes STAT3 nuclear translocation leading to Twist-1 induction.\",\n      \"method\": \"Overexpression and knockdown experiments; co-immunoprecipitation (JAK2/STAT3 complex); immunofluorescence (STAT3 nuclear translocation); reporter assays; in vivo metastasis model; C-terminal deletion constructs\",\n      \"journal\": \"Cell death and differentiation\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — Co-IP for complex formation, domain mapping, nuclear translocation imaging, and in vivo metastasis model; single lab, multiple methods\",\n      \"pmids\": [\"30903102\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2021,\n      \"finding\": \"MEST promotes lung cancer invasion and metastasis by interacting directly with VCP (valosin-containing protein); this interaction increases VCP–IκBα association, accelerating IκBα degradation and activating NF-κB signaling. VCP silencing abrogated MEST-driven NF-κB activation, placing MEST upstream of the VCP/IκBα/NF-κB pathway.\",\n      \"method\": \"Co-immunoprecipitation; mass spectrometry-based interactome; immunofluorescence colocalization; VCP siRNA epistasis; in vitro and in vivo metastasis assays\",\n      \"journal\": \"Journal of experimental & clinical cancer research\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — reciprocal Co-IP plus MS identification of VCP, epistasis by VCP silencing; single lab, multiple orthogonal methods\",\n      \"pmids\": [\"34560900\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2021,\n      \"finding\": \"Hypermethylation of the Mest promoter in Alzheimer's disease brain reduces Mest mRNA levels and activates Wnt signaling. CRISPR/Cas9 knockout of Mest in mouse embryonic stem cells and P19 cells causes neuronal differentiation arrest. shRNA-mediated Mest depletion in primary hippocampal neurons causes neurodegeneration, and depletion in primary cortical neurons induces tau phosphorylation at S199 and T231.\",\n      \"method\": \"Promoter methylation analysis in AD patient brains; CRISPR/Cas9 KO in mouse ES/P19 cells; lentiviral shMest in primary hippocampal and cortical neurons; phospho-tau Western blot; neurodegeneration assessment\",\n      \"journal\": \"Scientific reports\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — multiple loss-of-function models (CRISPR KO, shRNA), defined cellular phenotypes (differentiation arrest, tau phosphorylation); single lab\",\n      \"pmids\": [\"34625606\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2023,\n      \"finding\": \"MEST physically interacts with PURA and activates the SRCIN1/RASAL1-ERK-Snail signaling cascade to promote cancer invasion and metastasis in esophageal squamous cell carcinoma. Blockade of the MEST–PURA interaction with small molecule G699-0288 (identified by computational docking) significantly inhibits cancer metastasis.\",\n      \"method\": \"Genome-wide CRISPR/Cas9 screen; protein interactome (Co-IP/MS); RNA-seq; whole-genome methylation sequencing; surface plasmon resonance; modified ELISA; homology modeling; small-molecule functional assays in vitro and in vivo\",\n      \"journal\": \"EBioMedicine\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — CRISPR screen plus Co-IP/MS identification of PURA, SPR binding confirmation, and in vivo rescue; single lab, multiple orthogonal methods\",\n      \"pmids\": [\"37149929\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2016,\n      \"finding\": \"MEST protein is highly expressed in invasive extravillous trophoblast (EVT) cells. siRNA knockdown of MEST in HTR-8/SVneo cells significantly reduced cell invasion and migration as well as extravillous explant outgrowth, associated with downregulation of Twist, N-cadherin, and Vimentin. Decreased MEST expression correlated with isoform 2 promoter hypermethylation in placentas of missed abortions.\",\n      \"method\": \"Western blot; immunofluorescence; immunohistochemistry; siRNA knockdown; Matrigel invasion assay; Transwell migration; xCELLigence; bisulfite sequencing PCR for promoter methylation\",\n      \"journal\": \"Placenta\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — siRNA knockdown with multiple invasion/migration readouts; methylation-expression correlation in clinical samples; single lab\",\n      \"pmids\": [\"27697227\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2023,\n      \"finding\": \"Lewis Y (LeY) α1,3-fucosylation at Asn163 of MEST by FUT4 promotes trophoblast cell migration and invasion. Decreased LeY modification on MEST impairs its binding to translation initiation factor eIF4E2, thereby inhibiting translation initiation of implantation-related genes and causing trophoblast dysfunction associated with pregnancy failure.\",\n      \"method\": \"Lectin array; FUT4 siRNA knockdown; proteomics and translatomics; Co-IP of MEST with eIF4E2; site-directed glycosylation site identification; functional invasion/migration assays\",\n      \"journal\": \"Cell death & disease\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — identification of glycosylation site plus Co-IP of modified MEST with eIF4E2; functional assays; single lab, multiple methods\",\n      \"pmids\": [\"37798282\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2002,\n      \"finding\": \"In breast cancer, the switch from monoallelic to biallelic PEG1/MEST expression is caused by a promoter switch mechanism: isoform 1 remains imprinted (monoallelic) in both normal and tumor tissue, whereas upregulation of the normally minor isoform 2 (which is biallelically expressed) accounts for the apparent 'loss of imprinting' detected in invasive carcinomas.\",\n      \"method\": \"RT-PCR with isoform-specific primers; allele-specific expression analysis; real-time RT-PCR quantification of isoform ratio\",\n      \"journal\": \"Human molecular genetics\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — isoform-resolved allele-specific expression in matched normal/tumor pairs; single lab, mechanistically defines promoter-switch as the operative mechanism\",\n      \"pmids\": [\"12023987\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2002,\n      \"finding\": \"Peg1 (Mest) is expressed in myocardial trabeculae of the developing mouse heart, and mice lacking Peg1 show altered trabeculation pattern — increased compact myocardium thickness and reduced trabecular density — resembling human ventricular noncompaction cardiomyopathy.\",\n      \"method\": \"In situ hybridization; analysis of cardiac morphology in Peg1 KO mice by histology\",\n      \"journal\": \"Developmental dynamics\",\n      \"confidence\": \"Low\",\n      \"confidence_rationale\": \"Tier 3 / Weak — morphological phenotype in KO with no molecular mechanism identified; single lab, single histological method\",\n      \"pmids\": [\"12242721\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2015,\n      \"finding\": \"Mest (but not miR-335) is required for normal muscle regeneration: Mest+/- mice show retardation of body growth and decreased muscle growth during cardiotoxin-induced regeneration. Mest loss also affects expression of maternally expressed imprinted genes H19 and Igf2r in tibialis anterior muscle, suggesting Mest mediates muscle regeneration through regulation of imprinted gene networks.\",\n      \"method\": \"Mest+/- and miR-335-deficient mice; cardiotoxin-induced muscle regeneration; body weight measurement; gene expression analysis of imprinted gene network\",\n      \"journal\": \"PloS one\",\n      \"confidence\": \"Low\",\n      \"confidence_rationale\": \"Tier 3 / Weak — KO phenotype with pathway-level gene expression readouts but no direct molecular mechanism; single lab\",\n      \"pmids\": [\"26098312\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2018,\n      \"finding\": \"Peg1/Mest is expressed in mammary epithelial cells during gestation and knockdown in mammary epithelial cells suppresses alveoli formation and proliferation; overexpression in HC11 cells impairs lactogenic differentiation (β-casein induction), establishing a role for Mest in mammary gland maturation.\",\n      \"method\": \"Immunohistochemistry; immunofluorescence in 3D culture; siRNA knockdown; overexpression in HC11 cells; qRT-PCR for β-casein\",\n      \"journal\": \"Journal of cellular physiology\",\n      \"confidence\": \"Low\",\n      \"confidence_rationale\": \"Tier 3 / Weak — KD/OE with cellular phenotype (alveoli formation, differentiation) but no identified binding partner or molecular pathway; single lab\",\n      \"pmids\": [\"30144363\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2020,\n      \"finding\": \"MEST is expressed in the cytoplasm of periodontal ligament stem cells (PDLSCs); knockdown reduces stem cell markers (CD105, CD146, p75NTR, N-cadherin, NANOG), proliferative potential, and multilineage differentiation capacity, while overexpression in low-potency cells enhances stemness markers and differentiation capacity, establishing MEST as a regulator of PDLSC stemness.\",\n      \"method\": \"siRNA knockdown; lentiviral overexpression; flow cytometry for stem cell markers; multilineage differentiation assays (osteoblast, adipocyte, chondrocyte); immunofluorescence for subcellular localization\",\n      \"journal\": \"Stem cells international\",\n      \"confidence\": \"Low\",\n      \"confidence_rationale\": \"Tier 3 / Weak — reciprocal KD/OE with defined stem cell phenotype, but no molecular mechanism identified beyond marker expression; single lab\",\n      \"pmids\": [\"32724317\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2023,\n      \"finding\": \"NFIL3 transcription factor binds directly to the MEST DNA promoter to increase MEST transcription under low-shear-stress conditions; hydrogen sulfide inhibits this by sulfhydrylating NFIL3, reducing its binding to the MEST promoter and thereby suppressing MEST-driven endothelial-mesenchymal transition (EndMT) and atherosclerosis.\",\n      \"method\": \"ChIP-qPCR; luciferase reporter assay; NFIL3 knockdown; MEST overexpression/knockdown in vivo (AAV); sulfhydrylation assay; EndMT markers\",\n      \"journal\": \"Nitric oxide : biology and chemistry\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — direct NFIL3-MEST promoter binding confirmed by ChIP and reporter assay; in vivo AAV model; single lab, multiple methods\",\n      \"pmids\": [\"38049061\"],\n      \"is_preprint\": false\n    }\n  ],\n  \"current_model\": \"MEST/PEG1 is a paternally expressed imprinted gene belonging to the α/β-hydrolase fold family whose maternal allele is silenced by promoter CpG methylation maintained through a TIF1β–HP1–H3K9me3 heterochromatin mechanism; the active MEST protein inhibits Wnt/β-catenin signaling by blocking LRP6 glycosylation and membrane localization, promotes or inhibits adipogenesis in a species-dependent manner, supports neuronal migration via Wnt-Akt signaling and dopaminergic neuron survival, drives cancer invasion and metastasis through JAK2/STAT3/Twist-1 EMT induction and through interaction with VCP to activate NF-κB or with PURA to activate SRCIN1/RASAL1-ERK-Snail cascades, and undergoes α1,3-fucosylation at Asn163 that enables it to bind eIF4E2 and promote trophoblast invasion.\"\n}\n```","stage2_raw":"{\n  \"mechanistic_narrative\": \"MEST/PEG1 is a paternally expressed imprinted gene of the α/β-hydrolase fold family that functions as a developmental and signaling regulator whose silenced maternal allele is controlled by parent-of-origin-specific CpG methylation [#0, #1, #3]. The imprint is established during spermatogenesis and maintained at the promoter through a TIF1β(KAP1)–HP1 heterochromatin mechanism that enforces H3K9me3, H4K20me3, hypoacetylation, and DNA hypermethylation; disrupting the TIF1β–HP1 interaction reactivates the non-imprinted allele while leaving the imprinted allele's methylation intact [#6, #10]. Paternal loss of Mest in mice causes embryonic growth retardation, reduced survival, and abnormal maternal behavior, confirming the imprint is functional [#3]. At the molecular level MEST acts as a node in Wnt/β-catenin signaling, inhibiting the pathway by blocking glycosylation and plasma-membrane localization of the co-receptor LRP6 and thereby promoting β-catenin ubiquitination and degradation [#11]; through this and related signaling it supports neuronal migration and dopaminergic neuron survival via Wnt-Akt signaling [#14, #15] and regulates adipogenesis, promoting adipocyte size and lipid accumulation in mouse yet inhibiting differentiation in human cells [#8, #13, #16]. In cancer MEST drives EMT, invasion, and metastasis through multiple effector axes: IL-6/JAK2/STAT3/Twist-1 activation requiring its C-terminus [#17], direct interaction with VCP to accelerate IκBα degradation and activate NF-κB [#18], and interaction with PURA to engage the SRCIN1/RASAL1-ERK-Snail cascade [#20]. In trophoblast biology, Lewis-Y α1,3-fucosylation of MEST at Asn163 by FUT4 enables binding to the translation initiation factor eIF4E2 to promote invasion, and reduced MEST expression accompanies promoter hypermethylation in pregnancy failure [#21, #22].\",\n  \"teleology\": [\n    {\n      \"year\": 1995,\n      \"claim\": \"Establishing that PEG1/MEST is a paternally expressed imprinted gene of the α/β-hydrolase family defined the locus and predicted a hydrolase-type biochemical activity.\",\n      \"evidence\": \"cDNA subtraction between normal and parthenogenetic embryos with sequence analysis\",\n      \"pmids\": [\"7550314\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"No enzymatic substrate identified for the predicted hydrolase activity\", \"Family assignment is sequence-based, not biochemically demonstrated\"]\n    },\n    {\n      \"year\": 1997,\n      \"claim\": \"Mapping parent-of-origin CpG methylation showed the silent maternal allele is methylated and the active paternal allele unmethylated, defining the epigenetic basis of imprinting at this locus.\",\n      \"evidence\": \"Bisulfite sequencing, restriction analysis, and allele-specific RT-PCR in human tissues and mouse knock-in embryos\",\n      \"pmids\": [\"9192843\", \"9302270\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Did not identify the factors that read or maintain the methylation mark\"]\n    },\n    {\n      \"year\": 1998,\n      \"claim\": \"Paternal transmission of a Mest null allele produced growth retardation and abnormal maternal behavior reversibly silenced through the female germline, proving the imprint is biologically functional.\",\n      \"evidence\": \"Gene targeting in mice with reciprocal allele transmission and behavioral phenotyping\",\n      \"pmids\": [\"9771709\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Molecular effector linking Mest loss to growth and behavior not defined at this stage\"]\n    },\n    {\n      \"year\": 2000,\n      \"claim\": \"Single-cell methylation analysis across spermatogenesis showed the MEST imprint is established during male germ-cell development, locating the timing of imprint setting.\",\n      \"evidence\": \"Bisulfite sequencing of microdissected human germ cells at defined stages\",\n      \"pmids\": [\"10958657\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Enzymes establishing the male-germline imprint not identified\"]\n    },\n    {\n      \"year\": 2002,\n      \"claim\": \"Discovery that biallelic MEST in breast cancer arises from a promoter switch upregulating a biallelic isoform 2 rather than true loss of imprinting refined how the locus is dysregulated in disease.\",\n      \"evidence\": \"Isoform-specific allele-resolved RT-PCR in matched normal/tumor pairs\",\n      \"pmids\": [\"12023987\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Trigger for isoform-2 promoter activation in tumors unknown\"]\n    },\n    {\n      \"year\": 2008,\n      \"claim\": \"Identifying the TIF1β–HP1 heterochromatin complex as the maintainer of the repressive promoter state explained how the non-imprinted MEST allele is kept silent independently of the imprinted allele's methylation.\",\n      \"evidence\": \"ChIP for histone marks and HP1, siRNA of TIF1β, and allele-specific methylation analysis\",\n      \"pmids\": [\"18923144\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Does not explain how the imprinted maternal-allele methylation is maintained, which is TIF1β-insensitive\"]\n    },\n    {\n      \"year\": 2011,\n      \"claim\": \"Demonstrating that MEST blocks LRP6 glycosylation and membrane localization to inhibit Wnt/β-catenin signaling provided the first defined molecular mechanism for the protein.\",\n      \"evidence\": \"Wnt reporter assay, Co-IP, LRP6 glycosylation assay, β-catenin ubiquitination Western, and adipogenesis assay\",\n      \"pmids\": [\"21375506\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Whether MEST acts catalytically (as the α/β-hydrolase prediction implies) or as a scaffold in blocking LRP6 is unresolved\"]\n    },\n    {\n      \"year\": 2017,\n      \"claim\": \"In vivo knockdown and knockout placed MEST in neuronal migration and dopaminergic neuron survival via Wnt-Akt signaling, extending its Wnt regulatory role to the nervous system.\",\n      \"evidence\": \"In utero electroporation shRNA with live imaging, and Mest KO mice with TH immunohistochemistry, striatal dopamine HPLC, and behavioral testing\",\n      \"pmids\": [\"28501506\", \"28133444\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Direct molecular link between MEST and Wnt-Akt in neurons not biochemically resolved\", \"Mechanism of intronic miR-335 contribution incompletely separated from MEST protein effects\"]\n    },\n    {\n      \"year\": 2017,\n      \"claim\": \"Reciprocal mouse and human studies revealed a species-divergent role in adipogenesis, with MEST facilitating lipid accumulation and adipocyte expansion in mouse but inhibiting differentiation in human cells.\",\n      \"evidence\": \"3T3-L1 and human ASC overexpression/knockdown, transgenic and tissue-specific KO mice with metabolic profiling\",\n      \"pmids\": [\"15353408\", \"26119994\", \"28640866\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Basis for the opposite mouse-vs-human direction not mechanistically explained\"]\n    },\n    {\n      \"year\": 2021,\n      \"claim\": \"Identification of MEST–VCP and the upstream JAK2/STAT3/Twist-1 axis established distinct effector pathways through which MEST drives cancer EMT and metastasis.\",\n      \"evidence\": \"Co-IP/MS interactome, domain mapping, STAT3 nuclear translocation imaging, VCP epistasis, and in vivo metastasis models\",\n      \"pmids\": [\"30903102\", \"34560900\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"How a single protein engages multiple metastatic cascades (STAT3, NF-κB) is not unified\", \"Direct binding interfaces not structurally resolved\"]\n    },\n    {\n      \"year\": 2023,\n      \"claim\": \"Identification of MEST–PURA driving the SRCIN1/RASAL1-ERK-Snail cascade, and a small molecule blocking the interaction, validated MEST as a metastasis-promoting hub and a druggable target.\",\n      \"evidence\": \"Genome-wide CRISPR screen, Co-IP/MS, SPR binding, and small-molecule rescue in vitro and in vivo\",\n      \"pmids\": [\"37149929\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Structural basis of the MEST–PURA interaction inferred by homology modeling only\"]\n    },\n    {\n      \"year\": 2023,\n      \"claim\": \"Discovery of FUT4-mediated Lewis-Y fucosylation at Asn163 enabling MEST binding to eIF4E2 linked a post-translational modification to translational control of trophoblast invasion.\",\n      \"evidence\": \"Lectin array, FUT4 knockdown, glycosite mapping, Co-IP of modified MEST with eIF4E2, and invasion assays\",\n      \"pmids\": [\"27697227\", \"37798282\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Which implantation-related mRNAs are translationally controlled by the MEST–eIF4E2 complex not fully enumerated\"]\n    },\n    {\n      \"year\": null,\n      \"claim\": \"Whether MEST possesses genuine catalytic hydrolase activity, and how its scaffolding interactions (LRP6, VCP, PURA, eIF4E2) relate to its predicted α/β-hydrolase fold, remains unresolved.\",\n      \"evidence\": \"\",\n      \"pmids\": [],\n      \"confidence\": \"Low\",\n      \"gaps\": [\"No enzymatic substrate identified\", \"No structural model of the catalytic pocket\", \"Unclear whether one protein activity underlies its many context-specific roles\"]\n    }\n  ],\n  \"mechanism_profile\": {\n    \"molecular_activity\": [\n      {\"term_id\": \"GO:0016787\", \"supporting_discovery_ids\": [0]},\n      {\"term_id\": \"GO:0060089\", \"supporting_discovery_ids\": [11, 17, 18, 20]}\n    ],\n    \"localization\": [\n      {\"term_id\": \"GO:0005829\", \"supporting_discovery_ids\": [27]},\n      {\"term_id\": \"GO:0005886\", \"supporting_discovery_ids\": [11]}\n    ],\n    \"pathway\": [\n      {\"term_id\": \"R-HSA-162582\", \"supporting_discovery_ids\": [11, 17, 18, 20]},\n      {\"term_id\": \"R-HSA-1643685\", \"supporting_discovery_ids\": [17, 18, 20, 23]},\n      {\"term_id\": \"R-HSA-4839726\", \"supporting_discovery_ids\": [10]}\n    ],\n    \"complexes\": [],\n    \"partners\": [\"LRP6\", \"VCP\", \"PURA\", \"STAT3\", \"JAK2\", \"eIF4E2\", \"TRIM28\", \"NFIL3\"],\n    \"other_free_text\": []\n  }\n}","audit_flag":null,"evaluation":{"pairwise":"win","faith_supported":6,"faith_total":6,"faith_pct":100.0}}