Journal: Cell reports

Article Title: Amplification of MED30 at chromosome 8q24 reprograms MYC binding to low-affinity oncogenic enhancers in cancer cells

doi: 10.1016/j.celrep.2026.117498

Figure Lengend Snippet: (A) CoIP experiments in Tet-on MED30-3xHA Mia PaCa-2 cells. (B) Genome browser example of indicated mediator subunits and other factors in gained enhancer upon MED30 OE (+Dox), by CUT&Tag experiments in Mia PaCa-2 cells. (C) MED1, MED4, MED12, MED17, MED23, and MED30 tag density profile on MED30 OE gained peaks Mia PaCa-2 cells (CUT&Tag experiment). (D) Transcription level (PRO-seq) changes of the exemplified genes with gained mediator binding peaks nearby. Data are presented as mean ± SD. ** p < 0.01; two-tailed Student’s t test. (E) Tag density profile of indicated histone marks and transcription level (PRO-seq) on MED30 gained sites.

Article Snippet: MED12 , Bethyl , A300-774 A.

Techniques: Binding Assay, Two Tailed Test

Journal: PLOS One

Article Title: Impact of MED12 mutation and CDK8 activity on uterine leiomyoma growth and response to gonadotropin-releasing hormone agonist treatment

doi: 10.1371/journal.pone.0338485

Figure Lengend Snippet: (A) Representative western blot image; CTD-pS5 phosphorylated by the CDK8-MED12 complex and non-phosphorylated CTD protein (p–) are indicated in the upper part of the panel. Protein extracts were immunoprecipitated with anti-MED12 antibodies and immunoblotted with anti-CDK8 and anti-MED12 antibodies as indicated in the middle of the panel. Protein extracts were immunoblotted with anti-CDK8, anti-MED12, and anti-β-actin antibodies as indicated in the lower part of the image. (B–F) Graph of each protein band intensity according to MED12 status (WT or MUT) in GnRH agonist-treated (GnRH agonist+) and -untreated (GnRH agonist–) groups. * p < 0.05; ** p < 0.01. CTD, C-terminal domain (of the largest subunit of RNA polymerase II); CTD-pS5, C-terminal domain with phosphorylated 5th serine residue; GnRH, gonadotropin-releasing hormone; IP, immunoprecipitated; MUT, mutant; WT, wild type.

Article Snippet: Rabbit polyclonal anti-MED12 antibody (1 μg) (Bethyl Laboratories, Inc., Montgomery, TX) was cross-linked to 12.5 μL Dynabeads protein G (Thermo Fisher Scientific) and mixed with 380 μL (tissues) or 1 mL (cells) of protein extract at 4°C overnight.

Techniques: Western Blot, Immunoprecipitation, Residue, Mutagenesis

Journal: PLOS One

Article Title: Impact of MED12 mutation and CDK8 activity on uterine leiomyoma growth and response to gonadotropin-releasing hormone agonist treatment

doi: 10.1371/journal.pone.0338485

Figure Lengend Snippet: Primary leiomyoma cells were treated with Snx ± EP for 4 days followed by CDK8 phosphorylation assay and western blotting analysis. Results for the leiomyoma cells derived from two patients (LM1 and LM2) are shown. Data are normalized to vehicles and presented as the means of three independent experiments. (A) Representative western blot image: the CTD-pS5 protein is indicated in the upper part of the panel. Cell extracts were immunoprecipitated with an anti-MED12 antibody and immunoblotted with anti-CDK8 and anti-MED12 antibodies, as indicated in the middle of the panel. Protein extracts were immunoblotted with anti-CDK8, anti-MED12, and anti-ERα antibodies, as indicated in the lower part of the panel. Anti-β-actin was used as a loading control. (B) Quantification of phosphorylated CTD protein and immunoprecipitated protein. (C) Quantification of protein extracts. Protein levels were quantified, and expression data are presented relative to β-actin. The data are normalized to the control and presented as the mean of three independent experiments. Two-way ANOVA showed an interaction between Snx and EP at CTD-pS5 in LM2, and post-hoc test showed a significant difference between EP (–) Snx (–) and other groups. (*, p < 0.05; **, p < 0.01). CTD, C-terminal domain (of the largest subunit of RNA polymerase II); CTD-pS5, C-terminal domain with phosphorylated 5th serine residue; EP, 17-β estradiol (E2) + progesterone (P4); ERα, estrogen receptor alpha; IP, immunoprecipitated; Snx, Senexin B.

Article Snippet: Rabbit polyclonal anti-MED12 antibody (1 μg) (Bethyl Laboratories, Inc., Montgomery, TX) was cross-linked to 12.5 μL Dynabeads protein G (Thermo Fisher Scientific) and mixed with 380 μL (tissues) or 1 mL (cells) of protein extract at 4°C overnight.

Techniques: Phospho-proteomics, Western Blot, Derivative Assay, Immunoprecipitation, Control, Expressing, Residue

Journal: PLOS One

Article Title: Impact of MED12 mutation and CDK8 activity on uterine leiomyoma growth and response to gonadotropin-releasing hormone agonist treatment

doi: 10.1371/journal.pone.0338485

Figure Lengend Snippet: (A) Representative western blot image; CTD-pS5 phosphorylated by the CDK8-MED12 complex and non-phosphorylated CTD protein (p–) are indicated in the upper part of the panel. Protein extracts were immunoprecipitated with anti-MED12 antibodies and immunoblotted with anti-CDK8 and anti-MED12 antibodies as indicated in the middle of the panel. Protein extracts were immunoblotted with anti-CDK8, anti-MED12, and anti-β-actin antibodies as indicated in the lower part of the image. (B–F) Graph of each protein band intensity according to MED12 status (WT or MUT) in GnRH agonist-treated (GnRH agonist+) and -untreated (GnRH agonist–) groups. * p < 0.05; ** p < 0.01. CTD, C-terminal domain (of the largest subunit of RNA polymerase II); CTD-pS5, C-terminal domain with phosphorylated 5th serine residue; GnRH, gonadotropin-releasing hormone; IP, immunoprecipitated; MUT, mutant; WT, wild type.

Article Snippet: Four primary cultures of MED12 WT leiomyomas and two primary cultures of normal myometrium were isolated as previously described [ ] and stored at –145°C in Bambanker (Nippon Genetics Co., Ltd., Tokyo, Japan) until use.

Techniques: Western Blot, Immunoprecipitation, Residue, Mutagenesis

Journal: PLOS One

Article Title: Impact of MED12 mutation and CDK8 activity on uterine leiomyoma growth and response to gonadotropin-releasing hormone agonist treatment

doi: 10.1371/journal.pone.0338485

Figure Lengend Snippet: Primary leiomyoma cells were treated with Snx ± EP for 4 days followed by CDK8 phosphorylation assay and western blotting analysis. Results for the leiomyoma cells derived from two patients (LM1 and LM2) are shown. Data are normalized to vehicles and presented as the means of three independent experiments. (A) Representative western blot image: the CTD-pS5 protein is indicated in the upper part of the panel. Cell extracts were immunoprecipitated with an anti-MED12 antibody and immunoblotted with anti-CDK8 and anti-MED12 antibodies, as indicated in the middle of the panel. Protein extracts were immunoblotted with anti-CDK8, anti-MED12, and anti-ERα antibodies, as indicated in the lower part of the panel. Anti-β-actin was used as a loading control. (B) Quantification of phosphorylated CTD protein and immunoprecipitated protein. (C) Quantification of protein extracts. Protein levels were quantified, and expression data are presented relative to β-actin. The data are normalized to the control and presented as the mean of three independent experiments. Two-way ANOVA showed an interaction between Snx and EP at CTD-pS5 in LM2, and post-hoc test showed a significant difference between EP (–) Snx (–) and other groups. (*, p < 0.05; **, p < 0.01). CTD, C-terminal domain (of the largest subunit of RNA polymerase II); CTD-pS5, C-terminal domain with phosphorylated 5th serine residue; EP, 17-β estradiol (E2) + progesterone (P4); ERα, estrogen receptor alpha; IP, immunoprecipitated; Snx, Senexin B.

Article Snippet: Four primary cultures of MED12 WT leiomyomas and two primary cultures of normal myometrium were isolated as previously described [ ] and stored at –145°C in Bambanker (Nippon Genetics Co., Ltd., Tokyo, Japan) until use.

Techniques: Phospho-proteomics, Western Blot, Derivative Assay, Immunoprecipitation, Control, Expressing, Residue

Journal: Molecular Medicine

Article Title: Functional characterization of the MED12 p.Arg1138Trp variant in females: implications for neural development and disease mechanism

doi: 10.1186/s10020-025-01365-5

Figure Lengend Snippet: MED12 patient-derived iPSCs and protein expression. A Schematic indicating the MED12 variant location and known protein domains. B MED12 3D modelling utilizing AlphaMissense indicating position and change of amino acid. C and D Targeted amplicon sequencing of gDNA and cDNA showing read counts and the percentage of reads aligning to MED12 WT or MED12 VUS. E Immunofluorescent staining of the variant and WT iPSCs indicating cellular localization of MED12 protein. White bar, 60 μm

Article Snippet: Membranes were blocked overnight at 4 °C with Intercept ® (TBS) Blocking Buffer (LI-COR Biosciences), then incubated with rabbit anti-human MED12 monoclonal antibody (1:1000; clone BLR084G; Bethyl Laboratories, USA) and/or β-actin antibody (1:2000; MA5-15729; Life Technologies, Australia).

Techniques: Derivative Assay, Expressing, Variant Assay, Amplification, Sequencing, Staining

Journal: Molecular Medicine

Article Title: Functional characterization of the MED12 p.Arg1138Trp variant in females: implications for neural development and disease mechanism

doi: 10.1186/s10020-025-01365-5

Figure Lengend Snippet: Neural disease modelling for MED12_WT and MED12_VUS. iPSCs were induced to differentiate into neural progenitor cells and examined for changes in morphology, marker expression, and MED12 protein levels at indicated timepoints. A Light-microscopy images of NPCs at days 18 and day 24 showing bipolar cells with long dendrites. B Flow cytometry gating strategy for iPSCs and NPCs during neural cell differentiation. C and D Bar graphs show the percentage of live cells expressing stem or neural markers, respectively. Timepoints as indicated. MED12_WT (blue), and MED12_VUS, (red). Mixed-model two-way ANOVA with Bonferroni’s multiple comparison test. ( n = 3 group). * p < 0.05; ** p < 0.01. E MED12 western blot in NPCs at day 24 of neural cell differentiation, and F Bar graph indicates MED12 protein expression, normalized to β- Actin expression, in MED12_WT and MED12_VUS NPCs

Article Snippet: Membranes were blocked overnight at 4 °C with Intercept ® (TBS) Blocking Buffer (LI-COR Biosciences), then incubated with rabbit anti-human MED12 monoclonal antibody (1:1000; clone BLR084G; Bethyl Laboratories, USA) and/or β-actin antibody (1:2000; MA5-15729; Life Technologies, Australia).

Techniques: Marker, Expressing, Light Microscopy, Flow Cytometry, Cell Differentiation, Comparison, Western Blot

Journal: Molecular Medicine

Article Title: Functional characterization of the MED12 p.Arg1138Trp variant in females: implications for neural development and disease mechanism

doi: 10.1186/s10020-025-01365-5

Figure Lengend Snippet: Changes in MED12 and components of the MKM during neural cell differentiation. MED12_WT and MED12_VUS iPSCs were stimulated for neural cell differentiation for transcriptomics analysis. A Upset plots showing DEGs common to WT and VUS differentiation at days 18 and 24. B Comparison of NPCs to public NPCs sourced from the ARCHS4 dataset. C GSEA GO-BP geneset enrichment indicates upregulation of neural pathways at day 18 and day 24 in NPCs compared to respective iPSCs. D and E Box plots indicate down-regulation of stem cell markers at and up-regulation of neural cell markers at day 24. F Box plots indicate changes in transcript expression for components of the MKM. G Bar graph shows the change in the MED12/MED12L ratio during differentiation. Boxplots adjusted p-value < 0.05; Bar graph, One tailed, unpaired t-test, p < 0.05

Article Snippet: Membranes were blocked overnight at 4 °C with Intercept ® (TBS) Blocking Buffer (LI-COR Biosciences), then incubated with rabbit anti-human MED12 monoclonal antibody (1:1000; clone BLR084G; Bethyl Laboratories, USA) and/or β-actin antibody (1:2000; MA5-15729; Life Technologies, Australia).

Techniques: Cell Differentiation, Comparison, Expressing, One-tailed Test

Journal: Molecular Medicine

Article Title: Functional characterization of the MED12 p.Arg1138Trp variant in females: implications for neural development and disease mechanism

doi: 10.1186/s10020-025-01365-5

Figure Lengend Snippet: MED12 variant alters gene expression and neural development. MED12_WT and MED12_VUS iPSCs were differentiated into NPCs and transcriptomic profiles of MED12_VUS and MED12_WT NPCs were compared. A Heatmap of top differentially expressed genes. B Treeplot of GO-BP terms enriched when comparing NPCs. Colour indicates normalised enrichment score, representing overall direction of enrichment. Red (positive) represents overall up-regulation, blue (negative) represents overall down-regulation

Article Snippet: Membranes were blocked overnight at 4 °C with Intercept ® (TBS) Blocking Buffer (LI-COR Biosciences), then incubated with rabbit anti-human MED12 monoclonal antibody (1:1000; clone BLR084G; Bethyl Laboratories, USA) and/or β-actin antibody (1:2000; MA5-15729; Life Technologies, Australia).

Techniques: Variant Assay, Gene Expression

Journal: Molecular Medicine

Article Title: Functional characterization of the MED12 p.Arg1138Trp variant in females: implications for neural development and disease mechanism

doi: 10.1186/s10020-025-01365-5

Figure Lengend Snippet: Neural cells carrying the MED12 variant show altered cell growth, specification, and ribosomal complex formation. The MED12_WT and MED12_VUS iPSCs were stimulated for neural cell differentiation, and transcriptomics performed using GSEA and the GO-CC data set. Treeplot demonstrates enriched GO-CC terms when comparing MED12_VUS NPCs to MED12_WT NPCs at days 18 and 24. Terms are clustered based on similarity in gene set. Colour indicates direction of enrichment, with red representing overall upregulation of gene set, blue representing overall downregulation

Article Snippet: Membranes were blocked overnight at 4 °C with Intercept ® (TBS) Blocking Buffer (LI-COR Biosciences), then incubated with rabbit anti-human MED12 monoclonal antibody (1:1000; clone BLR084G; Bethyl Laboratories, USA) and/or β-actin antibody (1:2000; MA5-15729; Life Technologies, Australia).

Techniques: Variant Assay, Cell Differentiation