Journal: Molecular Cell

Article Title: Neuronal non-CG methylation is an essential target for MeCP2 function

doi: 10.1016/j.molcel.2021.01.011

Figure Lengend Snippet:

Article Snippet: Anti-NCOR1 antibody, rabbit polyclonal , Bethyl , Cat# A301-146A; RRID: AB_873086.

Techniques: Virus, Recombinant, Derivative Assay, Bisulfite Sequencing, RNA Sequencing, Knock-In, Methylation, Pull Down Assay, Cloning, Software

Journal: Proceedings of the National Academy of Sciences of the United States of America

Article Title: NCoR1-independent mechanism plays a role in the action of the unliganded thyroid hormone receptor

doi: 10.1073/pnas.1706917114

Figure Lengend Snippet: Congenital hypothyroid Pax8−/− mutant mice cannot be rescued by inactivating the NCoR1 through expression of NCoR1∆ID. (A) Percentage of mice of the indicated genotypes surviving until day P15. (B) Tabulated numbers of mice generated of the indicated genotype. Body weight (C) and length (D) of the indicated genotypes (n = 5 or 6 mice per genotype). Data are shown as mean ± SEM; ***P < 0.0001, ****P < 0.0001; ns, not significant; two-way ANOVA.

Article Snippet: Blots were probed for NCoR1 using rabbit polyclonal anti-NCoR1 antibody (A301-145A; Bethyl) or SMRT (06-891; Millipore) overnight at 4° and HRP-conjugated secondary antibody for 1 h at room temperature and were visualized using ECL Prime Western blotting detection agent (GE Healthcare).

Techniques: Mutagenesis, Expressing, Generated

Journal: Proceedings of the National Academy of Sciences of the United States of America

Article Title: NCoR1-independent mechanism plays a role in the action of the unliganded thyroid hormone receptor

doi: 10.1073/pnas.1706917114

Figure Lengend Snippet: Phenotype of L-NCoR1–KO mice. (A) Protein extracts from the liver of the indicated genotypes were blotted and probed with NCoR1-specific (A301-145A; Bethyl) or SMRT-specific (06-891; Millipore) antisera, and the membranes were stripped and reprobed with a RNA-polymerase II (RNApol2) antibody (05-952; EDM Millipore). (B) Total T4 was measured by ELISA in the indicated genotypes. (C) Liver weight-to-body weight ratio (LW/BW), serum cholesterol, and liver triglycerides were measured in the indicated genotypes; n = 6. Data are shown as mean ± SD; *P < 0.05, **P < 0.01, ****P < 0.0001; two-way ANOVA.

Article Snippet: Blots were probed for NCoR1 using rabbit polyclonal anti-NCoR1 antibody (A301-145A; Bethyl) or SMRT (06-891; Millipore) overnight at 4° and HRP-conjugated secondary antibody for 1 h at room temperature and were visualized using ECL Prime Western blotting detection agent (GE Healthcare).

Techniques: Enzyme-linked Immunosorbent Assay

Journal: Proceedings of the National Academy of Sciences of the United States of America

Article Title: NCoR1-independent mechanism plays a role in the action of the unliganded thyroid hormone receptor

doi: 10.1073/pnas.1706917114

Figure Lengend Snippet: Hypothyroidism represses the expression of T3-posivite targets independently of NCoR1. Liver mRNA levels of the indicated type 1 genes (A) or type 2 genes (B) in the shown genotypes were measured by qPCR (n = 6 mice per genotype). (C) Dio1 and Thrsp TRβ1-ChIP peaks (6) visualized in genome browser. Arrows indicate the targeted acetylation sites upstream (+) or downstream (−) of TRβ1-ChIP peaks that were used after ChIP with D. H3K9ac ChIP-PCR was run on chromatin of euthyroid and hypothyroid L-NCoR1–KO mice (n = 3 IP per genotype); anti-H3K9 acetyl (07-352; Millipore). (E) DNA libraries from euthyroid and hypothyroid WT and L-NCoR1–KO mice (n = 3 IP per genotype) were used as template for ChIP-PCR. Anti-H3K27ac (catalog no. 39133; Active Motif) was used for immunoprecipitation (IP). Data are shown as mean ± SD; *P < 0.05, **P < 0.01, ***P < 0.001, ****P < 0.0001; two-way ANOVA.

Article Snippet: Blots were probed for NCoR1 using rabbit polyclonal anti-NCoR1 antibody (A301-145A; Bethyl) or SMRT (06-891; Millipore) overnight at 4° and HRP-conjugated secondary antibody for 1 h at room temperature and were visualized using ECL Prime Western blotting detection agent (GE Healthcare).

Techniques: Expressing, Immunoprecipitation

Journal: Proceedings of the National Academy of Sciences of the United States of America

Article Title: NCoR1-independent mechanism plays a role in the action of the unliganded thyroid hormone receptor

doi: 10.1073/pnas.1706917114

Figure Lengend Snippet: Hypothyroidism represses the expression of T3-positive targets independently of SMRT and NCoR1. (A) Liver mRNA levels of the indicated genes in the shown genotypes were measured by qPCR (n = 5 mice per genotype). (B) H3K27acetyl ChIP-PCR was run on chromatin of euthyroid and hypothyroid L-SMRT–KO and SMRT-KO/NCoR1∆ID mice (n = 3 IP per genotype). Anti-H3K27ac (catalog no. 39133; Active Motif) was used for immunoprecipitation. Data are shown as mean ± SD; *P < 0.05, **P < 0.01, ****P < 0.0001; two-way ANOVA.

Article Snippet: Blots were probed for NCoR1 using rabbit polyclonal anti-NCoR1 antibody (A301-145A; Bethyl) or SMRT (06-891; Millipore) overnight at 4° and HRP-conjugated secondary antibody for 1 h at room temperature and were visualized using ECL Prime Western blotting detection agent (GE Healthcare).

Techniques: Expressing, Immunoprecipitation

Journal: Proceedings of the National Academy of Sciences of the United States of America

Article Title: NCoR1-independent mechanism plays a role in the action of the unliganded thyroid hormone receptor

doi: 10.1073/pnas.1706917114

Figure Lengend Snippet: Ligand-independent activation of T3-negative targets does not require NCoR1. (A) Euthyroid mice were fed chow, and hypothyroid mice were fed a PTU/LID diet for 21 d (n = 6). The liver mRNA levels of the indicated genotypes were measured by qPCR. (B) Euthyroid mice were fed chow, hypothyroid mice were fed a PTU/LID diet, and hyperthyroid mice were fed PTU/LID with T3 (2 μg/100 g) injected daily on last 4 d of the experiment (n = 5 or 6 mice per genotype), and liver mRNA levels were assessed by qPCR. (C) H3K27ac sites near TRβ1 peaks in the regulatory regions of Gsta1, Serpina7, and Vldlr were targeted by ChIP-PCR (n = 4 IP per genotype). Data are shown as mean ± SD; *P < 0.05, **P < 0.01, ***P < 0.001, ****P < 0.0001; two-way ANOVA.

Article Snippet: Blots were probed for NCoR1 using rabbit polyclonal anti-NCoR1 antibody (A301-145A; Bethyl) or SMRT (06-891; Millipore) overnight at 4° and HRP-conjugated secondary antibody for 1 h at room temperature and were visualized using ECL Prime Western blotting detection agent (GE Healthcare).

Techniques: Activation Assay, Injection

Journal: Proceedings of the National Academy of Sciences of the United States of America

Article Title: NCoR1-independent mechanism plays a role in the action of the unliganded thyroid hormone receptor

doi: 10.1073/pnas.1706917114

Figure Lengend Snippet: T3-negative targets retain the response to PTU and T3 in the absence of NCoR1 or TRβ1. (A) Euthyroid mice were fed chow, and hypothyroid mice were fed a PTU/LID diet for 21 d (n = 6 mice per genotype). The liver mRNA levels of the indicated genotypes were measured by qPCR. (B) Euthyroid mice were fed chow, hypothyroid mice were fed PTU/LID, and hyperthyroid mice fed PTU/LID with T3 (2 μg/100 g body weight) injected daily on last 4 d of the experiment (n = 5 or 6 mice per genotype). Liver mRNA levels were assessed by qPCR. Data are shown as mean ± SD; **P < 0.01, ****P < 0.0001; two-way ANOVA.

Article Snippet: Blots were probed for NCoR1 using rabbit polyclonal anti-NCoR1 antibody (A301-145A; Bethyl) or SMRT (06-891; Millipore) overnight at 4° and HRP-conjugated secondary antibody for 1 h at room temperature and were visualized using ECL Prime Western blotting detection agent (GE Healthcare).

Techniques: Injection

Journal: Proceedings of the National Academy of Sciences of the United States of America

Article Title: NCoR1-independent mechanism plays a role in the action of the unliganded thyroid hormone receptor

doi: 10.1073/pnas.1706917114

Figure Lengend Snippet: Hypothyroidism can modulate gene expression in an NCoR1-independent fashion. (A) The heat maps depict changes in gene expression (microarray) and H3K27 acetylation (CHIP-seq) caused by hypothyroidism. (A, 1) Down-regulated genes in livers of control and L-NCoR1ΔID/KO mice. (A, 2) Up-regulated genes in livers of control and L-NCoR1ΔID/KO mice. The yellow-to-red scale indicates a log2 fold change. Mice were fed a PTU/LID diet or chow diet (n = 3). (B, Top Row) The number of genes repressed (yellow) or activated (red) in WT or NCoR1ΔID mice on a chow vs. PTU diet. (Middle Row) The number and percentage of genes that have associated H3K27ac peaks within 100 kb of the TSS. (Bottom Row) The number and percentage of genes that also have intersecting TRβ1 ChIP peaks near (within 1 kb) of the H3K27ac peaks.

Article Snippet: Blots were probed for NCoR1 using rabbit polyclonal anti-NCoR1 antibody (A301-145A; Bethyl) or SMRT (06-891; Millipore) overnight at 4° and HRP-conjugated secondary antibody for 1 h at room temperature and were visualized using ECL Prime Western blotting detection agent (GE Healthcare).

Techniques: Gene Expression, Microarray, ChIP-sequencing, Control

Journal: Proceedings of the National Academy of Sciences of the United States of America

Article Title: NCoR1-independent mechanism plays a role in the action of the unliganded thyroid hormone receptor

doi: 10.1073/pnas.1706917114

Figure Lengend Snippet: NCoR1 controls basal expression of T3-target genes. (A) The heat map depicts changes in gene expression (microarray) and H3K27 acetylation (CHIP-seq) caused by disruption/deletion of NCoR1. (A, 1) Up-regulated genes in livers of euthyroid or hypothyroid mice. (A, 2) Down-regulated genes in livers of euthyroid or hypothyroid mice. The yellow-to-red scale indicates a log2 fold change. Mice were fed a chow or a PTU/LID diet (n = 3). (B) The table displays the number of changed genes in control vs. NCoR1ΔID in the euthyroid (chow diet) or hypothyroid (PTU/LID diet) conditions, the number and percentage of genes that had associated H3K27ac ChIP peaks in control vs. NCoR1-KO mice, and the number and percentage of genes that showed overlap between the TRβ1 ChIP peak and H3K27ac within 100 kb of the TSS.

Article Snippet: Blots were probed for NCoR1 using rabbit polyclonal anti-NCoR1 antibody (A301-145A; Bethyl) or SMRT (06-891; Millipore) overnight at 4° and HRP-conjugated secondary antibody for 1 h at room temperature and were visualized using ECL Prime Western blotting detection agent (GE Healthcare).

Techniques: Expressing, Gene Expression, Microarray, ChIP-sequencing, Disruption, Control

Journal: PLoS ONE

Article Title: SUMOylation of PPARγ by Rosiglitazone Prevents LPS-Induced NCoR Degradation Mediating Down Regulation of Chemokines Expression in Renal Proximal Tubular Cells

doi: 10.1371/journal.pone.0079815

Figure Lengend Snippet: HK-2 cells were transfected with siNCoR to knock down NCoR, or transfected with sicontrol as controls. Two days after transfection, IL-8 and MCP-1 mRNA were quantified after incubated without or with 10 µM RGL for 2 hours followed by stimulation with 1 μg/ml LPS for 4 hours in HK-2 cells and HK-2/NCoR-knockdown cells. (A) Knockdown efficience of NCoR mRNA by siRNA. (B) The expression of IL-8 mRNA in different groups. (C) The expression of MCP-1 mRNA in different groups. The results are representative of three independent experiments. * P <0.05 compared to the control group; # P <0.05 compared to the LPS-treated group.

Article Snippet: The membranes were blocked with TTBS buffer (50 mmol/L Tris HCl, 150 mmol/L NaCl, and 0.05% Tween 20; pH 7.5) containing 5% skim milk for 2 hours at room temperature, followed by incubating overnight at 4°C with a rabbit polyclonal anti-NFκB p50 antibody(1:1000 dilution; Santa Cruz, Inc., CA), a rabbit polyclonal anti-NFκB p65 antibody (1:1000 dilution; Santa Cruz, Inc., CA) or a rabbit polyclonal anti-NCoR antibody (1:1000 dilution; Santa Cruz, Inc., CA).

Techniques: Transfection, Knockdown, Incubation, Expressing, Control

Journal: PLoS ONE

Article Title: SUMOylation of PPARγ by Rosiglitazone Prevents LPS-Induced NCoR Degradation Mediating Down Regulation of Chemokines Expression in Renal Proximal Tubular Cells

doi: 10.1371/journal.pone.0079815

Figure Lengend Snippet: HK-2 cells were treated without or with 10 µM RGL for 2 hours and then stimulation with 1 μg/ml LPS for 30 minutes. Nuclear protein was extracted and NCoR was quantitated by immunoblotting analysis in different groups. The column bar graph shows the means ± SD of values obtained by blot densitometric analysis from three independent experiments. * P <0.05 compared to the control group; # P <0.05 compared to the LPS-treated group.

Article Snippet: The membranes were blocked with TTBS buffer (50 mmol/L Tris HCl, 150 mmol/L NaCl, and 0.05% Tween 20; pH 7.5) containing 5% skim milk for 2 hours at room temperature, followed by incubating overnight at 4°C with a rabbit polyclonal anti-NFκB p50 antibody(1:1000 dilution; Santa Cruz, Inc., CA), a rabbit polyclonal anti-NFκB p65 antibody (1:1000 dilution; Santa Cruz, Inc., CA) or a rabbit polyclonal anti-NCoR antibody (1:1000 dilution; Santa Cruz, Inc., CA).

Techniques: Western Blot, Control

Journal: PLoS ONE

Article Title: SUMOylation of PPARγ by Rosiglitazone Prevents LPS-Induced NCoR Degradation Mediating Down Regulation of Chemokines Expression in Renal Proximal Tubular Cells

doi: 10.1371/journal.pone.0079815

Figure Lengend Snippet: HK-2 cells were transfected with siPIAS1 to knock down PIAS1, or transfected with sicontrol as controls. Two days after transfection, IL-8 and MCP-1 mRNA were quantified after incubated with 10 µM RGL for 2 hours followed by stimulation with LPS 1 μg/ml for 4 hours in HK-2 cells and HK-2/PIAS1-knockdown cells. Quantification of mRNA level was performed by real-time PCR. (A) Knockdown efficience of PIAS1 mRNA by siRNA. (B) The expression of IL-8 mRNA in different groups. (C) The expression of MCP-1 mRNA in different groups. (D) Two days after transfection, Nuclear protein was extracted after incubated with 10 µM RGL for 2 hours followed by stimulation with 1 μg/ml LPS for 30 minutes in HK-2 cells. NCoR protein were quantitated by immunoblotting analysis in different groups. The results are representative of three independent experiments. * P <0.05 compared to the control group; # P <0.05 compared to the LPS-treated group.

Article Snippet: The membranes were blocked with TTBS buffer (50 mmol/L Tris HCl, 150 mmol/L NaCl, and 0.05% Tween 20; pH 7.5) containing 5% skim milk for 2 hours at room temperature, followed by incubating overnight at 4°C with a rabbit polyclonal anti-NFκB p50 antibody(1:1000 dilution; Santa Cruz, Inc., CA), a rabbit polyclonal anti-NFκB p65 antibody (1:1000 dilution; Santa Cruz, Inc., CA) or a rabbit polyclonal anti-NCoR antibody (1:1000 dilution; Santa Cruz, Inc., CA).

Techniques: Transfection, Knockdown, Incubation, Real-time Polymerase Chain Reaction, Expressing, Western Blot, Control