Journal: Science Advances

Article Title: ARID1A loss in neuroblastoma promotes the adrenergic-to-mesenchymal transition by regulating enhancer-mediated gene expression

doi: 10.1126/sciadv.aaz3440

Figure Lengend Snippet: ( A ) Kaplan-Meier survival analysis according to ARID1A expression with chi-square test. The cutoff value of ARID1A expression levels was determined by the Kaplan scanner tool in R2 web application. ( B ) Correlation analysis between ARID1A and MYCN in human neuroblastomas. Tumors are categorized as MYCN -amplified (blue), MYCN -nonamplified (green), and MYCN status not determined (n.d.) (pink). Correlation coefficients ( r ) and statistical significance levels were determined by linear regression with lm method in R, and data were plotted using ggplot2 in R. Shaded area indicates 95% confidence interval. Data used in (A) and (B) were obtained from Tumor Neuroblastoma SEQC-498-RPM-seqcnb1 in R2 database. ( C ) Cumulative frequency of neuroblastoma onset among stable transgenic and mutant lines by Kaplan-Meier analysis. The overall differences between the curves for various mutant lines versus EGFP;MYCN line are compared by the log-rank test. ( D to O ) Lack of detectable EGFP expression in the IRG of a 17-wpf EGFP transgenic fish (D) and EGFP expression in the tumors (arrows) arising in the IRG of a 45-wpf EGFP;MYCN fish (H) and a 26-wpf arid1aa +/− ;arid1ab +/− ;EGFP;MYCN fish (L). Hematoxylin and eosin (H&E) staining and immunohistochemical staining of tyrosine hydroxylase (TH) of the sagittal sections through the IRG of each fish are shown on the right. E, eye; G, gill; H, heart; I, intestine; IRG, interrenal gland; L, liver. Scale bars, 1 mm (D, E, H, I, L, and M) and 20 μm (F, G, J, K, N, and O). Photo credit: Ting Tao, Dana-Farber Cancer Institute.

Article Snippet: Anti-ARID1A rabbit polyclonal antibody was purchased from Novus Biologicals (#NB100-55334) and used to detect both human ARID1A and zebrafish Arid1aa.

Techniques: Expressing, Amplification, Transgenic Assay, Mutagenesis, Staining, Immunohistochemical staining

Journal: Science Advances

Article Title: ARID1A loss in neuroblastoma promotes the adrenergic-to-mesenchymal transition by regulating enhancer-mediated gene expression

doi: 10.1126/sciadv.aaz3440

Figure Lengend Snippet: ( A ) Western blot analysis of ARID1A protein expression in control (Ctrl), ARID1A +/− , ARID1A −/− #1, and ARID1A −/− #2 NGP cells. α-Tubulin was used as a loading control. ( B ) Cell growth was measured at days 1, 3, 5, and 7. Values are means ± SEM of triplicate experiments. RLU, relative light unit. ( C and D ) Transwell invasion and migration assay of the indicated cells. The fold change of the number of migrated cells passing through the membrane per field was relative to the mean for control NGP cells and was compared with the two-tailed unpaired t test (C). ** P < 0.01; *** P < 0.001. Cells were stained with 0.1% crystal violet (D). Scale bars, 100 μm. ( E ) Immunoprecipitation (IP) of the components of the BAF complexes using an anti-ARID1B antibody. PHF10 is present exclusively in the PBAF complexes and served as a negative control in this experiment; β-actin was used as a loading control. ( F ) Control luciferase shRNA (shLuc) or each of the ARID1B -targeting shRNAs (shARID1B #1 and #2) was induced by doxycycline (1.5 μg/ml) for 6 days in the indicated cells. The expression of ARID1B was detected by western blot analysis. α-Tubulin was used as a loading control. ( G ) Cell growth was measured at days 0, 3, 5, 7, and 9 after pretreatment with doxycycline (1.5 μg/ml) for 3 days to induce the expression of shLuc, shARID1B #1, or shARID1B #2. Values are means ± SEM of triplicate experiments.

Article Snippet: Anti-ARID1A rabbit polyclonal antibody was purchased from Novus Biologicals (#NB100-55334) and used to detect both human ARID1A and zebrafish Arid1aa.

Techniques: Western Blot, Expressing, Control, Migration, Membrane, Two Tailed Test, Staining, Immunoprecipitation, Negative Control, Luciferase, shRNA

Journal: Science Advances

Article Title: ARID1A loss in neuroblastoma promotes the adrenergic-to-mesenchymal transition by regulating enhancer-mediated gene expression

doi: 10.1126/sciadv.aaz3440

Figure Lengend Snippet: ( A ) Fold change (log 2 ) in SS18 ChIP-seq signals in ARID1A −/− #1 ( x value) or ARID1A −/− #2 ( y value) relative to control (Ctrl) NGP cells at each BAF binding site. Sites with weakened (<2/3×) signals in both ARID1A −/− #1 and ARID1A −/− #2 are indicated by blue, while sites with strengthened (>3/2×) signals in both ARID1A −/− #1 and ARID1A −/− #2 are indicated by red; others are indicated by black. ( B ) Fold change (log 2 ) in H3K27ac ChIP-seq signals in ARID1A −/− #1 ( x value) or ARID1A −/− #2 ( y value) relative to control (Ctrl) NGP cells at each promoter and enhancer. Promoters or enhancers with weakened (<1/2×) signals in both ARID1A −/− #1 and ARID1A −/− #2 are indicated by blue, while sites with strengthened (>2×) signals in both ARID1A −/− #1 and ARID1A −/− #2 are indicated by red; others are indicated by black. ( C ) Example of SS18, BRG1, H3K4me3, and H3K27ac ChIP-seq tracks and RNA sequencing (RNA-seq) tracks at TSS-distal BAF weakened leucine rich repeats and immunoglobulin like domains 3 ( LRIG3 ) locus (left) and strengthened immunoglobulin superfamily containing leucine rich repeat 2 ( ISLR2 ) locus (right) in control (red), ARID1A −/− #1 (blue), and ARID1A −/− #2 (black) NGP cells. ( D ) ChIP-seq profiles and heat maps of SS18, BRG1, H3K4me3, and H3K27ac in control (red), ARID1A −/− #1 (blue), and ARID1A −/− #2 (black) NGP cells and of ARID1A in control NGP cells (red), around all TSS-distal BAF binding sites. ( E ) Log 2 fold change of RPKM in RNA-seq between ARID1A mutants and control NGP cells for TSS-distal BAF target genes. Boxes indicate the median (horizontal line), 25th percentile, and 75th percentile; whiskers, distances from the largest and smallest value to each end of the box that are within 1.5× box length; dots, outliers. Data were analyzed with the Mann-Whitney test.

Article Snippet: Anti-ARID1A rabbit polyclonal antibody was purchased from Novus Biologicals (#NB100-55334) and used to detect both human ARID1A and zebrafish Arid1aa.

Techniques: ChIP-sequencing, Control, Binding Assay, RNA Sequencing, MANN-WHITNEY

Journal: Science Advances

Article Title: ARID1A loss in neuroblastoma promotes the adrenergic-to-mesenchymal transition by regulating enhancer-mediated gene expression

doi: 10.1126/sciadv.aaz3440

Figure Lengend Snippet: ( A ) H3K27ac and BRG1 ChIP-seq tracks and RNA-seq tracks at PHOX2B and FN1 loci in control (red), ARID1A −/− #1 (blue), and ARID1A −/− #2 (black) NGP cells. ( B ) Western blot analysis of ARID1A, an adrenergic maker (PHOX2B), and three mesenchymal markers (FN1, SNAI2, and VIM) in control (Ctrl), ARID1A −/− #1, and ARID1A −/− #2 NGP cells. α-Tubulin was used as a loading control. ( C ) mRNA expression (RPKM of RNA-seq) of core regulatory circuitries of transcription factors specific for adrenergic (ADRN core TFs) or mesenchymal (MES core TFs) cells in control (Ctrl), ARID1A −/− #1, and ARID1A −/− #2 NGP cells. ( D ) GSEA to determine the enrichment of a generic gene signature for mesenchymal tumor in ARID1A mutant NGP cells. Genes are ranked by score and plotted along the x axis as vertical black bars. NES, normalized enrichment score; KO, knockout. ( E ) Cell viability [% relative to N , N ′-dimethylformamide (DMF)–treated cells, y value] of control (red), ARID1A −/− #1 (blue), and ARID1A −/− #2 (black). NGP cells were measured after treatment with various concentrations of cisplatin (0 to 20,000 nM, x value) for 72 hours, and half-maximal inhibitory concentration (IC 50 ) values are indicated. Values are means ± SD of triplicate experiments.

Article Snippet: Anti-ARID1A rabbit polyclonal antibody was purchased from Novus Biologicals (#NB100-55334) and used to detect both human ARID1A and zebrafish Arid1aa.

Techniques: ChIP-sequencing, RNA Sequencing, Control, Western Blot, Expressing, Mutagenesis, Knock-Out, Concentration Assay

Journal: Frontiers in Oncology

Article Title: Downregulated ARID1A by miR-185 Is Associated With Poor Prognosis and Adverse Outcomes in Colon Adenocarcinoma

doi: 10.3389/fonc.2021.679334

Figure Lengend Snippet: Differential expression of ARID1A. (A) Differential ARID1A expression analysis across several cancer types (significant differences by the Mann-Whitney U test are marked with red*). Left, normal; right, tumor); (B) ARID1A expression include paired tumor and adjacent normal tissues from Gene chip data; (C) ARID1A expression include paired tumor and adjacent normal tissues from RNA-Seq data; (D) Significant different expression of ARID1A through metastases, normal, and tumors; (E–F) Experimental verification of ARID1A mRNA and protein levels in colon cancer cell line using qPCR and western blot, respectively.

Article Snippet: After blocking with 5% non-fat milk in Tris-buffered saline (1× TBS) containing 0.05% Tween-20, the membrane was incubated overnight at 4°C with rabbit antibodies against ARID1A (1:300, CUSABIO TECHNOLOGY), βactin (1:1000, Beijing Zhongshan Biotechnology Co., Ltd).

Techniques: Quantitative Proteomics, Expressing, MANN-WHITNEY, RNA Sequencing, Western Blot

Journal: Frontiers in Oncology

Article Title: Downregulated ARID1A by miR-185 Is Associated With Poor Prognosis and Adverse Outcomes in Colon Adenocarcinoma

doi: 10.3389/fonc.2021.679334

Figure Lengend Snippet: COAD patients with low ARID1A mRNA levels had a poorer prognosis and predictive value in the pathological stages of COAD. (A) Prognostic value of ARID1A in primary colon adenocarcinoma using Kaplan-Meier plotter from GEPIA2, OS, Overall survival, DFS, Disease free survival; (B) Prognostic value of ARID1A in metastases using Kaplan-Meier plotter from CanEvolve: M-OS, Metastases overall survival, M-DSS, disease-specific survival, and M-DFS, Metastases disease-free survival; (C) Pathologic M, pathologic N, and pathologic T, respectively; (D) Lymphatic invasion, microsatellite instability, and Vital status, respectively. * p < 0.05 and *** p < 0.001.

Article Snippet: After blocking with 5% non-fat milk in Tris-buffered saline (1× TBS) containing 0.05% Tween-20, the membrane was incubated overnight at 4°C with rabbit antibodies against ARID1A (1:300, CUSABIO TECHNOLOGY), βactin (1:1000, Beijing Zhongshan Biotechnology Co., Ltd).

Techniques:

Journal: Frontiers in Oncology

Article Title: Downregulated ARID1A by miR-185 Is Associated With Poor Prognosis and Adverse Outcomes in Colon Adenocarcinoma

doi: 10.3389/fonc.2021.679334

Figure Lengend Snippet: Mir-185 downregulates ARID1A in colon cancer. (A) ARID1A can be controlled by mir-185-5p; (B) Transcription factors that regulate ARID1A expression from Cistrome (Chip-Seq data); (C) Mir-185 is overexpressed in COAD; (D) ARID1A is negatively correlated with mir-185-5p in COAD; (E, F) Mir185 mimics and inhibitors results detected by qPCR; (G) Mir185 mimics and inhibitors results detected by western blot.

Article Snippet: After blocking with 5% non-fat milk in Tris-buffered saline (1× TBS) containing 0.05% Tween-20, the membrane was incubated overnight at 4°C with rabbit antibodies against ARID1A (1:300, CUSABIO TECHNOLOGY), βactin (1:1000, Beijing Zhongshan Biotechnology Co., Ltd).

Techniques: Expressing, ChIP-sequencing, Western Blot

Journal: Frontiers in Oncology

Article Title: Downregulated ARID1A by miR-185 Is Associated With Poor Prognosis and Adverse Outcomes in Colon Adenocarcinoma

doi: 10.3389/fonc.2021.679334

Figure Lengend Snippet: The correlation between ARID1A and immune cell infiltration. (A) The significant positive correlation between ARID1A and subtypes of immune cells (TIMER); (B) Kaplan-Meier curves for the immune infiltrates and ARID1A Expression. The correlation between ARID1A expression and the abundance of CD4+ T cells, B cells, and natural killer cells, respectively (TIMER).

Article Snippet: After blocking with 5% non-fat milk in Tris-buffered saline (1× TBS) containing 0.05% Tween-20, the membrane was incubated overnight at 4°C with rabbit antibodies against ARID1A (1:300, CUSABIO TECHNOLOGY), βactin (1:1000, Beijing Zhongshan Biotechnology Co., Ltd).

Techniques: Expressing

Journal: Frontiers in Oncology

Article Title: Downregulated ARID1A by miR-185 Is Associated With Poor Prognosis and Adverse Outcomes in Colon Adenocarcinoma

doi: 10.3389/fonc.2021.679334

Figure Lengend Snippet: Correlation analysis between ARID1A and gene biomarkers of immune cells in COAD (TIMER).

Article Snippet: After blocking with 5% non-fat milk in Tris-buffered saline (1× TBS) containing 0.05% Tween-20, the membrane was incubated overnight at 4°C with rabbit antibodies against ARID1A (1:300, CUSABIO TECHNOLOGY), βactin (1:1000, Beijing Zhongshan Biotechnology Co., Ltd).

Techniques:

Journal: Journal of Cellular and Molecular Medicine

Article Title: ARID1A downregulation promotes cell proliferation and migration of colon cancer via VIM activation and CDH1 suppression

doi: 10.1111/jcmm.17590

Figure Lengend Snippet: Primer lists and their sequences used in the study

Article Snippet: The membrane was incubated overnight at 4°C with rabbit antibodies against β‐actin (1:1000, Beijing Zhongshan Biotechnology Co., Ltd); ARID1A (1:300, CUSABIO TECHNOLOGY LLC); E‐cadherin Polyclonal antibody, cat on: 20874‐1‐AP; and Vimentin Polyclonal antibody, cat no: 10366‐1‐AP after blocking with 5% non‐fat milk in Tris‐buffered saline containing 0.05%Tween‐20 (1–1000).

Techniques: Sequencing, Negative Control

Journal: Journal of Cellular and Molecular Medicine

Article Title: ARID1A downregulation promotes cell proliferation and migration of colon cancer via VIM activation and CDH1 suppression

doi: 10.1111/jcmm.17590

Figure Lengend Snippet: Identified differentially expressed genes (DEGs)in HCT116‐ARID1A‐wild‐type vs. HCT116‐ARID1A‐knockout (A) Volcano plot of differentially expressed genes (B) Heat map of DEGs in control and knockout groups (C) upper part,KEGG and GO enrichment analysis of differentially upregulated genes. (C) lower part ,KEGG and GO enrichment analysis of differentially downregulated genes

Article Snippet: The membrane was incubated overnight at 4°C with rabbit antibodies against β‐actin (1:1000, Beijing Zhongshan Biotechnology Co., Ltd); ARID1A (1:300, CUSABIO TECHNOLOGY LLC); E‐cadherin Polyclonal antibody, cat on: 20874‐1‐AP; and Vimentin Polyclonal antibody, cat no: 10366‐1‐AP after blocking with 5% non‐fat milk in Tris‐buffered saline containing 0.05%Tween‐20 (1–1000).

Techniques: Knock-Out, Control

Journal: Journal of Cellular and Molecular Medicine

Article Title: ARID1A downregulation promotes cell proliferation and migration of colon cancer via VIM activation and CDH1 suppression

doi: 10.1111/jcmm.17590

Figure Lengend Snippet: Expression of EMT‐related markers (VIM) and correlation with ARID1A (A) Vimentin (VIM) overexpression was found in KO cell lines (B) VIM high expression is associated with a poor prognosis in COAD (C) VIM displays high expression in cells with lower ARID1A expression (SW620&RKO) and low expression in cells with a high ARID1A expression (HCT116&LoVo) (D) colon cancer samples were examined for co‐expression of Vimentin and ARID1A (E) GEPIA2 database visualization of the co‐expression of Vimentin and ARID1A in colon cancer tissue.

Article Snippet: The membrane was incubated overnight at 4°C with rabbit antibodies against β‐actin (1:1000, Beijing Zhongshan Biotechnology Co., Ltd); ARID1A (1:300, CUSABIO TECHNOLOGY LLC); E‐cadherin Polyclonal antibody, cat on: 20874‐1‐AP; and Vimentin Polyclonal antibody, cat no: 10366‐1‐AP after blocking with 5% non‐fat milk in Tris‐buffered saline containing 0.05%Tween‐20 (1–1000).

Techniques: Expressing, Over Expression

Journal: Journal of Cellular and Molecular Medicine

Article Title: ARID1A downregulation promotes cell proliferation and migration of colon cancer via VIM activation and CDH1 suppression

doi: 10.1111/jcmm.17590

Figure Lengend Snippet: Expression of EMT‐related marker (CDH1) and correlation with ARID1A (A) CDH1 is downregulated in ARID1A deficient cells (B) shows a Kaplan–Maier result of CDH1 in COAD (C and D) E‐cadherin expression correlates positively with ARID1A expression in COAD

Article Snippet: The membrane was incubated overnight at 4°C with rabbit antibodies against β‐actin (1:1000, Beijing Zhongshan Biotechnology Co., Ltd); ARID1A (1:300, CUSABIO TECHNOLOGY LLC); E‐cadherin Polyclonal antibody, cat on: 20874‐1‐AP; and Vimentin Polyclonal antibody, cat no: 10366‐1‐AP after blocking with 5% non‐fat milk in Tris‐buffered saline containing 0.05%Tween‐20 (1–1000).

Techniques: Expressing, Marker

Journal: Journal of Cellular and Molecular Medicine

Article Title: ARID1A downregulation promotes cell proliferation and migration of colon cancer via VIM activation and CDH1 suppression

doi: 10.1111/jcmm.17590

Figure Lengend Snippet: ARID1A sequence and mutation profiles in CRC cells used in the research

Article Snippet: The membrane was incubated overnight at 4°C with rabbit antibodies against β‐actin (1:1000, Beijing Zhongshan Biotechnology Co., Ltd); ARID1A (1:300, CUSABIO TECHNOLOGY LLC); E‐cadherin Polyclonal antibody, cat on: 20874‐1‐AP; and Vimentin Polyclonal antibody, cat no: 10366‐1‐AP after blocking with 5% non‐fat milk in Tris‐buffered saline containing 0.05%Tween‐20 (1–1000).

Techniques: Sequencing, Mutagenesis, Variant Assay

Journal: Journal of Cellular and Molecular Medicine

Article Title: ARID1A downregulation promotes cell proliferation and migration of colon cancer via VIM activation and CDH1 suppression

doi: 10.1111/jcmm.17590

Figure Lengend Snippet: ARID1A knockdown in HCT116 cell line (A) At 50 nM, 4 μl siRNA‐ARID1A, qPCR demonstrates no change of vimentin and E‐cadherin expression levels in HCT116 after 24 h transfection (B) ARID1A deficiency at 75 nM, 6 μl siRNA‐ARID1A for 24 h activates VIM and suppress CDH1 in HCT116 (C) qPCR results of ARID1A downregulation using 75 nM, 6‐μl siRNA for 48 h showed VIM and CDH1 expression in HCT116. ARID1A silencing altered VIM and E‐cadherin expression (D and E) VIM and CDH1 expression were altered in qPCR results of LS174‐T‐ARID1A knocked down for 24 and 48 h, respectively. ns, Non‐significant; *p < 0.05, **p < 0.001, and ***p < 0.0001

Article Snippet: The membrane was incubated overnight at 4°C with rabbit antibodies against β‐actin (1:1000, Beijing Zhongshan Biotechnology Co., Ltd); ARID1A (1:300, CUSABIO TECHNOLOGY LLC); E‐cadherin Polyclonal antibody, cat on: 20874‐1‐AP; and Vimentin Polyclonal antibody, cat no: 10366‐1‐AP after blocking with 5% non‐fat milk in Tris‐buffered saline containing 0.05%Tween‐20 (1–1000).

Techniques: Knockdown, Expressing, Transfection

Journal: Journal of Cellular and Molecular Medicine

Article Title: ARID1A downregulation promotes cell proliferation and migration of colon cancer via VIM activation and CDH1 suppression

doi: 10.1111/jcmm.17590

Figure Lengend Snippet: ARID1A silencing alters VIM and E‐cadherin expression (A and B) VIM and CDH1 expression were altered in WB results after ARID1A knocked down for 24 and 48 h.

Article Snippet: The membrane was incubated overnight at 4°C with rabbit antibodies against β‐actin (1:1000, Beijing Zhongshan Biotechnology Co., Ltd); ARID1A (1:300, CUSABIO TECHNOLOGY LLC); E‐cadherin Polyclonal antibody, cat on: 20874‐1‐AP; and Vimentin Polyclonal antibody, cat no: 10366‐1‐AP after blocking with 5% non‐fat milk in Tris‐buffered saline containing 0.05%Tween‐20 (1–1000).

Techniques: Expressing

Journal: Journal of Cellular and Molecular Medicine

Article Title: ARID1A downregulation promotes cell proliferation and migration of colon cancer via VIM activation and CDH1 suppression

doi: 10.1111/jcmm.17590

Figure Lengend Snippet: ARID1A knockdown promotes colon cell line migration and proliferation (A and B) The effect of ARID1A depletion on HCT116 and LS174T cell migration (C) The effect of ARID1A depletion on LS174T cell invasion (D) The proliferation of HCT116 cells was assessed at specified times following ARID1A siRNA transfection (E) Immunoprecipitated DNA from ChIP experiments with anti‐ARID1A antibody was examined by real‐time qPCR using E‐cadherin and Vimentin‐specific promoter primers. IP stands for Immunoprecipitated. *p < 0.05, **p < 0.001, and ***p < 0.0001. Statistical differences were calculated using an imageJ software.

Article Snippet: The membrane was incubated overnight at 4°C with rabbit antibodies against β‐actin (1:1000, Beijing Zhongshan Biotechnology Co., Ltd); ARID1A (1:300, CUSABIO TECHNOLOGY LLC); E‐cadherin Polyclonal antibody, cat on: 20874‐1‐AP; and Vimentin Polyclonal antibody, cat no: 10366‐1‐AP after blocking with 5% non‐fat milk in Tris‐buffered saline containing 0.05%Tween‐20 (1–1000).

Techniques: Knockdown, Migration, Transfection, Immunoprecipitation, Software

Journal: Journal of Cellular and Molecular Medicine

Article Title: ARID1A downregulation promotes cell proliferation and migration of colon cancer via VIM activation and CDH1 suppression

doi: 10.1111/jcmm.17590

Figure Lengend Snippet: List of 87 ARID1A interaction proteins in HCT116

Article Snippet: The membrane was incubated overnight at 4°C with rabbit antibodies against β‐actin (1:1000, Beijing Zhongshan Biotechnology Co., Ltd); ARID1A (1:300, CUSABIO TECHNOLOGY LLC); E‐cadherin Polyclonal antibody, cat on: 20874‐1‐AP; and Vimentin Polyclonal antibody, cat no: 10366‐1‐AP after blocking with 5% non‐fat milk in Tris‐buffered saline containing 0.05%Tween‐20 (1–1000).

Techniques: