arid1a antibody  (Bio-Techne corporation)

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