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Arid1b

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ARID1B is a protein that is part of the SWI/SNF chromatin remodeling complex. It plays a role in regulating gene expression by modifying chromatin structure.

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Lab products found in correlation

Arid1a, by Cell Signaling Technology (3 mentions) Smarca4 brg1, by Santa Cruz Biotechnology (3 mentions) Smarcc2 baf170, by Fortis Life Sciences (3 mentions) Smarcd1 baf60a, by Fortis Life Sciences (3 mentions) Smarce1 baf57, by Fortis Life Sciences (3 mentions) Smarcb1 snf5, by Fortis Life Sciences (3 mentions) Actl6a baf53a, by Fortis Life Sciences (3 mentions) Arid1a, by Santa Cruz Biotechnology (3 mentions) Bca kit, by Thermo Fisher Scientific (3 mentions) Baf57, by Fortis Life Sciences (2 mentions) Arid2, by Santa Cruz Biotechnology (2 mentions) Actin, by Cell Signaling Technology (2 mentions) Smarcc1 baf155 antibody, by Santa Cruz Biotechnology (2 mentions) Arid1a antibody, by Merck Group (2 mentions) Arid1b antibody, by Santa Cruz Biotechnology (2 mentions) Protein g dynabead, by Thermo Fisher Scientific (2 mentions) Ne per nuclear and cytoplasmic extraction kit, by Thermo Fisher Scientific (2 mentions) E cadherin, by Cell Signaling Technology (2 mentions) Protease inhibitor cocktail, by Roche (2 mentions) Ne per nuclear and cytoplasmic extraction reagent, by Thermo Fisher Scientific (2 mentions)

9 protocols using arid1b

1

Investigating Chromatin Remodeling Complexes

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Whole cell extracts of isogenic HCT116 cell lines were used in Western blots for E-Cadherin (Cell Signaling Technology: 24E10), ARID1A (Cell Signaling Technology: 12354), ARID1B (ABCAM ab57461) and ACTIN (Cell Signaling Technology: 5125). Nuclear extracts for co-immunoprecipitation were prepared using the NE-PER Nuclear and Cytoplasmic Extraction Kit (Thermo Scientific #78835). Nuclear extracts were diluted with RIPA buffer (Life Technologies 89900) to a final concentration of 1 mg/ml (with protease inhibitor cocktails, Roche). Each IP was incubated with SMARCC1/BAF155 antibody (Santa Cruz: sc9746), ARID1A antibody (Millipore PSG3), or ARID1B antibody (Santa Cruz 32762) overnight at 4°C. Protein G Dynabeads (Life Technologies 10009D) were added and incubated at 4°C for 3 h. Beads were then washed three times with RIPA buffer and resuspended in reducing SDS gel loading buffer. Antibodies to the following proteins were used in the immunoblots: ARID1A (Cell Signaling Technology: 12354); ARID1B (Abcam: ab54761); SMARCA4/BRG1 (Santa Cruz: sc17796); BRM (Cell Signaling Technology: 11966); SMARCC2/BAF170 (Bethyl Laboratories: A301-039A); SMARCD1/BAF60A (Bethyl Laboratories: A301-595A); SMARCE1/ BAF57 (Bethyl Laboratories: A300-810A); SMARCB1/SNF5 (Bethyl Laboratories: A301-087A); ACTL6A/BAF53A (Bethyl Laboratories: A301-391A); ACTIN (Cell Signaling Technology: 5125, 1:3,000).
Mathur R., Alver B.H., San Roman A.K., Wilson B.G., Wang X., Agoston A.T., Park P.J., Shivdasani R.A, & Roberts C.W. (2016). ARID1A loss impairs enhancer-mediated gene regulation and drives colon cancer in mice. Nature genetics, 49(2), 296-302.
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2

Investigating Chromatin Remodeling Complexes

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Whole cell extracts of isogenic HCT116 cell lines were used in Western blots for E-Cadherin (Cell Signaling Technology: 24E10), ARID1A (Cell Signaling Technology: 12354), ARID1B (ABCAM ab57461) and ACTIN (Cell Signaling Technology: 5125). Nuclear extracts for co-immunoprecipitation were prepared using the NE-PER Nuclear and Cytoplasmic Extraction Kit (Thermo Scientific #78835). Nuclear extracts were diluted with RIPA buffer (Life Technologies 89900) to a final concentration of 1 mg/ml (with protease inhibitor cocktails, Roche). Each IP was incubated with SMARCC1/BAF155 antibody (Santa Cruz: sc9746), ARID1A antibody (Millipore PSG3), or ARID1B antibody (Santa Cruz 32762) overnight at 4°C. Protein G Dynabeads (Life Technologies 10009D) were added and incubated at 4°C for 3 h. Beads were then washed three times with RIPA buffer and resuspended in reducing SDS gel loading buffer. Antibodies to the following proteins were used in the immunoblots: ARID1A (Cell Signaling Technology: 12354); ARID1B (Abcam: ab54761); SMARCA4/BRG1 (Santa Cruz: sc17796); BRM (Cell Signaling Technology: 11966); SMARCC2/BAF170 (Bethyl Laboratories: A301-039A); SMARCD1/BAF60A (Bethyl Laboratories: A301-595A); SMARCE1/ BAF57 (Bethyl Laboratories: A300-810A); SMARCB1/SNF5 (Bethyl Laboratories: A301-087A); ACTL6A/BAF53A (Bethyl Laboratories: A301-391A); ACTIN (Cell Signaling Technology: 5125, 1:3,000).
Mathur R., Alver B.H., San Roman A.K., Wilson B.G., Wang X., Agoston A.T., Park P.J., Shivdasani R.A, & Roberts C.W. (2016). ARID1A loss impairs enhancer-mediated gene regulation and drives colon cancer in mice. Nature genetics, 49(2), 296-302.
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3

Immunoblotting of Chromatin Remodeling Proteins

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Immunoblotting was performed as previously described17 (link). Antibodies to the following proteins were used in the immunoprecipitation and immunoblots: SMARCC1/BAF155 (1:3000 dilution, Santa Cruz: sc9746); ARID1A (1:3000 dilution, Cell Signaling Technology: 12354 and home-made); ARID1B (1:1000 dilution, Abcam: ab54761); SMARCA4/BRG1 (1:500 dilution, Santa Cruz: sc17796 and home-made); SMARCC2/BAF170 (1:3000 dilution, Bethyl Laboratories: A301-039A); SMARCD1/BAF60A (1:3000 dilution, Bethyl Laboratories: A301-595A); SMARCE1/ BAF57 (1:3000 dilution, Bethyl Laboratories: A300-810A); SMARCB1/SNF5 (1:3000 dilution, Bethyl Laboratories: A301-087A); ACTL6A/BAF53A (1:3000 dilution, Bethyl Laboratories: A301-391A); ACTIN-HRP (1:3000 dilution, Santa Cruz: sc-47778).
Wolf B.K., Zhao Y., McCray A., Hawk W.H., Deary L.T., Sugiarto N.W., LaCroix I.S., Gerber S.A., Cheng C, & Wang X. (2022). Cooperation of chromatin remodeling SWI/SNF complex and pioneer factor AP-1 shapes 3D enhancer landscapes. Nature structural & molecular biology, 30(1), 10-21.
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4

Characterizing Chromatin Remodeling Factors

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ARID1A (Sigma-Aldrich #HPA005456, western blot and IHC), ARID1A (Santa Cruz #sc-32761, western blot), ARID1B (Abcam #AB57461, western blot and IHC), ARID2 (Abiocode #R2380–1, western blot), Brg1 (Santa Cruz #sc-10768, western blot), BAF170 (Santa Cruz #sc-17838, western blot), BAF155 (Santa Cruz #sc-9746, western blot), BAF60b (Santa Cruz #sc-101162, western blot), BAF57 (Bethyl #A300–810A, western blot), BAF53 (Santa Cruz #sc-137063, western blot), BAF47 (Santa Cruz #sc-166165, western blot), BAF47 (CST #91735, western blot), BAF45d (Santa Cruz #sc-101106, western blot), Ki-67 (Abcam #AB15580, IHC), HNF4A (CST #3113, IHC), CK19 (Abcam #AB15463, IHC), EpCAM (CST #14452, IHC), Flag (CST #2368, western blot), Ty1 (Diagenode #C15200054, western blot and ChIP).
Wang Z., Chen K., Jia Y., Chuang J.C., Sun X., Lin Y.H., Celen C., Li L., Huang F., Liu X., Castrillon D.H., Wang T, & Zhu H. (2020). Dual ARID1A/ARID1B loss leads to rapid carcinogenesis and disruptive redistribution of BAF complexes. Nature cancer, 1(9), 909-922.
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5

Knockout Cell Lines Viability Assay

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We purchased HAP1 knockout cell lines from Horizon Discovery in August 2015 (Cambridge, UK) and the knockouts were confirmed using Western blot analysis. Cells were plated in triplicate at a density of 2500 cells/well, and grown in complete IMDM glutamax media (Gibco) supplemented with 10% FBS and 1% penicillin and streptomycin, in 5% CO2 at 37°C. We treated the cell lines with varying doses of docetaxel and paclitaxel. Cell viability was evaluated after 3, 4, and 8 days of treatment using CellTitre-Glo® (Promega) luminescent assay on a Victor X5 plate reader (Perkin Elmer). We used commercial antibodies: SMARCA4 (Santa Cruz sc-17796), SMARCA2 (Abcam ab15597), ARID1B (Abcam ab57461, gamma-tubulin (Sigma T5326), ARID1A (Santa Cruz sc-32761), ARID2 (Santa Cruz sc-166117), PBRM1 (Bethyl A-301-591A-M), and Lamin B1 (Abcam ab16048). We used Memcode Protein Stain Kit (Thermo Scientific) to detect transferred proteins to nitrocellulose membranes. We used secondary antibodies conjugated with Horseradish peroxidase (Interchim) and revealed the signal by chemiluminescence substrate from Pierce (SuperSignal West Pico).
Gurard-Levin Z.A., Wilson L.O., Pancaldi V., Postel-Vinay S., Sousa F.G., Reyes C., Marangoni E., Gentien D., Valencia A., Pommier Y., Cottu P, & Almouzni G. (2016). Chromatin regulators as a guide for cancer treatment choice. Molecular cancer therapeutics, 15(7), 1768-1777.
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6

Comprehensive Chromatin Remodeling Protein Analysis

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Primary antibodies used for western blotting are as follows: ARID1A (Santa Cruz Biotechnology, sc-32761), ARID1B (Abcam, ab57461), TBP (Thermo Scientific, MA1-21516), Brg1 (Santa Cruz Biotechnology, sc-17796), Brm (Bethyl Laboratories, A301-015A), BAF60A (Santa Cruz Biotechnology, sc-135843), BAF57 (Bethyl Laboratories, A300-810A), BAF53A (Novus Biologicals, NB100-61628), BAF180 (Bethyl Laboratories, A301-591A), MET (Cell Signaling Technology, #8198), V5 (Biorad, MCA1360). Antibodies used for ChIP: H3K27ac (Abcam, ab4729), H3K4me (Abcam, ab8895), H3K4me3 (Millipore, #05–745), H3K27me3 (Active Motif, #39155), FRA1 (Santa Cruz Biotechnology, sc-183x).
Kelso T.W., Porter D.K., Amaral M.L., Shokhirev M.N., Benner C, & Hargreaves D.C. (2017). Chromatin accessibility underlies synthetic lethality of SWI/SNF subunits in ARID1A-mutant cancers. eLife, 6, e30506.
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7

Quantitative Immunoblotting Analysis

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HRP-conjugated and fluorescent secondary antibody signals were captured by Odyssesy Fc Dual Mode Imaging System (Li-COR). Immunoblot signals were quantified using Image Studio Lite (Ver 3.1). Following antibodies were purchased: Arid1b (Abcam, ab57461, 1:1000), HDAC4 (Abcam, ab111318, 1:1000), β-catenin (BD Transduction, 610154, 1:2000), pβ-catenin (Ser-552, Cell Signaling, 9566, 1:1000), pβ-catenin (Ser-675, Cell Signaling, 4176, 1:1000), GAPDH (Cell Signaling, 2118L, 1:2000), and PSD-95 (NeuroMab, 75-028, 1:1000), SynGAP (Invitrogen, PA1-046, pan-SynGAP antibody, 1:1000), Arc (Synaptic system, 156 003, 1:500), MEF2A (Cell Signaling, 9736S, 1:500), pMEF2A (Ser-408, Cell Signaling, 9737S, 1:500), HRP conjugated anti-Rb IgG (Jackson, Cat # 711-035-152, 1:10,000), HRP conjugated anti-Ms IgG (Jackson, Cat # 715-035-150, 1:10,000), IRDye 800CW conjugated anti-Ms IgG (Li-Cor, Cat # 926-32212, 1:10000).
Kim H., Kim D., Cho Y., Kim K., Roh J.D., Kim Y., Yang E., Kim S.S., Ahn S., Kim H., Kang H., Bae Y, & Kim E. (2022). Early postnatal serotonin modulation prevents adult-stage deficits in Arid1b-deficient mice through synaptic transcriptional reprogramming. Nature Communications, 13, 5051.
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8

Western Blot Analysis of Chromatin Remodelers

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Cells were harvested with 0.05% tryspin/0.02% EDTA solution, and cell pellets washed with cold PBS. Proteins were extracted using NE-PER Nuclear and Cytoplasmic Extraction Reagents (Thermo Scientific catalog # 78835) as per manufacturer’s instructions. Protein concentrations were estimated by BCA kit (Thermo Scientific) on a microplate reader (SpectraMax M5). Comparable amounts of proteins in the nuclear extract were subjected to SDS-PAGE using NuPAGE 4%–12% Bis–Tris gradient gels (Invitrogen) and were transferred to PVDF membranes. Following overnight incubation with primary antibody at 4°C, membranes were incubated with goat anti-rabbit or goat anti-mouse secondary antibodies coupled to IRDye fluorophores for 1 hour at room temperature, and fluorescence was detected using Odyssey CLx Imaging System (LI-COR Biosciences). The following primary antibodies were used at 1:1000 dilution: ARID1A (Bethyl Labs # A301-040A), ARID1B (Abcam # ab57461), ARID2 (Bethyl Labs # A302-230A), SMARCB1 (BD Biosciences # 612111), SMARCC1 (Santa Cruz Biotechnology # sc-32763), β-ACTIN (Sigma # A2228).
Saqcena M., Leandro-Garcia L.J., Maag J.L., Tchekmedyian V., Krishnamoorthy G.P., Tamarapu P.P., Tiedje V., Reuter V., Knauf J.A., de Stanchina E., Xu B., Liao X.H., Refetoff S., Ghossein R., Chi P., Ho A.L., Koche R.P, & Fagin J.A. (2020). SWI/SNF complex mutations promote thyroid tumor progression and insensitivity to redifferentiation therapies.. Cancer discovery, 11(5), 1158-1175.
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9

Western Blot Analysis of Chromatin Remodelers

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Total protein was extracted from cells using cell lysis RIPA buffer and supplemented with a phosphatase and protease inhibitor cocktail; 60 µg of protein lysate was resolved by SDS–polyacrylamide gel electrophoresis (SDS-PAGE; 6%) and transferred to PVDF membranes, which were blocked with 5% non-fat milk in PBS-T and incubated overnight with the indicated antibody. The primary antibodies used are the following: anti-SMARCA4 (sc-17796, Santa Cruz Biotechnology, Dallas, TX, USA), SMARCA2 (D9E8B-XP, Cell Signaling Technology, Danvers, MA, USA), ARID1A (D2A8U, Cell Signaling Technology), ARID1B (ab57461, Abcam, Cambridge, UK), ARID2 (sc-166117, Santa Cruz Biotechnology), BRD9 (A303-781A, Bethyl Laboratories), and anti-BETA-ACTIN (A5441, Sigma Aldrich, Merck KGaA, Darmstadt, Germany). The membranes were then treated with HRP-conjugated anti-rabbit or anti-mouse secondary antibodies (P0448 and P0447, respectively, Dako, Agilent Technologies, Santa Clara, CA, USA). The target protein bands were visualized using Clarity Western ECL Substrate (BioRad, Hercules, CA, USA) and ImageQuant LAS4000 (GE Healthcare, Chicago, IL, USA). Protein bands were quantified using ImageJ software. We normalized the data with ACTB values and we transformed the data by applying the ‘log2 (normalized value +1)’.
Peinado P., Andrades A., Cuadros M., Rodriguez M.I., Coira I.F., Garcia D.J., Álvarez-Perez J.C., Baliñas-Gavira C., Arenas A.M., Patiño-Mercau J.R., Sanjuan-Hidalgo J., Romero O.A., Montuenga L.M., Carretero J., Sanchez-Cespedes M, & Medina P.P. (2020). Comprehensive Analysis of SWI/SNF Inactivation in Lung Adenocarcinoma Cell Models. Cancers, 12(12), 3712.
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