hdac class i ii inhibitor tsa  (MedChemExpress)

Journal: Nature Communications

Article Title: An HDAC6-dependent surveillance mechanism suppresses tau-mediated neurodegeneration and cognitive decline

doi: 10.1038/s41467-020-19317-4

Figure Lengend Snippet: a Immunoblotting of HEK-293A cells cotransfected for 48 h with WT tau, the acetyltransferase CBP, and WT HDAC6 (where indicated) using acetylation or phosphorylation specific tau antibodies. b – d The relative levels of modified tau within the MTBR (ac-280, ac-K311, p-S262, and p-S356) or outside the MTBR (AT8, p-S202, p-T205, and p-S396) were quantified by densitometry of tau PTMs normalized to total tau (Tau 12). e Cells expressing the indicated tau constructs (WT, 4Δ, PL, and PL4Δ) were treated with tubastatin A (TBST) to inhibit HDAC6 or cotransfected with WT HDAC6 to suppress tau acetylation followed by immunoblotting with acetylation or phosphorylation specific tau antibodies as in a . f – l Primary cortical neurons (E16) from PS19 derived tau-transgenic mice cultured for ten DIV were treated overnight with TBST at a concentration of 1–5 μM and analyzed by immunoblotting followed by quantification by densitometry of tau PTMs normalized to human, mouse, or total tau (K9JA). The arrows in f highlight overexpressed human tau migrating at ~65 kDa compared to mouse tau at ~50 kDa (see also Supplementary Fig. to distinguish between human and mouse tau). m Co-IP assay evaluating HDAC6 binding to WT tau, tau-P301L, and tau-P301L/4SE mutant containing S → E substitutions at all four MARK2-targeted phosphorylation sites (S262/S290/S324/S356E). Despite P301L tau showing greater binding to HDAC6, the addition of the MARK2 phosphorylation-mimic mutations reduced HDAC6 binding. n Co-IP assay evaluating tau-P301L mutant tau binding to HDAC6 in the presence of MARK2 or GSK-3β kinases. o HEK-293A cells co-transfected with WT tau, MARK2, or GSK-3β (where indicated) were analyzed by immunoblotting using acetylation or phosphorylation specific tau antibodies followed by quantification of ac-K311 and ac-Tubulin levels normalized to GAPDH levels in p . q HEK-293A cells co-transfected with WT and MARK2 were treated with a panel of HDAC inhibitors including tubastatin A (TBST, HDAC6 inhibitor, 5 μM), trichostatin A (TSA, class I/II HDAC inhibitor, 1 μM) or nicotinamide (NCA, class III HDAC inhibitor, 5 mM) and analyzed as described in o . Statistical tests: p value determined by two-sided unpaired t -test from n = 6 ( b – d ), n = 3 ( g – l ), and n = 4 ( p ) biologically independent experiments. Error bars represent means ± SEM ( b – d , g – l , p ). * p < 0.05; ** p < 0.01; *** p < 0.001; **** p < 0.0001. Source data are provided as a Source Data file.

Article Snippet: The following deacetylase inhibitors (Sigma) were added overnight at the following final concentrations: 1 μM trichostatin A (TSA, class I/II HDAC inhibitor), 5 μM tubastatin A (TBST, HDAC6 inhibitor), 5 mM nicotinamide (NCA, class III HDAC inhibitor).

Techniques: Western Blot, Modification, Expressing, Construct, Derivative Assay, Transgenic Assay, Cell Culture, Concentration Assay, Co-Immunoprecipitation Assay, Binding Assay, Mutagenesis, Transfection

Journal: Cell Death & Disease

Article Title: The epigenetic modifier trichostatin A, a histone deacetylase inhibitor, suppresses proliferation and epithelial–mesenchymal transition of lens epithelial cells

doi: 10.1038/cddis.2013.416

Figure Lengend Snippet: The HDAC inhibitor TSA suppressed proliferation of LECs by cell cycle arrest. ( a ) SRA01/04 and HLEB3 cell lines were treated with TSA at various concentrations (0.1, 0.2, 0.4, and 0.8 μ M) for 24 and 48 h, then the percentage of viable cells was determined by CCK-8 kit. * P< 0.05 versus the control group. ( b ) Cell cycle analysis of SRA01/04 and HLEB3 cell lines were quantified by PI staining followed by flow cytometry analyses after treatment for 24 h. Bar graphs represent the mean ±S.E.M. of three independent experiments. ( c ) The protein expression levels of cyclin D1, cyclin E1, CKD2, CDK4 CDK6, P21 and P27 were detected by western blot analysis after treatment for 24 h in SRA01/04 and HLEB3 cell lines. All experiments were repeated three times with similar results

Article Snippet: TSA (a class I and II HDAC inhibitor) was purchased from Sigma-Aldrich (Louis, MO, USA).

Techniques: CCK-8 Assay, Cell Cycle Assay, Staining, Flow Cytometry, Expressing, Western Blot