Journal: PLoS ONE

Article Title: In Cortical Neurons HDAC3 Activity Suppresses RD4-Dependent SMRT Export

doi: 10.1371/journal.pone.0021056

Figure Lengend Snippet: A ) GFP-SMRT FL export is insensitive to Leptomycin B and to mutation of a leucine-rich potential nuclear export sequence. Neurons were transfected with plasmids encoding GFP-SMRT FL or GFP-SMRT ΔLeu and treated, where indicated with Leptomycin B (20 ng/ml) for 1 h prior to treatment with TSA. Note that although redistribution was observed after 1–2 h treatment with TSA and other HDAC inhibitors, the effect observed was greater after 6–8 h and so all data presented in this manuscript relates to treatment with the indicated drugs for this time. * p <0.05 (n = 4). B) Neurons were transfected with plasmids encoding GFP-SMRT FL and 48 h after transfection the neurons were treated with TSA, VPA or Apicidin and the cellular localization of GFP-SMRT FL was analyzed. * p <0.05 (n = 3). C) Examples of the cellular localization of GFP-SMRT FL after treatments with the indicated HDAC inhibitors. Scale bar is 20 µm here and throughout the manuscript. D) Example pictures to demonstrate the efficacy of HDAC3-directed siRNA in knocking down endogenous HDAC3 expression in rat cortical neurons. Neurons were transfected with the siRNAs as indicated, plus peGFP to identify transfected cells. After 72 h, cells were fixed and HDAC3 expression analysed by immunofluorescence. White arrows point to transfected neurons. E) HDAC3 siRNA causes redistribution of SMRT to the cytoplasm. Neurons were transfected with plasmids encoding GFP-SMRT FL plus siRNA as indicated. SMRT localization was studied 72 h post-transfection. * p <0.05 (n = 5).

Article Snippet: For each well, 2.67 μl of lipofectamine and 0.65 μg of plasmid DNA was used. siRNA directed against HDAC3 (Santa Cruz sc-270161) or control siRNA (Dharmacon's control non-targeting siRNA #2 siRNA) was used at 100 nM.

Techniques: Mutagenesis, Sequencing, Transfection, Expressing, Immunofluorescence

Journal: Nature communications

Article Title: SCARB2 drives hepatocellular carcinoma tumor initiating cells via enhanced MYC transcriptional activity.

doi: 10.1038/s41467-023-41593-z

Figure Lengend Snippet: Fig. 6 | PMB binds with SCARB2 to suppress HCC by decreasing HDAC3- mediated MYC acetylation and MYC transcriptional activity. a–b Virtual screening of FDA-approved drugs to identify small molecules binding with SCARB2 (a), top 10 hits are listed (b). c The kinetics of the SCARB2-PMB interaction were determined by surface plasmon resonance (SPR) analysis. d The highest scoring docking model of the SCARB2 and PMB complex is shown. Top: surface of the PMB- SCARB2 complex. Bottom: 3D structure of the PMB (yellow)-SCARB2 complex. e The kinetics of the SCARB2-MYC interaction with or without PMB were deter- mined by SPR. f Structured illumination microscopic (SIM) images of vehicle- or PMB- treated HCCLM3 cells (1 h) stained for MYC and SCARB2. Scale bar, 10 μm. g The effect of PMB on the interaction of MYC and SCARB2 was evaluated by Co-IP assays. Extracts of DMSO and PMB-treated HCCLM3 cells were IP with an anti- SCARB2 Ab. h Representative images of MYC/HDAC3 colocalization foci in HCCLM3 cells before and after PMB treatment. Scale bar, 10 μm. i Effect of PMB on MYC acetylation. Extracts of DMSO and PMB-treated HCCLM3 cells were IP with an anti-MYC Ab. Acetylated MYC was detected by immunoblotting. j HCCLM3 cells

Article Snippet: Subsequently, 30-μl droplets containing the indicated dilutions of antibodies specific for MYC (R&D, AF3696, 1:100), SCARB2 (Abcam, ab176317, 1:100) or HDAC3 (Novus, NB500-126, 1:100) were placed on Parafilm in a dark humidified chamber.

Techniques: Activity Assay, Binding Assay, SPR Assay, Staining, Co-Immunoprecipitation Assay, Western Blot

Journal: Medical Oncology (Northwood, London, England)

Article Title: Valeric acid acts as a novel HDAC3 inhibitor against prostate cancer

doi: 10.1007/s12032-022-01814-9

Figure Lengend Snippet: VA suppresses the expression of HDAC3. In both PC-3 and DU145 cells, after 48-h treatment of VA, and the expression of HDAC3 was reduced significantly, compared with NC group ( P < 0.05) ( A – E ). Meanwhile, in both PC-3 and DU145 cells, after 48-h treatment of VA, the expression of HDAC3 protein was reduced, compared with NC group ( F )

Article Snippet: For establishing HDAC3-knockout clones, PC-3 and DU145 cells were both conducted using HDAC3 human gene-knockout kit (Crispr) (OriGene Technologies, USA) following the manufacturer’s instruction as described in our previous study [ ].

Techniques: Expressing

Journal: Medical Oncology (Northwood, London, England)

Article Title: Valeric acid acts as a novel HDAC3 inhibitor against prostate cancer

doi: 10.1007/s12032-022-01814-9

Figure Lengend Snippet: Effect of VA on proliferation of prostate cancer cell lines. VA has anti-proliferative effect on prostate cancer cells while being low toxic to normal cells and such effect could be reduced by HDAC3 gene knockout. The inhibition rates of PC‐3 in the presence of 100 μM VA ranged from 34.42 ± 5.9% to 56.08 ± 1.28% during 24 h to 96 h and displayed significantly higher inhibition rates when compared to the 50-μM VA group with the range from 16.52 ± 2.32% to 24.76 ± 2.32% ( P < 0.001, from 24 to 96 h) ( A ). Similar trends have been displayed in DU145 ( B ). Moreover, for RWPE-1 and RWPE-2, the inhibition rates of 50 μM VA group were significantly lower when compared to 100 μM group, respectively ( C and D ). The relative HDAC3 expression was significantly decreased in HDAC3-KO group than none transfection group in both PC-3 and DU145 cell lines ( E ). The inhibition rates were significantly decreased in CAXII-KO group compared to both non-transfection group and none-KO group, at 24, 48, 72, and 96 h, respectively, in PC-3 and DU145 cell lines ( F and G )

Article Snippet: For establishing HDAC3-knockout clones, PC-3 and DU145 cells were both conducted using HDAC3 human gene-knockout kit (Crispr) (OriGene Technologies, USA) following the manufacturer’s instruction as described in our previous study [ ].

Techniques: Gene Knockout, Inhibition, Expressing, Transfection

Journal: Medical Oncology (Northwood, London, England)

Article Title: Valeric acid acts as a novel HDAC3 inhibitor against prostate cancer

doi: 10.1007/s12032-022-01814-9

Figure Lengend Snippet: The representative 3D spheroid models of PC-3 ( A ) and DU145 ( B ) cells were treated by VA and NC, respectively. The cross-section area inhibition rates eventually climbed to 46.77 ± 19.62% and 48.07 ± 19.72% at 96 h for PC-3 and DU145 cells, respectively ( C ). The CASP3 SA (caspase-3 specific activity) has shown that VA evaluated the caspase-3 activity in both PC-3 and DU145 cells cultured in either 2D or 3D system, compared to respective NC ( P < 0.05) ( D ). Relative expression of E2F1 and E2F3 in VA group was significantly lower that of NC, respectively ( P < 0.001) ( E, F ), NC negative control

Article Snippet: For establishing HDAC3-knockout clones, PC-3 and DU145 cells were both conducted using HDAC3 human gene-knockout kit (Crispr) (OriGene Technologies, USA) following the manufacturer’s instruction as described in our previous study [ ].

Techniques: Inhibition, Activity Assay, Cell Culture, Expressing, Negative Control