Journal: Cells

Article Title: Identification of PARP-1, Histone H1 and SIRT-1 as New Regulators of Breast Cancer-Related Aromatase Promoter I.3/II

doi: 10.3390/cells9020427

Figure Lengend Snippet: PARP-1 functionally interacts with histone deacetylases (HDACs). If not indicated otherwise, cells were stimulated with forskolin. ( A ) pGL3-PII-522 wt-transfected 3T3-L1 cells were incubated with n-butyrate, without or with PJ34 ( n = 6; * p < 0.05 versus n-butyrate only; # p < 0.05 versus controls). ( B ) 3T3-L1 cells transfected with pGL3-PII-522 wt or pGL3-PII-522(T-241C) were incubated with panobinostat ( n = 8; * p < 0.05 versus no panobinostat; # p < 0.05 wt versus T-241C), or ( C ) with selisistat alone or in combination with PJ34 ( n = 8; * p < 0.05 versus no PJ34; # p < 0.05 wt versus T-241C). ( D , E ) Aromatase mRNA-expression or activity was measured in BAFs, which were treated with selisistat without or with PARP-1 inhibitor PJ34. For better comparison, the data from BAFs not treated with selisistat (control) were taken from E–H (control). SIRT-1 inhibition alone reduced aromatase expression and activity. ( D ) Control n = 3, selisistat n = 6; # p < 0.05 control versus selisistat; ( E ) n = 3; * p < 0.05 versus no selisistat; # p < 0.05 control versus selisistat). ( F ) The increased NAD + /NADH ratios were measured in forskolin-stimulated BAFs with the inhibition of PARP-1 (5 µM PJ34) or/and SIRT-1 (200 nM selisistat) ( n = 3; * p < 0.05 versus forskolin alone).

Article Snippet: Furthermore, the cells were treated with PARP-1 inhibitor PJ34 (Selleck Chemicals S7300, Houston, TX, USA), HDAC class I/IIa inhibitor n-butyrate (Sigma B5887, Taufkirchen, Germany), HDAC class I/II/IV inhibitor Panobinostat (LBH589, Selleck Chemicals S1030) and SIRT-1 inhibitor selisistat (EX527, Selleck Chemicals S1541).

Techniques: Transfection, Incubation, Expressing, Activity Assay, Comparison, Control, Inhibition

Journal: BMC Cancer

Article Title: Analysis of oncogenic activities of protein kinase D1 in head and neck squamous cell carcinoma

doi: 10.1186/s12885-018-4965-6

Figure Lengend Snippet: Effects of DNA methyltransferase inhibitor (Aza) and HDAC inhibitor (SAHA) on PKD1 expression. UPCI15B and UMSCC-1 cells were treated with SAHA and 5-aza-dC alone or in combination for 48 h. Cells were harvested for mRNA extraction and Western blotting. Levels of PKD1 transcripts were determined by real time qRT-PCR ( a ). Protein expression was analyzed by immunoblotting for PKD1 ( b and c ). GAPDH was used as loading control in both experiments. Representative data from one of three independent experiments are shown

Article Snippet: The histone deacetylase HDAC inhibitor suberoylanilide hydroxamic acid (SAHA) was purchased from Cayman Chemical (Ann Arbor, MI).

Techniques: Expressing, Extraction, Western Blot, Quantitative RT-PCR, Control

Journal: Cell Reports

Article Title: SIN3A histone deacetylase action counteracts MUS81 to promote stalled fork stability

doi: 10.1016/j.celrep.2024.113778

Figure Lengend Snippet: Sin3A prevents fork breakage in stressed conditions (A) Images of cells immunostained for chr-bound RAD51 (green) and FANCD2 (red) proteins. DNA stained with DAPI (blue). Scale bar, 10 μm. siRNAs and HU as indicated. Plots show number of FANCD2 (left) or RAD51 (right) foci per cell. Mean in black. Data are pooled from 3 different assays. >1,500 cells scored per condition. ∗∗∗ p < 0.0001; two-tailed Mann-Whitney test. Immunoblot detection of Sin3A. H3, loading control. (B) Images of cells immunostained for γH2AX (red) and chr-bound RPA (green) proteins. DNA stained with DAPI (blue). Scale bar, 25 μm. Treatment as in (A). Histograms show the percentage (mean + SD) of γH2AX (top), chr-bound RPA-positive cells (left), and double-positive cells. n = 3. >400 cells scored per condition and assay. Negative staining determined in untreated control cells. ∗ p = 0.0346 (top) and p = 0.0231 (bottom left); ∗∗ p = 0.0037; unpaired two-tailed Student’s t test. (C) Images of U2OS SEC-C (cells stably expressing Cas9) cells immunostained for γH2AX (red) protein. DNA stained with DAPI (blue). Scale bar, 10 μm. RNA guides (72 h) and HU (3 mM, 4 h) as indicated. Immunoblot detection of Sin3A in indicated samples. GAPDH, loading control. Plot shows distribution of γH2AX intensity values. Median in black. Data are pooled from 2 different assays. >140 cells scored per condition. ∗∗∗ p < 0.0001; two-tailed Mann-Whitney test. (D) Plot shows distribution of γH2AX intensity values in cells treated (4 h) with HU (3 mM) combined with sodium butyrate (NaB, 5 mM), trichostatin A (TSA, 250 nM), and romidepsin (50 nM). Median in black. Data are pooled from 4, 2, and 3 different assays, respectively. >1,100 cells scored per condition. ∗∗∗ p < 0.0001; two-tailed Mann-Whitney test. (E) Images of cells immunostained for chr-bound 53BP1 (green) protein. DNA was stained with DAPI (blue). Scale bar, 10 μm. siRNAs and HU as indicated. Plot shows number of 53BP1 foci per cell. Data are pooled from 3 different assays. >1,400 cells were scored per condition. ∗∗∗ p < 0.0001; two-tailed Mann-Whitney test. (F) Representative images of comet assay. Scale bar, 100 μm. siRNAs and HU as indicated. Histogram shows tail moment (mean + SD). n = 3. ∗∗ p = 0.0067; two-tailed unpaired Student’s t test. (G) Same as in (F) in indicated samples. Scale bar, 100 μm. Histogram shows tail moment (mean + SD). n = 4. n.s. p = 0.1087; two-tailed unpaired Student’s t test. Immunoblot detection of MTA2. GAPDH, loading control. siMTA2, MTA2 siRNA-transfected cells. HU (3 mM, 24 h) except for (C) and (D). siRNA transfection (72 h). All replicates are biological replicates. See also Figure S2 .

Article Snippet: Romidepsin (Class I HDACi) , Selleckchem , FR228.

Techniques: Staining, Two Tailed Test, MANN-WHITNEY, Western Blot, Control, Negative Staining, Stable Transfection, Expressing, Single Cell Gel Electrophoresis, Transfection

Journal: Cell Reports

Article Title: SIN3A histone deacetylase action counteracts MUS81 to promote stalled fork stability

doi: 10.1016/j.celrep.2024.113778

Figure Lengend Snippet:

Article Snippet: Romidepsin (Class I HDACi) , Selleckchem , FR228.

Techniques: Recombinant, Control, Protease Inhibitor, Imaging, Reverse Transcription, Plasmid Preparation, Software, Magnetic Beads, Blocking Assay, In Situ, Western Blot, Membrane