Journal: Molecular Cancer Research

Article Title: Mutual Exclusivity Analysis of Genetic and Epigenetic Drivers in Melanoma Identifies a Link Between p14ARF and RARβ Signaling

doi: 10.1158/1541-7786.mcr-13-0006

Figure Lengend Snippet: Figure 3. Cellular responses to ATRA in normal and p14ARF-depleted melanocytes. A, BrdUrd incorporation in primary human melanocytes grown for 48 hours in the presence or absence of ATRA (10 mmol/L) and in the presence or absence of LE135 (1 mmol/L). B, FACS analysis showing representative cell-cycle profiles of normal human melanocytes cultured for 10 days in the presence or absence of ATRA (10 mmol/L). C, ATRA-induced senescence in two cultures of normal human melanocytes (WT-1 and -2) and two independent pools of melanocytes expressing shRNAs against CDKN2A sequences (HEM-1 and -2). Cells were treated with ATRA (10 mmol/L) for 2 weeks, and the number of senescent cells was determined by SA-b-Gal positivity. Columns represent percentage of SA-b-Gal–positive cells. D, the same data as in (C), but with the increase in senescence in normal melanocytes set at 100%. Statistical analysis was conducted using a paired t test. , P < 0.05; ns, nonsignificant.

Article Snippet: LE135 was purchased from Santa Cruz Biotechnology.

Techniques: Cell Culture, Expressing

Journal: Biomedicines

Article Title: Histone H3 Acetylation Is Involved in Retinoid Acid-Induced Neural Differentiation through Increasing Mitochondrial Function

doi: 10.3390/biomedicines11123251

Figure Lengend Snippet: RA promotes histone acetylation through RA signaling during neural differentiation of SH-SY5Y cells. ( a – c ) RT-qPCR analysis of relative mRNA expression of RARα , RARβ , and RARγ in SH-SY5Y cells after RA treatment for 3 and 7 days. n = 3; ** p < 0.01; one-way ANOVA. ( d ) Co-immunoprecipitation was performed in SH-SY5Y cells cultured in maintenance medium or stimulated with RA for 3 and 7 days. Immunoprecipitation was conducted using anti-RARα followed by Western blot analysis of PCAF, CBP, and P300. ( e – g ) Western blot analysis of H3K14ac and H3ac in SH-SY5Y cells treated with DMSO or RA or pretreated with BMS195614/LE135 for 24 h followed by RA for 3 days. n = 3; * p < 0.05; one-way ANOVA.

Article Snippet: To identify the roles of key enzymes and receptors during neural differentiation, the HAT activity inhibitor C646 (Sigma, St. Louis, MO, USA), RARα antagonist BMS195614, and RARβ antagonist LE135 (Glpbio, Montclair, CA, USA) were supplemented during the RA treatment.

Techniques: Quantitative RT-PCR, Expressing, Immunoprecipitation, Cell Culture, Western Blot

Journal: Biomedicines

Article Title: Histone H3 Acetylation Is Involved in Retinoid Acid-Induced Neural Differentiation through Increasing Mitochondrial Function

doi: 10.3390/biomedicines11123251

Figure Lengend Snippet: Inhibition of RA signaling downregulates mitochondrial function and blocks neural differentiation. SH-SY5Y cells were treated with DMSO or RA or pretreated with BMS195614/LE135 for 24 h followed by RA for 3 days. ( a – d ) RT-qPCR analysis of relative mRNA expression of mitochondrial function-related genes ( NDUFS3 , ANT1 , ANT2 , ANT3 ). n = 3; * p < 0.05, ** p < 0.01; one-way ANOVA. ( e ) Changes in morphology and expression of neuronal differentiation markers Tuj1 (green) and MAP2 (red). Scale bar, 50 µm. Images in the black box show cells at a higher magnification. Red or white arrows show representative changes in morphology. ( f – h ) Differentiation rates and mean fluorescence intensity of Tuj1 and MAP2. n = 3; * p < 0.05, ** p < 0.01, *** p < 0.001, **** p < 0.0001; one-way ANOVA. ( i , j ) RT-qPCR analysis of relative mRNA expression of the neural differentiation markers Tuj1 and MAP2 . n = 3; * p < 0.05, ** p < 0.01; one-way ANOVA.

Article Snippet: To identify the roles of key enzymes and receptors during neural differentiation, the HAT activity inhibitor C646 (Sigma, St. Louis, MO, USA), RARα antagonist BMS195614, and RARβ antagonist LE135 (Glpbio, Montclair, CA, USA) were supplemented during the RA treatment.

Techniques: Inhibition, Quantitative RT-PCR, Expressing, Fluorescence