Journal: Clinical Epigenetics

Article Title: MicroRNA-101 is repressed by EZH2 and its restoration inhibits tumorigenic features in embryonal rhabdomyosarcoma

doi: 10.1186/s13148-015-0107-z

Figure Lengend Snippet: MiR-101 is directly targeted by EZH2 in RD cells. a , b Two independent ChIP assays on RD cells 72 h after EZH2 or CTR siRNA transfection showing the recruitment of EZH2 and histone H3 trimethylation on Lys27 (H3K27me3) levels on miR-101-2 promoter region and MCK regulatory regions. SMAD6 was the negative control gene. Rabbit IgG was used as a negative immunoprecipitation control. Histograms represent the percent of immunoprecipitated material relative to input DNA of the two independent experiments. c mRNA levels (RT-qPCR) of pri - miR-101-2 in RD cells 72 h after EZH2 siRNA treatment were normalized to GAPDH levels and expressed as fold increase over CTR siRNA. d Proposed model depicting the interplay between EZH2 and miR-101 in both normal myogenic differentiation ( left ) and eRMS ( right ). In muscle cells, when myogenesis is triggered, miR-101 is upregulated due to the lowering of EZH2 expression. Then, miR-101 directly inhibits EZH2 expression thus enforcing its own expression, driving late skeletal muscle differentiation. In eRMS, this circuit is dysregulated due to EZH2 over-expression, which leads to miR-101 down-regulation, thus maintaining the cells in an undifferentiated and proliferative state

Article Snippet: A human skeletal muscle differentiation model was obtained treating SkMC myoblasts for 14 days with a differentiating medium (DM) with appropriate supplements (C-23161 and C-39366, PromoCell GmbH, Heidelberg, Germany).

Techniques: Transfection, Negative Control, Immunoprecipitation, Quantitative RT-PCR, Expressing, Over Expression