Inhibition of EZH2 expression by miRNA-101 and miRNA-26, which are suppressed by MYC in NKTL cells. (A, left) MYC induction by tet-on. MYC overexpression was induced by treating cells with doxycycline. MYC expression was quantified by qRT-PCR analysis. (A, middle) Decreased levels of miR-101, miR-26a, and miR-26b in NKYS upon MYC induction by tet-on. (A, right) MYC induction by tet-on increases EZH2 mRNA levels. (B, left) Depletion of MYC by siRNAs results in induction of miR-101, mir-26a, and miR-26b transcription in NKYS cells. Cells were transfected with MYC siRNA or nontargeting siRNA as a control. Cells were harvested 48 hours after transfection for mRNA analysis of miR-101 and miR-26 gene levels by real-time PCR. (B, right) ChIP-qPCR for endogenous MYC binding to miR-101, miR-26a, and miR-26b genes. Fold enrichment in the ChIP experiment represents the signal obtained after MYC immunoprecipitation followed by qPCR amplified by primer pairs that spanned gene promoters. (C, left) Correlation between MYC activation index and EZH2 mRNA levels. Cross: Normal tissue; Plus sign: Normal NK; Triangle: NKTL; Circle: Cell Lines. R>0.95, P < 2.2*10^–16. (C, right) Scatterplot showing the correlation between IHC MYC staining and EZH2 expression. Spearman correlation coefficient r for MYC v EZH2 = 0.76; P < .0001.