Fig. 3: HDAC6 regulates EWSR1-FLI1 and endogenous EWSR1 expression through SP1/P300 binding to their promoters. | Oncogene

Fig. 3: HDAC6 regulates EWSR1-FLI1 and endogenous EWSR1 expression through SP1/P300 binding to their promoters.

From: Selective inhibition of HDAC6 regulates expression of the oncogenic driver EWSR1-FLI1 through the EWSR1 promoter in Ewing sarcoma

Fig. 3

A RT-qPCR analysis of miR-145 levels in SKNMC and WE68 EWS cell lines treated with BML-281. B RT-qPCR analysis of the expression levels of Let-7 family members in SKNMC and WE68 EWS cell lines treated with BML-281 treatment. C RT-qPCR analysis of EWSR1 mRNA expression of the non-translocated gene in SKNMC and WE68 EWS cell lines treated with BML-281. D Evaluation of EWSR1 promoter region-dependent transcriptional activation with luciferase reporter constructs containing the region of 2000 bp upstream of the EWSR1 transcription start site (TSS), in a dose-dependent BML-281-treatment of SKNMC cells for 4 h. E, F ChIP and qPCR analysis of SP1 and/or P300 binding levels at EWSR1 activating promoter region in SKNMC and WE68 EWS cell lines treated with BML-281. % Input indicates enrichment ratio of immunoprecipitated samples relative to input. The IgG antibody was used as a control for unspecific binding in ChIP assays. Binding sites of the primers used are shown schematically (F). G SP1 pull-down and evaluation of lysine acetylated levels against total SP1 protein expression in both SKNMC and WE68 cell lines treated for 24 h with increasing concentrations of BML-281 (IC50 and IC90). Numbers below blots represent densitometric quantification of bands, normalized to endogenous bands (total SP1) and referred to their respective controls (DMSO). *P < 0.05; **P < 0.01; ***P < 0.001.

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