Fig. 5: HDAC6 regulates RAD51 expression through Sp1.

A U87MG cells were transfected with control, HDAC1, HDAC2, and HDAC6 siRNA, respectively, for 72 h. The protein levels of RAD51, HDAC1, HDAC2, HDAC6, Ac-tubulin, and tubulin were analyzed by western blotting (upper panels). Result of the quantitative analysis (normalized to tubulin) of RAD51 from three independent experiments is shown (lower panel). B Schematic diagram shows the promoter region used for transcription factor-binding analysis. TSS transcription start site. C The mRNA expression levels of SP1, CEBPB, SRY, MYB, HSF1, HSF2, HLF, GATA1, and GATA2 in normal brain, primary, and recurrent tumor samples (TCGA-GBM dataset). D Distributions of Sp1-ChIP-seq reads mapped to the promoter region of RAD51. Visualization of the Sp1-ChIP-Seq data using the UCSC Genome Browser on Human (hg19) Assembly (https://genome.ucsc.edu/). The red dash line indicates TSS. E U87MG cells were treated with DMSO or 10 μM MPT0B291 following transfection with GFP or GFP-Sp1 plasmid. On the next day of treatment, the mRNA expression of RAD51 in cells was analyzed using qPCR. Quantitative result (normalized to GAPDH) from three independent experiments is shown. F P3 and P3-R cells were transfected with GFP-Sp1, siSp1, siControl, and siHDAC6, respectively, for 72 h. The protein levels of RAD51, Sp1, HDAC6, and tubulin were analyzed using western blotting (a). Quantitative results (normalized to tubulin) of RAD51 in P3 cells (b) and P3-R cells (c) from three independent experiments are shown. G P3-R cells were treated with 10 μM mithramycin A for 24 h. The protein levels of RAD51, γH2AX, Sp1, HDAC6, and tubulin were analyzed by western blotting (a). Quantitative results (normalized to tubulin) of RAD51 (b) and γH2AX (c) from three independent experiments are shown.