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. 2016 Jul 8;7(32):51503–51514. doi: 10.18632/oncotarget.10491

Figure 5. Elevated binding of BRD4 to the HOXA9 promoter in lrECM 3D culture.

Figure 5

(A) ChIP assays were carried out using a BRD4-specific antibody in 2D and lrECM 3D cultures of MDA-MB-231 cells on day 4. The BRD4 bound HOXA9 promoter was measured using qPCR and normalized to their corresponding input. A fold change of the BRD4-bound HOXA9 promoter in lrECM 3D culture over 2D culture was obtained by setting the values from 2D culture to one. (B) Similar to part A except that the BRD4-bound HOXA9 promoter was compared between lrECM 3D cultures treated with either JQ1 (250 nM) or DMSO. A fold change of the BRD4-bound HOXA9 promoter in the JQ1 group over the DMSO group was obtained by setting the values from the DMSO group to one. (C) Total cell RNA was extracted from MDA-MB-231 and Hs578T cells in 2D and lrECM 3D cultures on day 4. The mRNA levels of BRD4 were compared between 2D and lrECM 3D cultures using qRT-PCR. A fold change of BRD4 mRNA was obtained by normalizing to the housekeeping gene RPLP0 and setting the values from 2D culture to one. (D) Similar to part C except that the protein levels of BRD4 were measured using immunoblots. When presented, means and standard deviations were obtained from three independent experiments. * and ** indicate a P value < 0.05 and 0.01, respectively.