Ontrol. At 48 or 72 hour post-transfection, we quantified the changes in the BORISSchwarzenbach et al. BMC Cancer 2014, 14:796 http://www.biomedcentral.com/1471-2407/14/Page 7 ofAMAGE-A1 429 bp _ ?Actin 202 bp _MCF-BMAGE-A1 429 bp _ ?Actin 202 bp _BCMMAGE-A1 mRNA Expression ( )C80000 60000 40000 20000 2000 1500 1000 500MCF-p=0.p=0.p=0.001 p=0.basalAZATSAAZA + TSABorisFigure 1 Comparison of the MAG
Lyzed its impact on the modifications of histones bound at the MAGE-A1 promoters. To investigate the changes in the histone signature of MAGE-A1 promoter, it was compared in basal MCF-7 cells (no expression of MAGE-A1, Table 1) to the signature in MCF-7 cells stimulated by 5-aza-CdR with/without TSA or transfected with the expression plasmid encoding for BORIS. For these analyses we used antibodie
Lyzed its impact on the modifications of histones bound at the MAGE-A1 promoters. To investigate the changes in the histone signature of MAGE-A1 promoter, it was compared in basal MCF-7 cells (no expression of MAGE-A1, Table 1) to the signature in MCF-7 cells stimulated by 5-aza-CdR with/without TSA or transfected with the expression plasmid encoding for BORIS. For these analyses we used antibodie
L line BCM1, the expression of both genes is opposite: no levels of MAGE-A1 [2^(Ct) 1.07] and high levelsof BORIS [2^(Ct) 24.39]. We transiently transfected expression plasmid encoding BORIS into both cell lines, with negligible transcript levels of MAGE-A1, and quantified endogenous MAGE-A1 mRNA by RT (reverse transcription)-PCR and gel electrophoresis. As depicted in Figure 1, BORIS was able to
L line BCM1, the expression of both genes is opposite: no levels of MAGE-A1 [2^(Ct) 1.07] and high levelsof BORIS [2^(Ct) 24.39]. We transiently transfected expression plasmid encoding BORIS into both cell lines, with negligible transcript levels of MAGE-A1, and quantified endogenous MAGE-A1 mRNA by RT (reverse transcription)-PCR and gel electrophoresis. As depicted in Figure 1, BORIS was able to
Nhibitor TSA on the mRNA expression of MAGE-A1 gene and the other family members (MAGE-A2, -A3 and -A12) in different cell lines. Moreover, we assessed the methylation status of the MAGE-A promoters by sodium bisulfite mappingbefore and after stimulation with the demethylating agent 5-aza-CdR and/or TSA. While the methylation patterns clearly correlated with the basal MAGE RNA transcript levels, u
Ion levels of BORIS in MDA-MB-468, MCF-7 and BCM1 cells by RT-PCR and gel electrophoresis. As expected, we found a similar expression profile of BORIS mRNA (Figure 2) to that detected by quantitative real-time PCR (Table 1). However, gel electrophoresis and quantitative real time showed no and low expression levels of BORIS in MCF-7 cells, respectively, but the tendency was similar. The additional
Nhibitor TSA or after transient transfection of these cells with an expression plasmid encoding for BORIS were separated on an agarose gel. The bar chart shows the relative changes in mRNA expression levels of MAGE-A1 in MCF-7 cells by quantitative real-time PCR. The significant p-values are shown (C). H2O lane serves as a negative control. The housekeeping gene -Actin was selected as an internal