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. 2015 Jul 23;6(28):25784–25800. doi: 10.18632/oncotarget.4703

Figure 3. Effect of AF4, MLL, AF4-MLL and MLL-AF4 overexpression on SMAD3/SMAD4- and calcitriol/TGFβ-dependent response of the reporter gene constructs A. pN10 B. pN10cdsInJM and C. pGL3cdsInJM.

Figure 3

In order to study inducibility of the constructs by calcitriol/TGFβ, HeLa cells were co-transfected either with the pCGN control vector or with the expression vectors for the nuclear receptors SMAD3 and SMAD4. 16 hours after transfection, cells were incubated without or with calcitriol (50 nM) and TGFβ (1 ng/ml). After 40 hours, luciferase activity was determined. Results are given as RLU (left panels) which include normalization to transfection efficiency and as x-fold induction by SMAD3/SMAD4 over pCGN control (middle panels) or x-fold induction by calcitriol/TGFβ over SMAD3/SMAD4 (right panels). Each experiment was performed in triplicate. Results are presented as mean − S.E.M. of three independent experiments. Two-tailed t-test was performed to determine significant differences in relation to co-transfection of the backbone vector pTarget, indicated by asterisks (*P < 0.05, **P < 0.01, ***P < 0.001).