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. Author manuscript; available in PMC: 2013 Dec 9.

Published in final edited form as: Mol Cancer Ther. 2011 Mar 10;10(5):10.1158/1535-7163.MCT-10-0836. doi: 10.1158/1535-7163.MCT-10-0836

A). Transcriptional regulation of CCN1 promoter wild type and CCN1-promoter containing different deletions was assessed, using Luciferase-reporter assay. The constructs used were: 1) −565 (pGL3-ΔFOXO3a-Luc), 2) −176 (pGL3-Δp-176-Luc) and 3) −936 (pGL3-ΔAP1-Luc). *P<0.05 versus control cells (transfected with CCN1 full length) and ***P<0.001 versus control cells. B). Diagrammatic representation of the promoter of CCN1 with various selected transcription-binding sites. The FOXO3a binding motif (AAAGCAAACA) is showed in the schema. The plasmid vector containing luciferase reporter gene was used in all the reporter assays. pGL3-CCN1-Luc is the plasmid with full sequence of CCN1 promoter. C). The inhibitory effect of ZOL on CCN1-promoter is mediated by FOXO3a binding site. MDA-MB-231 cells were transfected with pGL3-CCN1-Luc promoter or pGL3-ΔFOXO3a-Luc containing the FOXO3a deletion and then treated with increases concentrations of ZOL for 24h. *P<0.05 versus control cells (untreated) transfected with pGL3-CCN1-Luc; #P<0.05 in cells transfected with pGL3-CCN1-Luc and treated with different concentrations of ZOL relative to control cells (untreated) transfected with the pGL3-CCN1-Luc.

A). Transcriptional regulation of CCN1 promoter wild type and CCN1-promoter containing different deletions was assessed, using Luciferase-reporter assay. The constructs used were: 1) −565 (pGL3-ΔFOXO3a-Luc), 2) −176 (pGL3-Δp-176-Luc) and 3) −936 (pGL3-ΔAP1-Luc). *P<0.05 versus control cells (transfected with CCN1 full length) and ***P<0.001 versus control cells. B). Diagrammatic representation of the promoter of CCN1 with various selected transcription-binding sites. The FOXO3a binding motif (AAAGCAAACA) is showed in the schema. The plasmid vector containing luciferase reporter gene was used in all the reporter assays. pGL3-CCN1-Luc is the plasmid with full sequence of CCN1 promoter. C). The inhibitory effect of ZOL on CCN1-promoter is mediated by FOXO3a binding site. MDA-MB-231 cells were transfected with pGL3-CCN1-Luc promoter or pGL3-ΔFOXO3a-Luc containing the FOXO3a deletion and then treated with increases concentrations of ZOL for 24h. *P<0.05 versus control cells (untreated) transfected with pGL3-CCN1-Luc; #P<0.05 in cells transfected with pGL3-CCN1-Luc and treated with different concentrations of ZOL relative to control cells (untreated) transfected with the pGL3-CCN1-Luc.