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. 2015 Jan 21;43(3):1609–1625. doi: 10.1093/nar/gkv026

Figure 2.

Figure 2.

ZBTB2 potently represses endogenous RelA/p65 expression in HEK293 cells by acting on the proximal GC-rich promoter element (bp, −30 to −1). (A) Transient transfection and transcription assays in HEK293 cells. HEK293 cells were co-transfected with the pGL2-RelA/p65-Luc reporter constructs indicated (−47; −401Δ(−30 to +44)) and Sp1 and/or ZBTB2 expression vectors. Luciferase activity was measured 48 h after transfection and normalized to total protein concentration. Bars represent standard deviations. *P < 0.05. (B) Western blot analysis of regulation of endogenous RelA/p65 expression by ZBTB2. HEK293 cells were transfected with pcDNA3 or pcDNA3-ZBTB2 expression plasmid or ZBTB2 siRNA. Whole cell extracts were separated by SDS-PAGE and analyzed using the antibodies indicated. GAPDH, control. (C) RT-qPCR analysis of regulation of endogenous RelA/p65 mRNA expression by ZBTB2. Cells were treated as in (B). Total mRNA was isolated from cells and the levels of ZBTB2 and RelA/p65 mRNA determined by RT-qPCR and normalized to 18S ribosomal RNA. *P < 0.05.