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. 2021 Feb 14;11(2):125. doi: 10.3390/jpm11020125

Figure 7.

Figure 7

Regulation of POU5F1 promoter by Elk-1. (A) Schematic diagram of predicted ets motifs ets1-4 on POU5F1 promoter. Motifs ets1-3 were predicted to be stronger binding motifs for Elk-1 and were individually deleted to generate POU5F1∆ets1-Luc, POU5F1∆ets2-Luc, and POU5F1∆ets3-Luc reporter plasmids. (B) Luciferase assay for (i) wildtype POU5F1-Luc and its deletion mutant reporters (ii) POU5F1∆ets1-Luc, (iii) POU5F1∆ets2-Luc, and (iv) POU5F1∆ets3-Luc in SK-N-BE (2) cells after transfection of expression plasmids with Elk1-VP16, Elk1-EN, or empty control plasmid pCDNA3.1 (left graphs) or co-transfection of silencing plasmids for scrRNA control or siElk-1 (right graphs). Luminometric measurements were normalized to Renilla-luc activity. ANOVA, Tukey’s multiple comparative tests, ** p < 0.01, *** p < 0.001 for left graphs (i–iv); unpaired two-tailed t-test, ** p < 0.01 and *** p < 0.001 for right graphs (i–iv). C. Luciferase assay for (i) wildtype POU5F1-Luc and its deletion mutant reporters (ii) POU5F1∆ets1-Luc, (iii) POU5F1∆ets2-Luc, and (iv) POU5F1 ets3-Luc in SH-SY5Y cells after transfection of expression plasmids with Elk1-VP16, Elk1-EN, or empty control plasmid pCDNA3.1 (left graphs) or co-transfection of silencing plasmids for scrRNA control or siElk-1 (right graphs). Luminometric measurements were normalized to Renilla-luc activity. ANOVA, Tukey’s multiple comparative tests, * p < 0.5, ** p < 0.01, *** p < 0.001 for left graphs (i–iv); unpaired two-tailed t-test, * p < 0.5, ** p < 0.01 for right graphs (i–iv). (D) Luciferase assay for (i) wildtype POU5F1-Luc and its deletion mutant reporters (ii) POU5F1∆ets1-Luc, (iii) POU5F1∆ets2-Luc, and (iv) POU5F1∆ets3-Luc in U87-MG cells after transfection of expression plasmids with Elk1-VP16, Elk1-EN, or empty control plasmid pCDNA3.1 (left graphs) or co-transfection of silencing plasmids for scrRNA control or siElk-1 (right graphs). Luminometric measurements were normalized to Renilla-luc activity. ANOVA, Tukey’s multiple comparative tests, * p < 0.5, ** p < 0.01, *** p < 0.001 for left graphs (i–iv); unpaired two-tailed t-test, ** p < 0.01, *** p < 0.001 for right graphs (i–iv).