Skip to main content
. 2019 Jul 11;294(35):12957–12974. doi: 10.1074/jbc.RA119.008100

Figure 2.

Figure 2.

p53 activates KAISO gene transcription by acting on the p53RE1 located in the 5′-upstream regulatory region. A, four potential p53REs identified in the KAISO gene: 5′-upstream regulatory region, intron 1, exon 2 coding sequence, and 3′-UTR. B, structure of the four pG5-Luc reporter plasmid constructs with the KAISO p53REs and transient transcription assays. The upstream activation sequence (UAS) (bp +1 to +97) was removed from pG5-Luc reporter plasmid. H1299 p53-null cells were transiently co-transfected with pG5–1x(p53RE)-Luc reporter plasmid and WT p53 expression vector. All assays were performed in triplicate. Error bars, S.D. C, HCT116 p53+/+ cells were transfected with pGL2-KAISO-Luc-4.6 kb containing the KAISO promoter (bp −4441 to +165). After treatment with etoposide for 24 h, luciferase activity was measured and normalized to total cellular protein. Data presented are the average of three independent assays. Error bars, S.D. *, p < 0.05. D, transient transcription analysis of the p53REs of the KAISO gene fusion reporter plasmid constructs. pG5–2x(p53RE1)-Luc or pG5–4x(p53RE3)-Luc and WT p53 expression vector or four hot spot p53 mutant expression vectors were transiently co-transfected in H1299 p53-null cells, and luciferase activities were measured. Luciferase activities were normalized to co-expressed β-gal activity, and data are presented as the average of three independent assays. Error bars, S.D. *, p < 0.05; **, p < 0.01; n.s., not significant. WB, Western blotting.