Figure 2

FATS is a transcriptional target of p53. (A) The p53-RE sequence in the promoter of human FATS gene (GenBank accession number: NM_001004298). The sequences of SRY-REs were also shown. (B) The sequence (-746 to +33) around TSS of human FATS was amplified by PCR and a luciferase reporter 746-luc was constructed subsequently. The sequence of a p53-RE was highlighted, and the primer sequences were underlined. (C) 746-luc (0.5 μg) was transfected with or without p53 (0.1 μg) by Lipofactamine (Invitrogen) into U87 cells carrying wild-type p53. A reporter pRL-TK (0.1 μg, Promega) was used as internal control for trasnfection. After 24 h, dual luciferase assay was performed [10], and the firefly lucifease activity derived from 746-luc was normalized by Renilla luciferase activity derived from pRL-TK. Results of three independent experiments were averaged and plotted as relative luciferase activity. Error bars refer to standard deviation (± SD). (D) U87 or MEF cells were treated with 1% formaldehyde for 10 min, respectively, and subjected to chromatin immunoprecipitation (ChIP) analysis. Primer sequences were 5'-CAAGCGATTGTCCTGTCTCA-3' and 5'-GAGACCAGCCTGACCAACAT-3' (human FATS); 5'-CCGCCTCAAAAGGATAAGGT-3' and 5'-GCAGGAACAATTACAACAATATGC-3' (mouse FATS), 5'-GTGTCCACCTCAGGGTGTCT-3' and 5'-CTTCAGGGCTTCGCTTATTG-3' (no p53-RE at hFATS exon 3).