Figure 2

Conservation of the KLK3 ( PSA ) promoter and ChIP verification of WT1 and SP1 binding. (A) Alignment of predicted TFBS (based on human sequences) in the KLK3 gene promoter of multiple genomes shows the conservation of two overlapping WT1 binding sites (solid box), an EGR1 site (dashed box), an SP1 site (dash-dotted box), and an SP2 site (double dash-dotted box). WT1 sites (human 1332–1348 and 1336–1352) are conserved between human, chimpanzee, macaque, and cow and they overlap an SP2 site (human 1347–1361) conserved between human, chimpanzee, and cow. An EGR1 site (human 1400–1416) overlaps an SP1 site (human 1404–1418) and both are conserved between human, chimpanzee, macaque, and dog. (B) The binding of WT1 and SP1 TFs to native chromatin obtained from WT1-transfected LNCaP cells was confirmed by ChIP. Lane 1 shows the no DNA PCR control and lane 2 shows PCR amplified input DNA. Lanes 3, 4, and 5 show PCR amplified DNA immunoprecipitated by IgG (no antibody control), SP1 or WT1 antibodies, respectively.