Coregulation of DNMT1, Sp1, and NF-κB. (A) Western blot analysis of DNMT1 in 293T cells stably transfected with empty vector (Pinco) or Sp1-myc-tagged-Pinco. (B) Western blot analysis of DNMT1 expression in 293T cells transiently transfected with empty vector (Pinco) or NF-κB (p65)-Pinco. (C) Mithramycin A depleted Sp1 expression and down-regulated DNMT1 expression in MV4-11 cells. (D) Mithramycin A down-regulates both Sp1 and DNMT1 expression in AML primary cells. (E) Sp1 shRNA concurrently decreased Sp1 and DNMT1 proteins (left panel: immunoblot gel; right panel: quantification graph). MV4-11 cells were transfected with pSuper.retro.neo + GFP vector with DNA stuffer sequences (vector) or Sp1 shRNAs (1 or 2) and cultured for 2 weeks. Total cell lysates were subjected to Western blot for Sp1 and DNMT1.