FIG 6.
NF-κB family members partner with chromatinized PKC-θ on inducible EMT and CSC genes. (A) CD44high/CD24low CSCs FACS analysis of MCF-IM cells transfected with either mock siRNA, p50 siRNA, or p65 siRNA and subsequently either nonstimulated (NS) or stimulated (ST). (B and C) mRNA expression by real-time PCR of CD44 and IL-6, respectively, of samples as described for panel A. (D and E) p50 and p65 ChIP assays in MCF-IM cells across the CD44 and IL-6 promoters, respectively. (F) p50 and p65 ChIP assays in HMLE-IM cells across the CD44 promoter region. (G) Enrichment of DNA on the CD44 promoter seen when sequential ChIP was performed first with anti-p65 antibody and then with anti-p50 antibody. (H and I) Sequential ChIP as described for panel G, except first with anti-Pol II and then with either anti-p50 (H) or anti-p65 (I) antibody. (J and K) Sequential ChIP as for panel H, except first with either anti-p65 antibody (J) or anti-p50 antibody (K) and then with anti-PKC-θ antibody. All results represent the means ± the standard errors from three independent experiments (n = 3). *, P < 0.05.