Regulation of C/EBPα gene expression by canonical Notch signaling pathway. Real-time PCR (A) and immunoblot analyses (B) of C/EBPα gene expression in control vector or Cre-infected RBP/J^lox/lox (R KO) 1°MK cultured in low or high Ca²⁺ media. (C) Same experiment as in A, except NICD-induced C/EBPα gene expression is measured. All experiments were performed three or more times, error bars represent standard deviation, and quantifications and details were as described in Fig. 2. (D) Real-time PCR and immunoblot analyses of C/EBPα gene expression in E18.5 WT versus K14-Cre/RBP/J^lox/lox cKO (R cKO) epidermis in vivo. (E) C/EBPα immunofluorescence. Bars, 20 μm. (F) WT and DKO keratinocytes were infected with lentivirus harboring either an empty vector (control) or a vector expressing a shRNA targeting ∼60% RBP/J knockdown. Infected cells were selected with puromycin and real-time PCR analyses were performed on these cells cultured at high Ca²⁺ conditions.