Fig 6.

Ligand activation of PPARβ/δ represses CDK1 and E2F1 by increasing recruitment of p107/130 to E2F4 binding sites. Wild-type and Pparβ/δ-null cells were mock infected or Hras infected for 2 days. Cells were cultured with or without 1 μM GW0742 for 24 h. (A) Western blot analysis of cytosol (columns C) and nuclear (columns N) extracts from HRAS-expressing keratinocytes. Expression levels were normalized to β-actin. The average ratios of nuclear to cytoplasmic protein (N/C) are shown. (B and C) Promoter occupancy of acetylated histone 4 (Ac-H4), E2F1, p130, p107, and E2F4 was examined by ChIP analysis of the mouse CDK1 (B) or E2F1 (C) promoter. For the CDK1 and E2F1 promoters, the distal E2F1 activator binding site and the proximal E2F4 repressor binding site are depicted as two blue boxes. The CHR binding site is depicted as the yellow box. The relative positions of the PCR products used for ChIP analysis are shown by the lines with double arrows. (D) Promoter analysis of the mouse CDK1 promoter. Mutations in the distal activator E2F1 binding site, the proximal repressor E2F4 binding site, and the proximal CHR binding site are illustrated. For all data sets, n = 3 independent samples. Values represent means ± SEM. *, significantly different from HRAS-expressing wild-type control, P ≤ 0.05.