FIGURE 1.

FPP mediates activation and function of GR in human keratinocytes. A, FPP activates GR. Primary human keratinocytes stained with anti-Ser(P)²¹¹ GR antibody reveal localization and activation of GR. This antibody recognizes ligand-induced phosphorylation at Ser²¹¹. The nuclei were visualized by propidium iodide staining. Weak Ser(P)²¹¹ GR immunoreactivity was observed in the cytoplasm and nuclei of untreated cells. In contrast, strong signal was evident in the nuclei of all treated cells: DEX, FPP, and ZGA, which elevates endogenous FPP levels. Mevastatin abolishes ZGA-induced activation of GR by preventing accumulation of endogenous FPP. B, FPP activates GR-mediated transcriptional regulation in keratinocytes. Transfection experiments of primary human keratinocytes with the K6-CAT and GRE-CAT reporters are shown. The data are presented as relative CAT activity, a measure of actual CAT activity normalized for total protein. The results show that DEX, ZGA, or FPP treatment lead to repression of the K6 promoter. Similar to DEX, FPP or ZGA also stimulated the GRE-CAT reporter. C, FPP targets the K6 promoter through GR. HEK were either untreated (−) or treated with 1 μm DEX or 10 μm FPP. Similar amounts of genomic DNA (Input) was used in each treatment. K6 promoter sequences were amplified by PCR after immunoprecipitation with anti-GR antibody or rabbit IgG.