Cellular and Biophysical Pipeline for the Screening of Peroxisome Proliferator-Activated Receptor Beta/Delta Agonists: Avoiding False Positives

Figure 1

Principle of the in vitro PPARß/δ transactivation assay. This assay is based on the transient transfection of the 293T cell line with three plasmids: pBIND-PPARß/δ encoding a chimera of Gal4-DBD and PPARß/δ-LBD genes (PPAR/GAL4); pGRE-LUC, which owns one GAL4 response element upstream of a firefly luciferase reporter gene (); and the pRL vector, which constitutively express Renilla luciferase (). The transfected cells express both PPARß/δ and Renilla luciferase constitutively. When the transfected cell is exposed to a molecule that works as a ligand (+ligand), such as GW0742, PPARß/δ moves into the nucleus, binds to GRE-LUC, and triggers the expression of , which is the expected PPARß/δ activation effect in this assay. The reporter-gene expression correlates with the bioactivity of PPARß/δ in the sample. Note. For simplicity, only PPARß/δ monomer binding to GRE has been depicted.