Identification and Molecular Characterization of Peroxisome Proliferator-Activated Receptor δ as a Novel Target for...

PPARδ belongs to the peroxisome proliferator-activated receptor (PPAR) family of nuclear receptors. Upon activation by an agonist, PPARδ controls a variety of physiological processes via regulation of its target genes. 15-Deoxy-Δ^12,14-prostaglandin J₂ (15d-PGJ₂) is a cyclopentenone prostaglandin that features an electrophilic, α,β-unsaturated ketone (an enone) in the cyclopentenone ring. Many of 15d-PGJ₂'s biological effects result from covalent interaction between C₉ and the thiol group of a catalytic cysteine (Cys) in target proteins. In this study, we investigated whether 15d-PGJ₂ activates PPARδ by forming a covalent adduct. Our data show that 15d-PGJ₂ activates PPARδ's transcriptional activity through formation of a covalent adduct between its endocyclic enone at C₉ and Cys249 in the receptor's ligand-binding domain. As expected, no adduct formation was seen following a Cys-to-Ser mutation at residue 249 (C249S) of PPARδ or with a PGD₂/PGJ₂ analogue that lacks the electrophilic C₉. Furthermore, the PPARδ C249S mutation weakened induction of the receptor's DNA binding activity by 15d-PGJ₂, which highlights the biological significance of our findings. Calculated chemical properties as well as data from molecular orbital calculations, reactive molecular dynamics simulations, and intrinsic reaction coordinate modeling also supported the selectivity of 15d-PGJ₂'s C₉ toward PPARδ's Cys thiol. In summary, our results provide the molecular, chemical, and structural basis of 15d-PGJ₂-mediated PPARδ activation, designating 15d-PGJ₂ as the first covalent PPARδ ligand to be identified.