Identification and Molecular Characterization of Peroxisome Proliferator-Activated Receptor δ as a Novel Target for Covalent Modification by 15-Deoxy-Δ12,14-prostaglandin J2

PPAR delta belongs to the peroxisome proliferator-activated receptor (PPAR) family of nuclear receptors. Upon activation by an agonist, PPAR delta controls a variety of physiological processes via regulation of its target genes. 15-Deoxy-Delta(12,14)-prostaglandin J(2) (15d-PGJ(2)) is a cyclopentenone prostaglandin that features an electrophilic, alpha,beta-unsaturated ketone (an enone) in the cyclopentenone ring. Many of 15d-PGJ(2)'s biological effects result from covalent interaction between C-9 and the thiol group of a catalytic cysteine (Cys) in target proteins. In this study, we investigated whether 15d-PGJ(2) activates PPAR delta by forming a covalent adduct. Our data show that 15d-PGJ(2) activates PPAR delta's transcriptional activity through formation of a covalent adduct between its endocyclic enone at C-9 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 PPAR delta or with a PGD(2)/PGJ(2) analogue that lacks the electrophilic C-9. Furthermore, the PPAR delta C249S mutation weakened induction of the receptor's DNA binding activity by 15d-PGJ(2), 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-PGJ(2)'s C-9 toward PPAR delta's Cys thiol. In summary, our results provide the molecular, chemical, and structural basis of 15d-PGJ(2)-mediated PPAR delta activation, designating 15d-PGJ(2) as the first covalent PPAR delta ligand to be identified.