Dependence of Peroxisome Proliferator-activated Receptor Ligand-induced Mitogen-activated Protein Kinase Signaling on Epidermal Growth Factor Receptor Transactivation

Abstract

Peroxisome proliferator-activated receptors (PPARs) are nuclear hormone receptors that function as ligand-activated transcription factors regulating lipid metabolism and homeostasis. In addition to their ability to regulate PPAR-mediated gene transcription, PPARα and γ ligands have recently been shown to induce activation of mitogen-activated protein kinases (MAPKs), which in turn phosphorylate PPARs, thereby affecting transcriptional activity. However, the mechanism for PPAR ligand-dependent MAPK activation is unclear. In the current study, we demonstrate that various PPARα (nafenopin) and γ (ciglitazone and troglitazone) agonists rapidly induced extracellular signal-regulated kinase (Erk) and/or p38 phosphorylation in rat liver epithelial cells (GN4). The selective epidermal growth factor receptor (EGFR) kinase inhibitors, PD153035 and ZD1839 (Iressa), abolished PPARα and γ agonist-dependent Erk activation. Consistent with this, PPAR agonists increased tyrosine autophosphorylation of the EGFR as well as phosphorylation at a putative Src-specific site, Tyr845. Experiments with the Src inhibitor, PP2, and the antioxidant N-acetyl-L-cysteine revealed critical roles for Src and reactive oxygen species as upstream mediators of EGFR transactivation in response to PPAR ligands. Moreover, PPARα and γ ligands increased Src autophosphorylation as well as kinase activity. EGFR phosphorylation, in turn, led to Ras-dependent Erk activation. In contrast, p38 activation by PPARα and γ ligands occurred independently of Src, oxidative stress, the EGFR, and Ras. Interestingly, PPARα and γ agonists caused rapid activation of proline-rich tyrosine kinase or Pyk2; Pyk2 as well as p38 phosphorylation was reduced by intracellular Ca2+ chelation without an observable effect on EGFR and Erk activation, suggesting a possible role for Pyk2 as an upstream activator of p38. In summary, PPARα and γ ligands activate two distinct signaling cascades in GN4 cells leading to MAPK activation.