Rosiglitazone Induces Caveolin-1 by PPARγ-dependent and PPRE-independent Mechanisms: The Role of EGF Receptor Signaling and its Effect on Cancer Cell Drug Resistance

Background: Caveolin-1, a key component of plasma membrane caveolae, has been implicated in the regulation of cancer cell growth and survival. Peroxisome proliferator-activated receptor-y (PPARy) is a ligand-activated nuclear receptor which plays a pivotal role in many cellular processes. Activation of PPARy by its ligand rosiglitazone up-regulates caveolin-1 mRNA and protein in human carcinoma cells. Materials and Methods: We have used specific signaling inhibitors to dissect the mechanisms of caveolin-1 mRNA and protein induction by rosiglitazone, determined by RT-PCR and Western blotting, respectively. ROS generation was measured by flow cytometry and cell survival was determined by the MTT assay. Results: We show that in HT-29 human colon cancer cells the induction of caveolin-1 by rosiglitazone is inhibited by the EGF receptor (EGFR) blacker AG1478. Moreover, rosiglitazone stimulates EGFR phosphorylation, while direct activation of EGFR by EGF up-regulates caveolin-1 mRNA. Inhibitors of Src and the Mekl-Erkl/2 and p38 MAP kinase pathways also inhibit up-regulation of caveolin-1 by rosiglitazone. Furthermore, rosiglitazone stimulates formation of superoxide anions, whereas induction of caveolin-1 expression by rosiglitazone is attenuated by the antioxidant N-acetyl-cysteine. Finally, rosiglitazone increases the resistance of HT-29 cells to doxorubicin and to hydrogen peroxide. The caveolin-1 gene promoter lacks a canonical PPARy response element (PPRE) and a PPRE-reporter construct is not sensitive to EGF or EGFR inhibition. Conclusion: Our findings indicate that up-regulation of caveolin-1 by rosiglitazone requires superoxide formation and the activation of Src, EGFR, and the Mekl-Erkl/2 and p38 MAP kinase pathways. We suggest a novel mode of action of PPARy ligands in the regulation of caveolin-1, and possibly other genes devoid of a PPRE in their promoters, which involves the coordinate activation of PPARy and intracellular signaling pathways.