Docosahexaenoic Acid Inhibits Superoxide Dismutase 1 Gene Transcription in Human Cancer Cells: The Involvement of Peroxisome Proliferator-Activated Receptor alpha and Hypoxia-Inducible Factor-2 alpha Signaling

Docosahexaenoic acid (DHA; n-3, 22:6) is known to have anticancer activity, but its mechanisms of action remain to be further elucidated. We recently demonstrated that DHA down-regulates superoxide dismutase (SOD) 1 gene expression, thereby weakening cellular antioxidant forces and enhancing cytotoxicity in various human cancer cells. The objective of this study was to investigate the mechanism of the inhibitory effect of DHA on SOD-1 gene expression in human cancer cells. A reporter gene assay indicated that DHA suppresses SOD-1 gene transcription in a time- and concentration-dependent manner in human cancer cells. Pretreatment with vitamin E did not block the inhibitory effect of DHA, indicating that this suppression does not depend on lipid peroxidation. The suppressive effect of DHA on SOD-1 gene transcription could be mimicked by the peroxisome proliferator-activator receptor (PPAR) alpha ligand clofibrate but not the PPAR gamma ligand troglitazone, suggesting the involvement of PPAR alpha signaling. Deletion analysis of the key DNA binding elements in the SOD-1 gene promoter identified the distal hypoxia response element (HRE), but not the peroxisome proliferator response element or nuclear factor-kappa B element, as essential for the suppressive effects of DHA. Coimmunoprecipitation confirmed that PPAR alpha, but not PPAR gamma, forms a complex with hypoxia-inducible factor (HIF)-2 alpha in cancer cells. Chromatin immunoprecipitation analysis indicated that both DHA and clofibrate reduce HIF-2 alpha binding to the HRE. Thus, we have identified the distal HRE in the SOD-1 gene promoter that mediates the suppression on the transcription of this gene by DHA, and we have demonstrated the involvement of PPAR alpha and HIF-2 alpha signaling in this event.