Perfluorooctane sulfonic acid (PFOS) is widely distributed in the environment including in the tissues of wildlife and humans, however, its mechanism of action remains unclear. Here, the Affymetrix rat genome U34A genechip was used to identify alterations in gene expression due to PFOS exposure. Rat hepatoma cells were treated with PFOS at 2-50mg/L (4-100μM) for 96h. Sprague-Dawley rats were orally dosed with PFOS at 5mg/kg/day for 3 days or 3 weeks. Genes that were significantly (P <0.0025) induced were primarily genes for fatty acid metabolizing enzymes, cytochrome P450s, or genes involved in hormone regulation. Consistent expression profiles were obtained for replicate exposures, for short-term and long-term in vivo exposures, and for acute and chronic exposures. One major pathway affected by PFOS was peroxisomal fatty acid β-oxidation, which could be explained by the structural similarity between PFOS and endogenous fatty acids.