Dietary polyunsaturated fatty acids (PUFAs) are a source of energy and structural components for cells. PUFAs also have dramatic effects on gene expression by regulating the activity or abundance of four families of transcription factor, including peroxisome proliferator activated receptor (PPAR) (α, β and γ), liver X receptors (LXRs) (α and β), hepatic nuclear factor-4 (HNF-4)α and sterol regulatory element binding proteins (SREBPs) 1 and 2. These transcription factors play a major role in hepatic carbohydrate, fatty acid, triglyceride, cholesterol and bile acid metabolism. Non-esterified fatty acids or fatty acid metabolites bind to and regulate the activity of PPARs, LXRs and HNF-4. In contrast, PUFAs regulate the nuclear abundance of SREBPs by controlling the proteolytic processing of SREBP precursors, or regulating transcription of the SREBP-1c gene or turnover of mRNASREBP-1c. The n3 and n6 PUFAs are feed-forward activators of PPARs, while these same fatty acids are feedback inhibitors of LXRs and SREBPs. Saturated fatty acyl coenzyme A thioesters activate HNF-4α, while coenzyme A thioesters of PUFAs antagonize HNF-4α action. Understanding how fatty acids regulate the activity and abundance of these and other transcription factors will likely provide insight into the development of novel therapeutic strategies for better management of whole body lipid and cholesterol metabolism.