Fatty acid synthase (FAS) is a key enzyme of hepatic lipogenesis responsible for the synthesis of long chain saturated fatty acids. This enzyme is mainly regulated at the transcriptional level by nutrients and hormones. In particular, glucose, insulin and T3 increase FAS activity whereas glucagon, saturated and polyunsaturated fatty acids decrease it. In the present study, we show that, in liver, T3 and insulin were able to activate FAS enzymatic activity, mRNA expression and gene transcription. We have localized the T3 response element (TRE) that mediates the T3 genomic effect, on the FAS promoter between -741 and -696 bp that mediates the T3 genomic effect. We show that both T3 and insulin regulate FAS transcription via this sequence. The TRE binds a TR/RXR heterodimer, even in the absence of hormone and this binding is increased in response to T3 and/or insulin treatment. The use of H7, a serine/threonine kinase inhibitor, reveals that a phosphorylation mechanism is implicated in the transcriptional regulation of FAS in response to both hormones. Specifically, we show that T3 is able to modulate FAS transcription via a non-genomic action targeting the TRE through the activation of a PI3-kinase-Erk1/2-MAPK dependent pathway. Insulin also targets the TRE sequence, probably via the activation of two parallel pathways: Ras/Erk1/2 MAPK and PI3-kinase/Akt. Finally, our data suggest that the non-genomic actions of T3 and insulin are probably common to several TREs, as we observed similar effects on a classical DR4 consensus sequence.