Within the lymphocyte lineage, GATA-3 is a major transcription factor implicated in the regulation of Th1/Th2 differentiation by promoting the expression of the Th2 cytokines, such as IL-4, IL-5, IL-10 and IL-13. Although the role of GATA-3 in the development of the Th2 lineage has been extensively described in the literature, the molecular mechanisms underlying its activity remain to be clarified. Here, we investigated whether GATA-3 might be regulated by reversible acetylation. In vivo, GATA-3 associates with class I and III HDACs. Biochemical studies unraveled the specific association of GATA-3 with the class III member SIRT1. Association with SIRT1 leads to the inhibition of GATA-3-induced IL-5 transcription. Using siRNA, we further show that SIRT1 promotes destabilization of GATA-3. Interestingly, nicotinamide, a specific inhibitor of SIRTs had no effect on the ability of SIRT1 to destabilize GATA-3 and to repress its transcriptional activity. In addition, a catalytic-defective mutant of SIRT1 (H363Y) shows similar effects to wild-type SIRT1, demonstrating that the deacetylase activity of SIRT1 is not required for its regulation of GATA-3. For the first time, our study indicates that SIRT1 is functionally linked to GATA-3. Moreover, our results also suggest that some important SIRT1 functions may not require its deacetylase activity.