Purpose Overexpression of cyclooxygenase 2 (COX-2) is thought to promote survival of transformed cells. Transforming growth factor beta (TGF-beta) exerts anti-proliferative effects on a broad range of epithelial cells. In the current study, we investigated whether TGF-beta can regulate COX-2 expression in A549 human lung adenocarcinoma cells, which are TGF-beta-responsive and overexpress COX-2. Materials and Methods Western blotting, Northern blotting, and mRNA stability assays were performed to demonstrate that COX-2 protein and mRNA expression were suppressed by TGF-beta. We also evaluated the effects of tristetraprolin (UP) on COX-2 mRNA using RNA interference. Results We demonstrated that COX-2 mRNA and protein expression were both significantly suppressed by TGF-beta. An actinomycin D chase experiment demonstrated that COX-2 mRNA was more rapidly degraded in the presence of TGF-beta, suggesting that TGF-beta-induced inhibition of COX-2 expression is achieved via decreased mRNA stability. We also found that TGF-beta rapidly and transiently induced the expression of UP, a well-known mRNA destabilizing factor, before suppression of COX-2 mRNA expression was observed. Using RNA interference, we confirmed that increased UP levels play a pivotal role in the destabilization of COX-2 mRNA by TGF-beta. Furthermore, we showed that Smad3 is essential to UP-dependent down-regulation of COX-2 expression in response to TGF-beta. Conclusion The results of this study show that TGF-beta down-regulated COX-2 expression via mRNA destabilization mediated by Smad3/TTP in A549 cells.