Transforming growth factor-beta 1 (TGF-beta1) and activin A (ActA) induce similar intracellular signaling mediated by the mothers against decapentaplegic homolog (SMAD) proteins. TGF-beta1 is a potent antimitogenic factor for thyroid follicular cells, while the role of ActA is not clear. In our study, the proliferation of TPC-1, the papillary thyroid carcinoma cell line, was reduced by both recombinant ActA and TGF-beta1. Due to the concomitant expression of TGF-beta1 and ActA in thyroid tumors, we investigated the effects of either TGF-beta1 or ActA gene silencing by RNA interference in TPC-1 cells in order to distinguish the specific participation of each in proliferation and intracellular signaling. An increased proliferation and reduced SMAD2, SMAD3, and SMAD4 mRNA expression were observed in both TGF-beta1 and ActA knockdown cells. Recombinant TGF-beta1 and ActA increased the expression of inhibitory SMAD7, whereas they reduced c-MYC. Accordingly, we detected a reduction in SMAD7 expression in knockdown cells while, unexpectedly, c-MYC was reduced. Our data indicate that both TGF-beta1 and ActA generate SMADs signaling with each regulating the expression of their target genes, SMAD7 and c-MYC. Furthermore, TGF-beta1 and ActA have an antiproliferative effect on thyroid papillary carcinoma cell, exerting an important role in the control of thyroid tumorigenesis.