Renal cell carcinoma (RCC) is the most prevalent cancer of the kidney. In human RCC cells, we recently showed that insulin-like growth factor I (IGF-I) has growth-promoting effects regulated by IGF-binding protein 3 (IGFBP-3). In this study, the analysis was expanded to include the interaction between the IGF and transforming growth factor-beta (TGF-beta) systems in the human RCC cells Caki-2 (from a primary tumor) and SK-RC-52 (from a metastasis). Functional effects such as cell proliferation, TGF-beta receptor (TbetaR) signaling, and IGFBP-3 levels were monitored after stimulation with various concentrations of IGF-I, TGF-beta, and IGFBP-3. In addition, human RCC tissues as well as experimental human RCC tumors were analyzed for cellular expression of phosphorylated Smad2 by immunohistochemistry. TGF-beta regulated the endogenous IGFBP-3 levels in these RCC cells as neutralizing anti-TGF-beta(1-3) antibodies strongly reduced the basal IGFBP-3 level. In addition, IGF-I increased the IGFBP-3 levels five- to eightfold with TGF-beta acting in synergy to enhance the IGFBP-3 levels 12- to 17-fold. Neutralizing TGF-beta(1-3) activity circumvented the growth inhibitory effects of IGFBP-3 seen in SK-RC-52, whereas it inhibited the growth-promoting effects of IGFBP-3 in Caki-2. Moreover, IGF-I interacted directly with TGF-beta activation of the TbetaR complex by enhancing phosphorylation and nuclear translocation of Smad2. This study demonstrates a direct interaction of the IGF and TGF-beta systems in human renal carcinoma cells. The observations that IGF-I enhances the TGF-beta signaling and that TGF-beta promotes IGFBP-3 production and thus influence the biological activity of IGF may be of importance for future therapeutic options.