To investigate the regulatory function of chaperone interacting protein CHIP in TGF-β signal pathway, a tetracycline induced CHIP expression stable cell line (Mv1Lu-Tet off-CHIP) was generated. In this cell line model, it was observed that overexpression of CHIP dramatically reduced the endougenous Smad2/3 protein level. Luciferase reporter analysis showed that CHIP inhibited the Smads induced transcription activity. Furthermore, Western blot results indicated that CHIP could downregulate gene expression of JunB, a quick response gene induced by TGF-β. These results provided new evidences that CHIP might be a novel signal protein negatively regulating TGF-β signal pathway.