Smad2 and Smad3 (Smad2/3) are key intracellular signal transducers for TGF-β signaling, and their transcriptional activities are controlled through reversible phosphorylation and nucleocytoplasmic shuttling. However, the precise mechanism underlying nuclear export of Smad2/3 remains elusive. Here we report the essential function of RanBP3 in selective nuclear export of Smad2/3 in the TGF-β pathway. RanBP3 directly recognizes dephosphorylated Smad2/3, which results from the activity of nuclear Smad phosphatases, and mediates nuclear export of Smad2/3 in a Ran-dependent manner. As a result, increased expression of RanBP3 inhibits TGF-β signaling in mammalian cells and Xenopus embryos. Conversely, depletion of RanBP3 expression or dominant-negative inhibition of RanBP3 enhances TGFβ-induced antiproliferative and transcriptional responses. In conclusion, our study supports a definitive role for RanBP3 in mediating Smad2/3 nuclear export and terminating TGF-β signaling. © 2009 Elsevier Inc. All rights reserved.