AbstractUbiquitin modification of the TGF-β pathway components is emerging as a key mechanism of TGF-β pathway regulation. To limit TGF-β responses, TGF-β signaling is regulated through a negative feedback loop whereby the E3 ligase SMURF2 targets the TGF-β receptor (TβR) complex for ubiquitin-mediated degradation. Counteracting this process, a number of deubiquitinating (DUBs) enzymes have recently been identified that deubiquitinate and stabilize the TβR. However the precise mechanism by which these DUBs act on TβR function remains poorly defined. Here, we demonstrate that apart from targeting the TβR complex directly, USP15 also deubiquitinates SMURF2 resulting in enhanced TβR stability and downstream pathway activation. Through proteomic analysis, we show that USP15 modulates the ubiquitination of Lys734, a residue required for SMURF2 catalytic activity. Our results show that SMURF2 is a critical target of USP15 in the TGF-β pathway and may also explain how USP15 and SMURF2 target multiple complementary protein complexes in other pathways.