Function And Mechanism Of OTUD1 In Cancer Metastasis

Metastasis is the main cause of death in cancer patients but the underlying mechanisms are largely unclear.Here,we built up a loss-of-function screen system by using mice as filter to read out metastasis.By screening shRNA library of deubiquitinating enzymes,we identified OTU domain-containing protein 1(OTUD1)as a novel metastasis-repressing factor.This approach can be easily expanded to other types of cancers.We envision that by elucidating this multistep metastatic process,vulnerabilities that are being exposed could be exploited to prevent metastasis.OTUD1 silenced cancer cells show mesenchymal and stem-cell-like characteristics.Further investigation revealed that OTUD1 directly deubiquitylates the TGF-? pathway inhibitor SMAD7 and prevents its degradation.OTUD1 not only stabilizes SMAD7 by removing Lys-48 poly-ubiquitination,but also removes Lys33-linked poly-ubiquination of SMAD7 and thereby enables SMAD7 to recruit the E3-ubiquitin ligase SMURF2.The Lys33-linked ubiquitin modification on Lys residue 220 in SMAD7 is in close proximity of the PY-motif in SMAD7 that mediates the interaction with E3 ubiquitin ligase SMURF2.Structural modelling of ubiquitin bound to SMAD7 at position 220 reveals that PY interaction motif is shielded,and this is further confirmed by biochemical and cellular experiments.Lys33-linked ubiquitination is an atypical linkage which is less known,and our study provides novel conceptual insight into how this uncommon reversible modification controls a specific protein-protein interaction that determines TGF-? type ? receptor stability at the cell surface.Using intracardial xenograft metastasis models and clinical database and patient sample analysis we confirmed the metastasis repressing effect of OTUD1.Low expression and genetic loss of OTUD1 was found to correlate with poor prognosis in breast and other types of cancers.Moreover,consistent with our biochemical and cell biological studies,we found that the protein expression of OTUD1 correlated with its target SMAD7 in clinical samples of breast cancer patients.Importantly,OTUD1 enriches 70 core targets of BRCA1 in the analysis of breast cancer patients.Similar with BRCA1 and BRCA2,expression of OTUD1 was inhibited by RAS oncogene in p53 null cells.These findings will provide new insights into the molecular mechanisms of TGF-?pathway.Thus,OTUD1 represses breast cancer metastasis by mitigating TGF-?-induced pro-oncogenic responses via deubiquitylating SMAD7.