Coyaud E, Mis M, Laurent EM, Dunham WH, Couzens AL, Robitaille M, Gingras AC, Angers S, Raught B

The identification of ubiquitin E3 ligase substrates has been challenging, due in part to low-affinity, transient interactions, the rapid degradation of targets and the inability to identify proteins from poorly soluble cellular compartments. SCF(β-TrCP1) and SCF(β-TrCP2) are well-studied ubiquitin E3 ligases that target substrates for proteasomal degradation, and play important ...

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roles in Wnt, Hippo, and NFκB signaling. Combining 26S proteasome inhibitor (MG132) treatment with proximity-dependent biotin labeling (BioID) and semiquantitative mass spectrometry, here we identify SCF(β-TrCP1/2) interacting partners. Based on their enrichment in the presence of MG132, our data identify over 50 new putative SCF(β-TrCP1/2) substrates. We validate 12 of these new substrates and reveal previously unsuspected roles for β-TrCP in the maintenance of nuclear membrane integrity, processing (P)-body turnover and translational control. Together, our data suggest that β-TrCP is an important hub in the cellular stress response. The technique presented here represents a complementary approach to more standard IP-MS methods and should be broadly applicable for the identification of substrates for many ubiquitin E3 ligases.

Date: Jul. 01, 2015

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