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Mapping the proximity interaction network of the Rho-family GTPases reveals signalling pathways and regulatory mechanisms

Nat Cell Biol. 2020; 
Bagci H, Sriskandarajah N, Robert A, Boulais J, Elkholi IE, Tran V, Lin ZY, Thibault MP, Dubé N, Faubert D, Hipfner DR, , Gingras AC, Côté JF, 0,
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Proteins, Expression, Isolation and Analysis Cells were lysed in CHAPS buffer (30mM Tris-HCl, pH7.5, 150mM NaCl, 5mM MgCl2 and 1% CHAPS), and cleared lysates were incubated for 3h at room temperature with glutathione beads (L00206-10ml, GenScript) or with Flag-M2 agarose (A2220, Sigma-Aldrich). Get A Quote

摘要

Guanine nucleotide exchange factors (RhoGEFs) and GTPase-activating proteins (RhoGAPs) coordinate the activation state of the Rho family of GTPases for binding to effectors. Here, we exploited proximity-dependent biotinylation to systematically define the Rho family proximity interaction network from 28 baits to produce 9,939 high-confidence proximity interactions in two cell lines. Exploiting the nucleotide states of Rho GTPases, we revealed the landscape of interactions with RhoGEFs and RhoGAPs. We systematically defined effectors of Rho proteins to reveal candidates for classical and atypical Rho proteins. We used optogenetics to demonstrate that KIAA0355 (termed GARRE here) is a RAC1 interactor. A functiona... More

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