CUL3-KBTBD6/KBTBD7 Ubiquitin Ligase Cooperates with GABARAP Proteins to Spatially Restrict TIAM1-RAC1 Signaling

Abstract

The small Rho GTPase RAC1 is an essential regulator of cellular signaling that controls actin rearrangements and cell motility. Here, we identify a novel CUL3 RING ubiquitin ligase complex, containing the substrate adaptors KBTBD6 and KBTBD7, that mediates ubiquitylation and proteasomal degradation of TIAM1, a RAC1-specific GEF. Increasing the abundance of TIAM1 by depletion of KBTBD6 and/or KBTBD7 leads to elevated RAC1 activity, changes in actin morphology, loss of focal adhesions, reduced proliferation, and enhanced invasion. KBTBD6 and KBTBD7 employ ATG8 family-interacting motifs to bind preferentially to GABARAP proteins. Surprisingly, ubiquitylation and degradation of TIAM1 by CUL3KBTBD6/KBTBD7 depends on its binding to GABARAP proteins. Our study reveals that recruitment of CUL3KBTBD6/KBTBD7 to GABARAP-containing vesicles regulates the abundance of membrane-associated TIAM1 and subsequently spatially restricted RAC1 signaling. Besides their role in autophagy and trafficking, we uncovered a previously unknown function of GABARAP proteins as membrane-localized signaling scaffolds.