Extraction of active RhoGTPases by RhoGDI regulates spatiotemporal patterning of RhoGTPases

Abstract

20The RhoGTPases are characterized as membrane-associated molecular switches cycling 21 between active, GTP-bound and inactive, GDP-bound states. However, 90-95% of RhoGTPases 22 are maintained in a soluble form by RhoGDI, which is generally viewed as a passive shuttle for 23 inactive RhoGTPases. Our current understanding of RhoGTPase:RhoGDI dynamics has been 24 limited by two experimental challenges: direct visualization of the RhoGTPases in vivo and 25 reconstitution of the cycle in vitro. We developed methods to directly image vertebrate 26 RhoGTPases in vivo or on lipid bilayers in vitro. Using these tools, we identified pools of active 27 and inactive RhoGTPase associated with the membrane, showed that RhoGDI can actively 28 extract both inactive and active RhoGTPases, and that the extraction of active RhoGTPase 29 contributes to their spatial regulation around wounds. In contrast to the textbook model of the 30 RhoGTPase cycle, these results indicate that RhoGDI actively contributes to spatiotemporal 31 patterning by removing active RhoGTPases from the plasma membrane. 32 33 34 35 36 37 38 39 40 41

Targeting Rho GTPase Signaling Networks in Cancer

“…For the classical Rho GTPases, this cycling between GTP-and GDP-bound states is tightly regulated by guanine nucleotide exchange factors (GEFs) and GTPase-activating proteins (GAPs) (Figure 2). Rho GTPases can also be regulated by guanine nucleotide dissociation inhibitor (GDI) binding, which sequesters Rho GTPases in the cytoplasm (Golding et al, 2019).…”

Animal Cell Cytokinesis: The Rho-Dependent Actomyosin-Anilloseptin Contractile Ring as a Membrane Microdomain Gathering, Compressing, and Sorting Machine

“…M-phase GAP (MP-GAP:ARHGAP11A in mammals, RGA-3/4 in C. elegans) also acts to globally suppress Rho and restrict cortical contractility to the equator (Zanin et al, 2013), although no ortholog exists in Drosophila. Finally, an additional level of regulation of the Rho GTPase cycle comes from guanine nucleotide dissociation inhibitors (GDIs), which can directly extract active Rho from the plasma membrane (Golding et al, 2019).…”

Structural basis for p50RhoGAP BCH domain–mediated regulation of Rho inactivation

“…The lipid-binding pocket that is required, but not sufficient, for RhoA binding highlights the multiple levels of regulation that exist for RhoA binding to its target protein, in addition to the well-known function of the lipidated tails of GTPases for membrane insertion. Notably, the BCH domain's preference for different nucleotide-bound or mutant RhoA is similar, but not identical, to that of the Rho GDP-dissociation inhibitor (RhoGDI), which acts to extract lipidated GTPases from the membrane (40,41). Indeed, while it remains unclear whether p50RhoGAP itself can extract RhoA from the membrane in a manner similar to that of other RhoGDIs, it is plausible that p50RhoGAP accomplishes the capture, retention, and inactivation mechanism proposed in the present study: RhoA binding to both the lipid-binding pocket and the other Rho-binding motif maintains a necessary local concentration of RhoA for its inactivation by the adjacent GAP domain following an appropriate signal.…”