Identification of a mitotic Rac-GEF, Trio, that counteracts MgcRacGAP function during cytokinesis.
Bottom Line: Moreover, Trio depletion specifically rescues the cytokinesis failure induced by MgcRacGAP depletion.Of importance, we demonstrate that this rescue is mediated by the Trio-Rac1 pathway, using GEF-dead mutants of Trio and a specific inhibitor of Rac1 activation by Trio.Overall this work identifies for the first time a GEF controlling Rac1 activation in dividing cells that counteracts MgcRacGAP function in cytokinesis.
Affiliation: Signaling and Cytoskeleton Dynamics Group, University of Montpellier, 34293 Montpellier, France.Show MeSH
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Mentions: To directly test whether Trio could function as a GEF of Rac1 in dividing cells, the amount of activated Rac1 was monitored by pull-down assay in synchronized mitotic cells. Compared to control siRNA–treated cells, Trio depletion reduced by half the amount of activated Rac1 in mitotic cells (Figure 3A), showing that Trio activates Rac1 in mitosis. Strikingly, Trio depletion (Figure 3B) led to defects in F-actin cytoskeleton remodeling in both interphase (unpublished data) and dividing cells (Figure 3C). More specifically, in anaphase cells, the F-actin staining at the cortex was significantly reduced in Trio-depleted cells compared with control cells (Figure 3C, MP). This decrease in F-actin staining was particularly obvious at the bottom plane (Figure 3, C–E, BP and inset), where cells are in contact with the substrate. Of interest, Trio depletion phenocopied the depletion of Rac1, consistent with a role for the Trio-Rac1 pathway in controlling F-actin remodeling in dividing cells (Figure 3, B–E). Trio depletion also phenocopied the depletion of actin-related protein 3 (Arp3; Figure 3, B–E), a component of the Arp2/3 complex whose depletion disrupts the Arp2/3 complex (Steffen et al., 2006). Arp2/3 is known to nucleate branched actin filaments downstream of Rac1 (Pollard, 2007). Taken together, these results suggest that Trio could control F-actin remodeling during cell division by controlling the Rac1-Arp2/3 pathway.
Affiliation: Signaling and Cytoskeleton Dynamics Group, University of Montpellier, 34293 Montpellier, France.