Inducible recruitment of Cdc42 or WASP to a cell-surface receptor triggersactin polymerization and filopodium formation

Background: Cdc42, a GTP-binding protein of the Rho family, controls actin cytosketetal organization and helps to generate actin-based protruding stru ctures, such as filopodia. In vitro, Cdc42 regulates actin polymerization b y facilitating the creation of free barbed ends - the more rapidly growing ends of actin filaments - and subsequent elongation at these ends. The Wisk ott-Aldrich syndrome protein, WASP, which has a pleckstrin-homology domain and a Cdc42/Rac-binding motif, has been implicated in cell signaling and cy toskeleton reorganization. We have investigated the consequences of local r ecruitment of activated Cdc42 or WASP to the plasma membrane, Results: We used an activated Cdc42 protein that could be recruited to an e ngineered membrane receptor by adding rapamycin as a bridge, and added anti body-coupled beads to aggregate these receptors. Inducible recruitment of C dc42 to clusters of receptors stimulated actin polymerization, resulting in the formation of membrane protrusions. Cdc42-induced protrusions were enri ched in the vasodilator-stimulated phosphoprotein VASP and the focal-adhesi on-associated proteins zyxin and ezrin. The Cdc42 effector WASP could also induce the formation of protrusions, albeit of different morphology. Conclusions: This is the first demonstration that the local recruitment of activated Cdc42 or its downstream effector, WASP, to a membrane receptor in whole cells is sufficient to trigger actin polymerization that results in the formation of membrane protrusions. Our data suggest that Cdc42-induced actin-based protrusions result from the local and serial recruitment of cyt oskeletal proteins including zyxin, VASP, and ezrin.