Cip4 integrates membrane trafficking and actin dynamics through WASP and WAVE. First, Cip4 promotes membrane invaginations and triggers the vesicle scission by recruiting Dynamin to the neck of nascent vesicles. Second, Cip4 recruits WASP and WAVE proteins to induce actin polymerization, supporting vesicle scission and providing the force for vesicle movement.

During epithelial development cells become polarized along their apical-basal axis and some epithelia also exhibit polarity in the plane of the tissue. Mutations in the gene encoding a Drosophila Pak family serine/threonine kinase, dPak, disrupt the follicular epithelium that covers developing egg chambers during oogenesis. The follicular epithelium normally exhibits planar polarized organization of basal F-actin bundles such that they lie perpendicular to the anterior-posterior axis of the egg chamber, and requires contact with the basement membrane for apical-basal polarization. During oogenesis, dPak becomes localized to the basal end of follicle cells and is required for polarized organization of the basal actin cytoskeleton and for epithelial integrity and apical-basal polarity. The receptor protein tyrosine phosphatase Dlar and integrins, all receptors for extracellular matrix proteins, are required for polarization of the basal F-actin bundles, and for correct dPak localization in follicle cells. dpak mutant follicle cells show increased beta(Heavy)-spectrin levels, and we speculate that dPak regulation of beta(Heavy)-spectrin, a known participant in the maintenance of membrane domains, is required for correct apical-basal polarization of the membrane. We propose that dPak mediates communication between the basement membrane and intracellular proteins required for polarization of the basal F-actin and for apical-basal polarity.

We have characterized Drosophila melanogaster ACK (DACK), one of two members of the ACK family of nonreceptor tyrosine kinases in Drosophila. The ACKs are likely effectors for the small GTPase Cdc42, but signaling by these proteins remains poorly defined. ACK family tyrosine kinase activity functions downstream of Drosophila Cdc42 during dorsal closure of the embryo, as overexpression of DACK can rescue the dorsal closure defects caused by dominant-negative Dcdc42. Similar to known participants in dorsal closure, DACK is enriched in the leading edge cells of the advancing epidermis, but it does not signal through activation of the Jun amino-terminal kinase cascade operating in these cells. Transcription of DACK is responsive to changes in Dcdc42 signaling specifically at the leading edge and in the amnioserosa, two tissues involved in dorsal closure. Unlike other members of the ACK family, DACK does not contain a conserved Cdc42-binding motif, and transcriptional regulation may be one route by which Dcdc42 can affect DACK function. Expression of wild-type and kinase-dead DACK transgenes in embryos, and in the developing wing and eye, reveals that ACK family tyrosine kinase activity is involved in a range of developmental events similar to that of Dcdc42.Cdc42 is a member of the Rho family of Ras-related small GTPases originally identified through a mutation in Saccharomyces cerevisiae that affects formation of the bud site. The Cdc42 protein is required for the assembly of a ring of F-actin filaments in the neck of the bud (1). Subsequent work in mammalian fibroblasts demonstrated that Cdc42 drives the formation of F-actin-rich filopodia (40, 50), and numerous later studies have confirmed that Cdc42 regulates the actin cytoskeleton and, as a consequence, cell shape (65). Cdc42 participates in a diverse range of cellular processes including membrane trafficking, transcription, cell growth, and Ras-mediated transformation (65). The various effects of Cdc42 are presumed to be mediated through the interaction of the activated, GTP-bound form of the protein with downstream effectors.Given the important events controlled by Cdc42, intensive efforts have been made to elucidate the signaling pathways activated by this GTPase. This work has largely focused on identifying proteins that interact with GTP-bound Cdc42. Two such proteins are ACK-1 and ACK-2, closely related mammalian nonreceptor tyrosine kinases that bind GTP-bound Cdc42 and not its inactive GDP-bound form (44, 67). ACK-1 and ACK-2 cannot bind either version of the closely related Rho family GTPases Rac1 and RhoA, and these kinases represent likely effectors in Cdc42-specific signaling.To date, much of what is known about Rho family signaling has come from biochemical and cell biological work, but it is now being studied with genetic approaches in a number of model organisms, including Drosophila melanogaster. The Drosophila homolog of Cdc42, Dcdc42, has been studied by using dominantly acting mutant transgenes and loss-of-function mutations. This work has in...

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The Rho family small GTPases Rho, Rac, and Cdc42 regulate cell shape and motility through the actin cytoskeleton. These proteins cycle between a GTP-bound "on" state and a GDP-bound "off" state and are negatively regulated by GTPase-activating proteins (GAPs), which accelerate the small GTPase's intrinsic hydrolysis of bound GTP to GDP. Drosophila RhoGAP68F is similar to the mammalian protein p50RhoGAP/Cdc42GAP, which exhibits strong GAP activity toward Cdc42. We find that, despite the strong similarities between RhoGAP68F and p50RhoGAP/Cdc42GAP, RhoGAP68F is most effective as a GAP for RhoA. These in vitro data are supported by the in vivo analysis of mutants in RhoGAP68F. We demonstrate through the characterization of two alleles of the RhoGAP68F gene that RhoGAP68F participates in gastrulation of the embryo, a morphogenetic event driven by cell constriction that involves RhoA signaling. We propose that RhoGAP68F functions as a regulator of RhoA signaling during gastrulation.