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The Rho GTPases are essential regulators of basic cellular processes, including cell migration, cell contraction and cell division. Most studies still involve just the three canonical members, RhoA, Rac1 and Cdc42, although the Rho GTPases comprise at least 20 members. The aim of this review is to highlight some of the recent advances in our knowledge regarding the less-studied Rho members, with the focus on RhoD and Rif. The phenotypic alterations to cell behaviour that are triggered by RhoD and Rif suggest that they have unique impacts on cytoskeletal dynamics that distinguish them from the well-studied members of the Rho GTPases. In addition, RhoD has a role in the regulation of intracellular transport of vesicles. Taken together, the available data indicate that RhoD and Rif have functions as master regulators in the integration of cytoskeletal reorganisation and membrane trafficking.

The Rho GTPases have mainly been studied in association with their roles in the regulation of actin filament organization. These studies have shown that the Rho GTPases are essential for basic cellular processes, such as cell migration, contraction, and division. In this paper, we report that RhoD has a role in the organization of actin dynamics that is distinct from the roles of the better-studied Rho members Cdc42, RhoA, and Rac1. We found that RhoD binds the actin nucleation-promoting factor WASp homologue associated with actin Golgi membranes and microtubules (WHAMM), as well as the related filamin A-binding protein FILIP1. Of these two RhoD-binding proteins, WHAMM was found to bind to the Arp2/3 complex, while FILIP1 bound filamin A. WHAMM was found to act downstream of RhoD in regulating cytoskeletal dynamics. In addition, cells treated with small interfering RNAs for RhoD and WHAMM showed increased cell attachment and decreased cell migration. These major effects on cytoskeletal dynamics indicate that RhoD and its effectors control vital cytoskeleton-driven cellular processes. In agreement with this notion, our data suggest that RhoD coordinates Arp2/3-dependent and FLNa-dependent mechanisms to control the actin filament system, cell adhesion, and cell migration.

The Cdc42-like GTPase Wnt responsive Cdc42 homolog 1 (Wrch1) has several atypical features; it has an N-terminal proline-rich extension that confers binding to SH3 domains, and it harbors an extremely high intrinsic nucleotide exchange activity, which overrides the normal GTPase activity. As a result, Wrch1 resides mainly in the active, GTP-loaded conformation under normal cellular conditions. We have previously shown that ectopic expression of Wrch1 in fibroblasts resulted in an altered cell morphology visible as a formation of filopodia, a loss of stress fibers, and a reduction in focal adhesions. Here, we show that Wrch1 binds to the nonreceptor tyrosine kinase Pyk2. The interaction required Wrch1 to be in a GTP conformation and also required an intact N-terminal proline-rich extension as well as an intact effector loop. Wrch1 requires Pyk2 in imposing the cytoskeletal effects, seen as the formation of filopodia, since treatment of cells with a Pyk2-specific small interfering RNA abrogated this response. Interestingly, we found that the presence and activity of Src were needed for the formation of a Wrch1-Pyk2 complex as well as for the Wrch1-induced formation of filopodia. We propose a model in which Pyk2 and Src function to coordinate the Wrch1-dependent effects on cytoskeletal dynamics.

Pombe Cdc15 homology (PCH) proteins have emerged in many species as important coordinators of signalling pathways that regulate actomyosin assembly and membrane dynamics. For example, the prototype PCH protein, Cdc15p of Schizosaccharomyces pombe, has a role in assembly of the contractile ring, which is needed to separate dividing cells. Recently, mammalian PCH proteins have been found to bind phospholipids and to participate in membrane deformation. These findings suggest that PCH proteins are crucial linkers of membrane dynamics and actin polymerization, for example, during the internalization of transmembrane receptors. Intriguingly, some members of the PCH protein family are mutated in neurodegenerative and inflammatory diseases, which has implications for the identification of cures for such disorders.

Transforming growth factor-beta (TGF-beta) is a potent regulator of cell growth and differentiation in many cell types. The Smad signaling pathway constitutes a main signal transduction route downstream of TGF-beta receptors. We studied TGF-beta-induced rearrangements of the actin filament system and found that TGF-beta 1 treatment of PC-3U human prostate carcinoma cells resulted in a rapid formation of lamellipodia. Interestingly, this response was shown to be independent of the Smad signaling pathway; instead, it required the activity of the Rho GTPases Cdc42 and RhoA, because ectopic expression of dominant negative mutant Cdc42 and RhoA abrogated the response. Long-term stimulation with TGF-beta 1 resulted in an assembly of stress fibers; this response required both signaling via Cdc42 and RhoA, and Smad proteins. A known downstream effector of Cdc42 is p38(MAPK); treatment of the cells with the p38(MAPK) inhibitor 4-(4-fluorophenyl)-2-(4-methylsulfinylphenyl)-5-(pyridyl)1H-imidazole (SB203580), as well as ectopic expression of a kinase-inactive p38(MAPK), abrogated the TGF-beta-induced actin reorganization. Moreover, treatment of cells with the inhibitors of the RhoA target-protein Rho-associated coiled-coil kinase (+)-R-trans-4-(aminoethyl)-N-(4-pyridyl) cyclohexanecarboxamide (Y-27632) and 1-5(-isoquinolinesulfonyl)homopiperazine (HA-1077), as well as ectopic expression of kinase-inactive Rho coiled-coil kinase-1, abrogated the TGF-beta 1-induced formation of stress fibers. Collectively, these data indicate that TGF-beta-induced membrane ruffles occur via Rho GTPase-dependent pathways, whereas long-term effects require cooperation between Smad and Rho GTPase signaling pathways.

BAR proteins comprise a heterogeneous group of multi-domain proteins with diverse biological functions. The common denominator is the Bin-Amphiphysin-Rvs (BAR) domain that not only confers targeting to lipid bilayers, but also provides scaffolding to mold lipid membranes into concave or convex surfaces. This function of BAR proteins is an important determinant in the dynamic reconstruction of membrane vesicles, as well as of the plasma membrane. Several BAR proteins function as linkers between cytoskeletal regulation and membrane dynamics. These links are provided by direct interactions between BAR proteins and actin-nucleation-promoting factors of the Wiskott-Aldrich syndrome protein family and the Diaphanous-related formins. The Rho GTPases are key factors for orchestration of this intricate interplay. This review describes how BAR proteins regulate the activity of Rho GTPases, as well as how Rho GTPases regulate the function of BAR proteins. This mutual collaboration is a central factor in the regulation of vital cellular processes, such as cell migration, cytokinesis, intracellular transport, endocytosis, and exocytosis. [ABSTRACT FROM PUBLISHER]

RhoD belongs to the Rho GTPases, a protein family responsible for the regulation and organization of the actin cytoskeleton, and, consequently, many cellular processes like cell migration, cell division and vesicle trafficking. Here, we demonstrate that the actin cytoskeleton is dynamically regulated by increased or decreased protein levels of RhoD. Ectopic expression of RhoD has previously been shown to give an intertwined weave of actin filaments. We show that this RhoD-dependent effect is detected in several cell types and results in a less dynamic actin filament system. In contrast, RhoD depletion leads to increased actin filament-containing structures, such as cortical actin, stress fibers and edge ruffles. Moreover, vital cellular functions such as cell migration and proliferation are defective when RhoD is silenced. Taken together, we present data suggesting that RhoD is an important component in the control of actin dynamics and directed cell migration. [ABSTRACT FROM AUTHOR]