The Rho Family GTPase Rif Induces Filopodia through mDia2

Mammalian Cell Biology Laboratory, Department of Biochemistry, School of Medical Sciences, University of Bristol, Bristol BS8 1TD, United Kingdom.
Current Biology (Impact Factor: 9.57). 02/2005; 15(2):129-33. DOI: 10.1016/j.cub.2005.01.011
Source: PubMed


Eukaryotic cells produce a variety of specialized actin-rich surface protrusions. These include filopodia-thin, highly dynamic projections that help cells to sense their external environment. Filopodia consist of parallel filaments of actin, bundled by actin crosslinking proteins. The filaments are oriented with their rapidly growing "barbed" ends at the protruding tip and their slowly growing "pointed" ends at the base. Extension occurs by polymerization at the tip and is controlled by regulation of filament capping. The Rho GTPase Cdc42 is a key mediator of filopodia formation, which it regulates through binding CRIB domain-containing effectors. Cdc42 binds and activates the WASP proteins, which in turn activate the actin-nucleating complex Arp2/3. It also binds and activates IRSp53, which recruits the Ena/WASP family protein Mena to the filopodial tip and protects elongating actin filaments from capping. Previously, we identified another Rho family GTPase, Rif, as a potent stimulator of filopodial protrusion through a mechanism that does not require Cdc42. Here we characterize the differences between filopodia induced by these two small GTPases and show that the Rif effector in this pathway is the Diaphanous-related formin mDia2. Thus, Rif and Cdc42 represent two distinct routes to the induction of filopodia-producing structures with both shared and unique properties.

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Available from: Harry Mellor, Oct 06, 2015
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    • "Interestingly, inside filopodia, GFP-Simiate localizes mainly to the head (Figure 4A) as does endogenous Simiate in filopodia from neuronal growth cones (cp. Figure 5A), supporting the idea that Simiate is indeed involved in the exploration behavior of filopodia by regulating Actin polymerization in the filopodia tip (cp. Berg and Cheney, 2002; Svitkina et al., 2003; Lebrand et al., 2004; Yamagishi et al., 2004; Millard et al., 2005; Pellegrin and Mellor, 2005; Schirenbeck et al., 2006; Dent et al., 2007; Goh et al., 2011). "
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    ABSTRACT: The Actin cytoskeleton constitutes the functional base for a multitude of cellular processes extending from motility and migration to cell mechanics and morphogenesis. The latter is particularly important to neuronal cells since the accurate functioning of the brain crucially depends on the correct arborization of neurons, a process that requires the formation of several dozens to hundreds of dendritic branches. Recently, a model was proposed where different transcription factors are detailed to distinct facets and phases of dendritogenesis and exert their function by acting on the Actin cytoskeleton, however, the proteins involved as well as the underlying molecular mechanisms are largely unknown. Here, we demonstrate that Simiate, a protein previously indicated to activate transcription, directly associates with both, G- and F-Actin and in doing so, affects Actin polymerization and Actin turnover in living cells. Imaging studies illustrate that Simiate particularly influences filopodia dynamics and specifically increases the branching of proximal, but not distal dendrites of developing neurons. The data suggests that Simiate functions as a direct molecular link between transcription regulation on one side, and dendritogenesis on the other, wherein Simiate serves to coordinate the development of proximal and distal dendrites by acting on the Actin cytoskeleton of filopodia and on transcription regulation, hence supporting the novel model.
    Frontiers in Cellular Neuroscience 04/2014; 8(1):99. DOI:10.3389/fncel.2014.00099 · 4.29 Impact Factor
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    • "Mouse diaphanous 2, mDia2, is a closely related formin protein. mDia2 was first identified as a protein which interacted with small GTPases [6] and is involved in filopodia formation in mammalian cells [7], [8]. In mouse oocytes with no recognizable centrosomes [9], [10], microtubule organizing centers (MTOCs) serve the function of microtubule formation. "
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    ABSTRACT: The mouse diaphanous 2 (mDia2) protein belongs to the formin family and has been shown to nucleate actin filaments and stabilize microtubules, thus indicating a role in cytoskeleton organization. Our previous study, which showed that mDia2 specifically localizes to spindle poles of metaphase I mouse oocytes and NIH3T3 cells, provided the first evidence of its spindle pole-associated cellular function. In the present study, we aim to determine whether spindle pole proteins, such as mDia2 and pericentrin, can be used to monitor the status of spindle poles in cryopreserved mouse oocytes. We show herein that mDia2 exhibits an overlapping distribution with pericentrin, which is a crucial component of centrosomes and microtubule organizing centers (MTOCs). In vitrified-warmed oocytes, the overlapping distribution of mDia2 and pericentrin was immediately detected after thawing, thereby suggesting that mDia2 maintains a tight association with the spindle pole machinery. Interestingly, we observed that microtubules extend from mDia2 clusters in cytoplasmic MTOCs after thawing. This result suggests that mDia2 is a major MTOC component that is closely associated with pericentrin and that it plays a role in microtubule growth from MTOCs. Collectively, our results provide evidence that mDia2 is a novel marker of spindle pole dynamics before and after cryopreservation.
    PLoS ONE 09/2013; 8(9):e75729. DOI:10.1371/journal.pone.0075729 · 3.23 Impact Factor
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    • "They are characteristically long finger-like projections within which the actin filaments are tightly bundled, and are thought to function as gradient sensors to orient the migrating cell [13]–[16] and to provide traction force [17], [18] through adhesion proteins [19]. Moreover, a specific set of proteins give them a unique character; Ena/VASP proteins preventing capping of the polymerizing barbed ends [20]–[22], myosin X transporting cargo like Mena/VASP [23] to the filopodial tips [24], IRSp53 deforming the membrane through its inverse BAR (I-BAR) domain [25], fascin cross-linking actin filaments [26], [27] and formins like mDia2 promoting polymerization of long unbranched actin filaments [28]–[30]. Still, the molecular mechanisms and signaling pathways involved in filopodial induction are not fully understood [30]. "
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    ABSTRACT: All modes of cell migration require rapid rearrangements of cell shape, allowing the cell to navigate within narrow spaces in an extracellular matrix. Thus, a highly flexible membrane and a dynamic cytoskeleton are crucial for rapid cell migration. Cytoskeleton dynamics and tension also play instrumental roles in the formation of different specialized cell membrane protrusions, viz. lamellipodia, filopodia, and membrane blebs. The flux of water through membrane-anchored water channels, known as aquaporins (AQPs) has recently been implicated in the regulation of cell motility, and here we provide novel evidence for the role of AQP9 in the development of various forms of membrane protrusion. Using multiple imaging techniques and cellular models we show that: (i) AQP9 induced and accumulated in filopodia, (ii) AQP9-associated filopodial extensions preceded actin polymerization, which was in turn crucial for their stability and dynamics, and (iii) minute, local reductions in osmolarity immediately initiated small dynamic bleb-like protrusions, the size of which correlated with the reduction in osmotic pressure. Based on this, we present a model for AQP9-induced membrane protrusion, where the interplay of water fluxes through AQP9 and actin dynamics regulate the cellular protrusive and motile activity of cells.
    PLoS ONE 04/2013; 8(4):e59901. DOI:10.1371/journal.pone.0059901 · 3.23 Impact Factor
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