The Rho Family GTPase Rif Induces Filopodia through mDia2

Pellegrin, Stéphanie; Mellor, Harry

doi:10.1016/j.cub.2005.01.011

Cited by 263 publications

(283 citation statements)

References 23 publications

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“…We therefore tested a variant of Isof7 that is artificially activated by deletion of its autoinhibitory DAD domain (GFP.Isof7∆DAD), but is unlikely to be expressed endogenously. Consistent with previous reports for DIAPH3 as well as for analogous constructs of the DIAPH3 mouse ortholog mDia2 [32][33][34], expression of Isof7∆DAD resulted in the formation of multiple filopodia but not blebs independently of whether or not endogenous DIAPH3 was depleted. In sharp contrast, expression of isoform 1 fused to GFP (GFP.Isof1, encompassing residues 264-1112) in DIAPH3 knockdown cells induced efficient PM blebbing that was sensitive to inhibition of ROCK ( Figure 4A and B, data not shown).…”

Section: Pm Blebbing Involves a Specific Diaph3 Isoformsupporting

confidence: 91%

“…Of note, introduction of a H181D mutation known to disrupt the interaction between the mDia2-GBD and Cdc42 [34,40], did not affect filopodia induction and failed to promote bleb formation by Isof7∆DAD (see Isof7∆DAD H181D, Figure 7B). Together, these results reveal the GBD as a key determinant for subcellular targeting and protrusion specificity of activated human DIAPH3 and further define isoform 1 as a non-GTPase-regulated protein that is specifically involved in PM blebbing.…”

Section: Aph3 Variantsmentioning

confidence: 96%

“…Dimerization and actin nucleation-deficient mutants were generated in analogy to mouse mDia2 [35] by site-directed mutagenesis using the DIAPH3 expression constructs for Isof7, Isof7∆DAD, Isof1 and Isof1 A274V as template, respectively. The Cdc42-binding mutant of DIAPH3 isoform 7 ∆ DAD (Isof7 ∆ DAD H181D) was generated in analogy to mouse mDia2 [34] by site-directed mutagenesis of DIAPH3 isoform 7 ∆ DAD construct. N-and C-terminal DIAPH3 deletion constructs were generated by PCR amplification and were cloned into pEGFP-C1.…”

Section: Expression Constructs Reagents and Antibodiesmentioning

confidence: 99%

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Differing and isoform-specific roles for the formin DIAPH3 in plasma membrane blebbing and filopodia formation

et al. 2011

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Plasma membrane (PM) blebs are dynamic actin-rich cell protrusions that occur, e.g., during cytokinesis, amoeboid cell motility and cell attachment. Using a targeted siRNA screen against 21 actin nucleation factors, we identify a novel and essential role of the human diaphanous formin DIAPH3 in PM blebbing during cell adhesion. Suppression of DIAPH3 inhibited blebbing to promote rapid cell spreading involving β1-integrin. Multiple isoforms of DIAPH3 were detected on the mRNA and protein level of which isoforms 3 and 7 were the largest and most abundant isoforms that however did not induce formation of actin-rich protrusions. Rather, PM blebbing specifically involved the low abundance isoform 1 of DIAPH3 and activation of isoform 7 by deletion of the diaphanous-autoregulatory domain caused the formation of filopodia. Dimerization and actin assembly activity were essential for induction of specific cell protrusions by DIAPH3 isoforms 1 and 7. Our data suggest that the N-terminal region comprising the GTPasebinding domain determined the subcellular localization of the formin as well as its protrusion activity between blebs and filopodia. We propose that isoform-selective actin assembly by DIAPH3 exerts specific and differentially regulated functions during cell adhesion and motility.

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Section: Pm Blebbing Involves a Specific Diaph3 Isoformsupporting

confidence: 91%

Section: Aph3 Variantsmentioning

confidence: 96%

Section: Expression Constructs Reagents and Antibodiesmentioning

confidence: 99%

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Differing and isoform-specific roles for the formin DIAPH3 in plasma membrane blebbing and filopodia formation

et al. 2011

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“…Over 30 formins have been described to date, with more than 15 family members found in vertebrates (1). Formin activity is required in vivo for a diverse array of cellular functions, such as stress fiber formation, endosome motility, cell motility, cytokinetic ring formation, cell-cell junction assembly, filopodia formation, induction of cell polarity, and activation of the MAL/serum response factor signaling pathway (2)(3)(4)(5)(6)(7)(8)(9)(10)(11)(12)(13). It is thought that at the core of all of these activities is the ability of formins to regulate actin cytoskeletal dynamics.…”

mentioning

confidence: 99%

Interaction of the N- and C-terminal Autoregulatory Domains of FRL2 Does Not Inhibit FRL2 Activity

Vaillant

Copeland

Davis

et al. 2008

Journal of Biological Chemistry

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Formin homology proteins are a highly conserved family of cytoskeletal remodeling proteins best known for their ability to induce the formation of long unbranched actin filaments. They accomplish this by nucleating the de novo polymerization of F-actin and also by acting as F-actin barbed end "leaky cappers" that allow filament elongation while antagonizing the function of capping proteins. More recently, it has been reported that the FH2 domains of FRL1 and mDia2 and the plant formin AFH1 are able to bind and bundle actin filaments via distinct mechanisms. We find that like FRL1, FRL2 and FRL3 are also able to bind and bundle actin filaments. In the case of FRL3, this activity is dependent upon a proximal DAD/WH2-like domain that is found C-terminal to the FH2 domain. In addition, we show that, like other Diaphanousrelated formins, FRL3 activity is subject to autoregulation mediated by the interaction between its N-terminal DID and C-terminal DAD. In contrast, the DID and DAD of FRL2 also interact in vivo and in vitro but without inhibiting FRL2 activity. These data suggest that current models describing DID/ DAD autoregulation via steric hindrance of FH2 activity must be revised. Finally, unlike other formins, we find that the FH2 and N-terminal dimerization domains of FRL2 and FRL3 are able to form hetero-oligomers.Formin homology proteins (formins) are a highly conserved family of cytoskeletal regulatory proteins. Over 30 formins have been described to date, with more than 15 family members found in vertebrates (1). Formin activity is required in vivo for a diverse array of cellular functions, such as stress fiber formation, endosome motility, cell motility, cytokinetic ring formation, cell-cell junction assembly, filopodia formation, induction of cell polarity, and activation of the MAL/serum response factor signaling pathway (2-13). It is thought that at the core of all of these activities is the ability of formins to regulate actin cytoskeletal dynamics. This is achieved through the activity of two conserved formin homology domains, FH1 and FH2. FH1 consists of proline-rich repeats of varying sizes that can serve as ligands for Src homology 3 and WW domains as well as the small actin-binding protein profilin (14,15). FH2 domains form a dimer that induces the polymerization of long, unbranched filaments. It does this by nucleating de novo actin polymerization and by associating with the F-actin barbed end as a "leaky capper," allowing filament elongation while antagonizing the function of capping proteins (16). However, not all formins nucleate or elongate F-actin with equal potency. For example, mDia1 is an extremely efficient nucleator of polymerization, and its association with the barbed end of actin filaments accelerates elongation (16,17). In contrast, the isolated FH2 domain of FHOD1 does not induce actin polymerization in vivo (18). In addition, FH2 activity may also be influenced by the activity of adjacent domains. For example, in INF2, a WH2 G-actin binding motif antagonizes the nucleation activ...

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“…In selected environments, most of the components in the actin polymerization apparatus, when overexpressed or experimentally activated, have been reported to increase the growth of filopodia or microvilli (2)(3)(4)(5)(6)(7). This suggests that there is a strong intrinsic potential in cells to display these extensions, ready for implementation by factors even outside the effector and signaling chains directly related to actin.…”

mentioning

confidence: 99%

Hyaluronan Synthesis Induces Microvillus-like Cell Surface Protrusions

Kultti

Rilla

Tiihonen

et al. 2006

Journal of Biological Chemistry

124

134

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Hyaluronan synthases (HASs) are plasma membrane enzymes that simultaneously elongate, bind, and extrude the growing hyaluronan chain directly into extracellular space. In cells transfected with green fluorescent protein (GFP)-tagged Has3, the dorsal surface was decorated by up to 150 slender, 3-20-m-long microvillus-type plasma membrane protrusions, which also contained filamentous actin, the hyaluronan receptor CD44, and lipid raft microdomains. Enzymatic activity of HAS was required for the growth of the microvilli, which were not present in cells transfected with other GFP proteins or inactive GFP-Has3 mutants or in cells incubated with exogenous soluble hyaluronan. The microvilli induced by HAS3 were gradually withered by introduction of an inhibitor of hyaluronan synthesis and rapidly retracted by hyaluronidase digestion, whereas they were not affected by competition with hyaluronan oligosaccharides and disruption of the CD44 gene, suggesting independence of hyaluronan receptors. The data bring out the novel concept that the glycocalyx created by dense arrays of hyaluronan chains, tethered to HAS during biosynthesis, can induce and maintain prominent microvilli.Vertebrate cells display slender plasma membrane protrusions like filopodia, microspikes, and microvilli, the formation and maintenance of which are thought to depend on the dynamics of a bundle of actin filaments in the core of these extensions (1). In selected environments, most of the components in the actin polymerization apparatus, when overexpressed or experimentally activated, have been reported to increase the growth of filopodia or microvilli (2-7). This suggests that there is a strong intrinsic potential in cells to display these extensions, ready for implementation by factors even outside the effector and signaling chains directly related to actin. Indeed, at the molecular level, mechanisms that induce the growth of microvilli in vivo are still somewhat obscure (8). We present a novel factor, active hyaluronan synthesis, apparently unrelated to any of the previously described signaling or actin polymerization processes, that triggers extensive microvillus formation in several cell types. Interestingly, the process is primarily driven by the cell surface glycocalyx rather than intracellular systems.Hyaluronan is an ubiquitous pericellular and extracellular matrix glycosaminoglycan important in embryonic development (9), wound healing (10), inflammation (11, 12), mammalian fertilization (13), and cancer (14). It is involved in cell adhesion (15), migration (16 -19), proliferation and epithelial-mesenchymal transition (9), resistance to apoptosis, and multidrug resistance (14). Some of its effects are thought to be mediated by an ability to form a hydrated pericellular matrix, a coat that can modify cellular interactions with other matrix components and neighboring cells, directly regulating processes such as differentiation and migration (20). Some of the biological signals elicited by hyaluronan are dependent on its CD44 receptor on cell su...

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The Rho Family GTPase Rif Induces Filopodia through mDia2

Cited by 263 publications

References 23 publications

Differing and isoform-specific roles for the formin DIAPH3 in plasma membrane blebbing and filopodia formation

Differing and isoform-specific roles for the formin DIAPH3 in plasma membrane blebbing and filopodia formation

Interaction of the N- and C-terminal Autoregulatory Domains of FRL2 Does Not Inhibit FRL2 Activity

Hyaluronan Synthesis Induces Microvillus-like Cell Surface Protrusions

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