Dynacortin Is a Novel Actin Bundling Protein That Localizes to Dynamic Actin Structures

Robinson, Douglas N.; Ocon, Stephani S.; Rock, Ronald S.; Spudich, James A.

doi:10.1074/jbc.m112144200

Cited by 25 publications

(22 citation statements)

References 30 publications

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“…Moreover, PTEN and cortexillin I have PtdIns(4,5)P 2 -binding domains, which suggest that these proteins are posterior and furrow residing proteins (Iijima et al, 2004;Stock et al, 1999). Conversely, dynacortin becomes highly enriched in cortical protrusions and at leading edge of polarized cells and likely contributes to actin assembly (Girard et al, 2004;Kabacoff et al, 2007;Robinson et al, 2002).…”

Section: Discussionmentioning

confidence: 99%

“…Activated Ras, PI3K2 and PKBA are localized at the leading edge of chemotaxing cells in cAMP gradients and help regulate the actin cytoskeleton (Kamimura et al, 2008;Sasaki and Firtel, 2009). Coronin, dynacortin and LimE are all actin-binding proteins that localize at the leading edge of a migrating cell but interact with actin differently: coronin crosslinks F-actin filaments (de Hostos et al, 1991) and dynacortin bundles F-actin filaments at the leading edge (Girard et al, 2004;Kabacoff et al, 2007;Robinson et al, 2002) while LimE binds and forms a complex with Rac and Factin and other actin-binding proteins at the cortex of cell projections (Prassler et al, 1998). Conversely, the tumor suppressor and phosphatase and tensin homolog (PTEN) has been shown to be distributed on the lateral and trailing edge of the plasma membrane of the cell (Funamoto et al, 2002;Iijima and Devreotes, 2002).…”

Section: Introductionmentioning

confidence: 99%

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Delineating the core regulatory elements crucial for directed cell migration by examining folic-acid-mediated responses

Srinivasan

Wright

Hames

et al. 2013

Journal of Cell Science

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SummaryDictyostelium discoideum shows chemotaxis towards folic acid (FA) throughout vegetative growth, and towards cAMP during development. We determined the spatiotemporal localization of cytoskeletal and signaling molecules and investigated the FA-mediated responses in a number of signaling mutants to further our understanding of the core regulatory elements that are crucial for cell migration. Proteins enriched in the pseudopods during chemotaxis also relocalize transiently to the plasma membrane during uniform FA stimulation. In contrast, proteins that are absent from the pseudopods during migration redistribute transiently from the PM to the cytosol when cells are globally stimulated with FA. These chemotactic responses to FA were also examined in cells lacking the GTPases Ras C and G. Although Ras and phosphoinositide 3-kinase activity were significantly decreased in Ras G and Ras C/G nulls, these mutants still migrated towards FA, indicating that other pathways must support FA-mediated chemotaxis. We also examined the spatial movements of PTEN in response to uniform FA and cAMP stimulation in phospholipase C (PLC) null cells. The lack of PLC strongly influences the localization of PTEN in response to FA, but not cAMP. In addition, we compared the gradient-sensing behavior of polarized cells migrating towards cAMP to that of unpolarized cells migrating towards FA. The majority of polarized cells make U-turns when the cAMP gradient is switched from the front of the cell to the rear. Conversely, unpolarized cells immediately extend pseudopods towards the new FA source. We also observed that plasma membrane phosphatidylinositol 3,4,5-trisphosphate [PtdIns(3,4,5)P 3 ] levels oscillate in unpolarized cells treated with Latrunculin-A, whereas polarized cells had stable plasma membrane PtdIns(3,4,5)P 3 responses toward the chemoattractant gradient source. Results were similar for cells that were starved for 4 hours, with a mixture of polarized and unpolarized cells responding to cAMP. Taken together, these findings suggest that similar components control gradient sensing during FA-and cAMP-mediated motility, but the response of polarized cells is more stable, which ultimately helps maintain their directionality.

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Section: Discussionmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

Delineating the core regulatory elements crucial for directed cell migration by examining folic-acid-mediated responses

Srinivasan

Wright

Hames

et al. 2013

Journal of Cell Science

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“…Low Centrifugal Force Sedimentation of Cross-linked Actin Filaments-Bundling of F-actin was determined by standard sedimentation assays (17)(18)(19). 10 M G-actin was polymerized in buffer F in the presence of 0.4 M human gelsolin (cytoskeleton) to get similar filament sizes and then stabilized with 10 M phalloidin.…”

Section: Methodsmentioning

confidence: 99%

The V-ATPase Subunit C Binds to Polymeric F-actin as Well as to Monomeric G-actin and Induces Cross-linking of Actin Filaments

Vitavska

Merzendorfer

Wieczorek

2005

Journal of Biological Chemistry

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Previously, we have shown that the V-ATPase holoenzyme as well as the V 1 complex isolated from the midgut of the tobacco hornworm (Manduca sexta) exhibits the ability of binding to actin filaments via the V 1 subunits B and C (Vitavska, O., Wieczorek, H., and Merzendorfer, H. (2003) J. Biol. Chem. 278, 18499 -18505). Since the recombinant subunit C not only enhances actin binding of the V 1 complex but also can bind separately to F-actin, we analyzed the interaction of recombinant subunit C with actin. We demonstrate that it binds not only to F-actin but also to monomeric G-actin. With dissociation constants of ϳ50 nM, the interaction exhibits a high affinity, and no difference could be observed between binding to ATP-G-actin or ADP-G-actin, respectively. Unlike other proteins such as members of the ADF/cofilin family, which also bind to G-as well as to F-actin, subunit C does not destabilize actin filaments. On the contrary, under conditions where the disassembly of F-actin into G-actin usually occurred, subunit C stabilized F-actin. In addition, it increased the initial rate of actin polymerization in a concentration-dependent manner and was shown to cross-link actin filaments to bundles of varying thickness. Apparently bundling is enabled by the existence of at least two actin-binding sites present in the N-and in the C-terminal halves of subunits C, respectively. Since subunit C has the possibility to dimerize or even to oligomerize, spacing between actin filaments could be variable in size.V-ATPases are ubiquitous and highly conserved proton pumps that acidify specific organelles such as endosomes, lysosomes, or secretory vesicles in every eukaryotic cell (1). They also are found in plasma membranes of many specialized animal cells where they either are involved in pH homeostasis or in membrane energization (2). V-ATPases consist of two complexes, a peripheral V 1 complex whose catalytic part faces the cytosol and a membrane-bound proton-conducting V 0 complex. In the midgut of the tobacco hornworm (Manduca sexta), the V 1 complex of the plasma membrane V-ATPase contains eight different subunits, A-H, whereas the V 0 complex consists of the four different subunits a and c-e (3). Under special physiological conditions V-ATPase activity is down-regulated by reversible dissociation of the V 1 complex from the membrane as was shown in the tobacco hornworm as well as in yeast (4,5).Subunit C appears to be released into the cytoplasm during this process, because the purified V 1 complex lacks most of it (3, 6).Dissociation of subunit C from the V 1 complex and its support of holoenzyme reassembly indicate that this subunit may play a crucial role in the regulation of V-ATPases. Another role, previously detected by us in the M. sexta midgut, is its ability to bind to the actin cytoskeleton (7). In feeding tobacco hornworms, actin filaments co-localize with the V-ATPase at the apical membrane of midgut goblet cells. Like in osteoclasts (8), actin binding occurs via the V 1 subunit B; however, binding to F-actin al...

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“…Because enlazin was discovered in the same genetic suppressor selection as dynacortin, we speculated that enlazin might assemble into punctate surface structures, which are probably the Dictyostelium equivalents of focal adhesions (Robinson et al, 2002b). To test this, we examined wt:GFPenlazin cells by TIRF microscopy.…”

Section: Enlazin Is a Cortically Enriched Proteinmentioning

confidence: 99%

Enlazin, a Natural Fusion of Two Classes of Canonical Cytoskeletal Proteins, Contributes to Cytokinesis Dynamics

Octtaviani¹,

Effler²,

Robinson³

2006

MBoC

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Cytokinesis requires a complex network of equatorial and global proteins to regulate cell shape changes. Here, using interaction genetics, we report the first characterization of a novel protein, enlazin. Enlazin is a natural fusion of two canonical classes of actin-associated proteins, the ezrin-radixin-moesin family and fimbrin, and it is localized to actin-rich structures. A fragment of enlazin, enl-tr, was isolated as a genetic suppressor of the cytokinesis defect of cortexillin-I mutants. Expression of enl-tr disrupts expression of endogenous enlazin, indicating that enl-tr functions as a dominantnegative lesion. Enlazin is distributed globally during cytokinesis and is required for cortical tension and cell adhesion. Consistent with a role in cell mechanics, inhibition of enlazin in a cortexillin-I background restores cytokinesis furrowing dynamics and suppresses the growth-in-suspension defect. However, as expected for a role in cell adhesion, inhibiting enlazin in a myosin-II background induces a synthetic cytokinesis phenotype, frequently arresting furrow ingression at the dumbbell shape and/or causing recession of the furrow. Thus, enlazin has roles in cell mechanics and adhesion, and these roles seem to be differentially significant for cytokinesis, depending on the genetic background.

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Dynacortin Is a Novel Actin Bundling Protein That Localizes to Dynamic Actin Structures

Cited by 25 publications

References 30 publications

Delineating the core regulatory elements crucial for directed cell migration by examining folic-acid-mediated responses

Delineating the core regulatory elements crucial for directed cell migration by examining folic-acid-mediated responses

The V-ATPase Subunit C Binds to Polymeric F-actin as Well as to Monomeric G-actin and Induces Cross-linking of Actin Filaments

Enlazin, a Natural Fusion of Two Classes of Canonical Cytoskeletal Proteins, Contributes to Cytokinesis Dynamics

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