Polarized bundles of actin filaments within microvilli of fertilized sea urchin eggs.

Burgess, David R.; Schroeder, Thomas E.

doi:10.1083/jcb.74.3.1032

Cited by 202 publications

(86 citation statements)

References 13 publications

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“…Fascin is recruited to the cortical cytoskeleton to form microvilli at fertilization of eggs (8) and filopodia of coelomocytes, phagocytic cells from the coelomic cavity (9,10). Those structures have actin bundle cores with an 11-nm transverse crossbanding pattern (11,12), which is also observed in actin bundles formed with fascin in vitro (7). Thus, those studies have demonstrated that echinoid fascin plays crucial roles in forming actin bundles.…”

Section: ؊1mentioning

confidence: 83%

Identification of an Actin Binding Region and a Protein Kinase C Phosphorylation Site on Human Fascin

Ono

Yamakita

Yamashiro

et al. 1997

Journal of Biological Chemistry

172

195

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Fascin is a 55-58-kDa actin-bundling protein, the actin binding of which is regulated by phosphorylation (Yamakita, Y., Ono, S., Matsumura, F., and Yamashiro, S. . Second, we identified the phosphorylation site of fascin as Ser-39 by sequencing a tryptic phosphopeptide purified by chelating column chromatography followed by C-18 reverse phase high performance liquid chromatography. Peptide map analyses revealed that the purified peptide represented the major phosphorylation site of in vivo as well as in vitro phosphorylated fascin. The mutation replacingSer-39withAlaeliminatedthephosphorylation-dependent regulation of actin binding of fascin, indicating that phosphorylation at this site regulates the actin binding ability of fascin.

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Section: ؊1mentioning

confidence: 83%

Identification of an Actin Binding Region and a Protein Kinase C Phosphorylation Site on Human Fascin

Ono

Yamakita

Yamashiro

et al. 1997

Journal of Biological Chemistry

172

195

View full text Add to dashboard Cite

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“…In other studies (9,42) bundle formation was studied at pHs ranging from 7.3 to 7.8. Whether this pH difference with respect to the more optimal low pH conditions (6.8-7 .0) would alter the results of kinetic 65 4 THE JOURNAL OF CELL 810LOGY " VOLUME 92, 1982 80 0 FIGURE 12 The amount of protein found in the pellet is dependent on the pH of the solution . As the pH increases from 6.6 to 7.6 the amount of protein sedimented in the microfuge (dots) or the airfuge (circles) decreases.…”

Section: Ca ++ Dependence Of Bundlingmentioning

confidence: 99%

Partial reconstruction of the microvillus core bundle: characterization of villin as a Ca(++)-dependent, actin-bundling/depolymerizing protein

Matsudaira¹,

Burgess²

1982

The Journal of Cell Biology

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The brush border, isolated from chicken intestine epithelial cells, contains the 95,000 relative molecular mass NO polypeptide, villin . This report describes the purification and characterization of villin as a Ca"-dependent, actin bundling/depolymerizing protein .The 100,000 g supernatant from a Ca" extract of isolated brush borders is composed of three polypeptides of 95,000 (villin), 68,000 (fimbrin), and 42,000 M r (actin) . Villin, following purification from this extract by differential ammonium sulfate precipitation and ion-exchange chromatography, was mixed with skeletal muscle F-actin . Electron microscopy of negatively stained preparations of these villin-actin mixtures showed that filament bundles were present . The viscosity, sedimentability, and ultrastructural morphology of filament bundles are dependent on the villin :actin molar ratio, the pH, and the free Ca" concentration in solution . At low free Ca" (<10-6 M), the amount of protein in bundles, when measured by sedimentation, increased as the villin: actin molar ratio increased and reached a plateau at approximately a 4:10 ratio . This behavior correlates with the conversion of single actin filaments into filament bundles as detected in the electron microscope . At high free Ca" (>10-6 M), there was a decrease in the apparent viscosity in the villin-actin mixtures to a level measured for the buffer. Furthermore, these Ca" effects were correlated with the loss of protein sedimented, the disappearance of filament bundles, and the appearance of short fragments of filaments . Bundle formation is also pH-sensitive, being favored at mildly acidic pH . A decrease in the pH from 7 .6 to 6.6 results in an increase in sedimentable protein and also a transformation of loosely associated actin filaments into compact actin bundles . These results are consistent with the suggestions that villin is a bundling protein in the microvillus and is responsible for the Ca"-sensitive disassembly of the microvillar cytoskeleton . Thus villin may function in the cytoplasm as a major cytoskeletal element regulating microvillar shape .The brush border microvillus is a model biological system for investigations aimed at understanding how a cell maintains its shape. Ultrastructural studies have shown that the microvillus contains a core of bundled microfilaments which extend into the underlying terminal web (44,45) . This core bundle is associated with the overlying plasma membrane by two types of attachments. The first type laterally connects the membrane along the length of the bundle by means of cross filaments (35,39,44,45). In the second type of membrane attachment, the microfilaments insert end-on into the electron-dense cap at the tips of the microvilli. Earlier studies had suggested that both types of attachments as well as the bundling protein in the microvillus core were composed of alpha-actinin (3,44,48) . However, recent studies have shown instead that alpha-actinin is not a structural component of the microvillus but rather is a component of the te...

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“…Although cytochalasin D (CD) has multiple effects, in vitro it has been shown to bind primarily to the barbed end ofactin filaments (4,17,18,28). In vivo, CD blocks the peptide-induced increase in actin polymerization (35).…”

Section: Fnorleleuphe Treatment Increases Free Barbed Ends In Pmn Lysmentioning

confidence: 99%

An actin-nucleating activity in polymorphonuclear leukocytes is modulated by chemotactic peptides.

Carson¹,

Weber²,

Zigmond³

1986

The Journal of Cell Biology

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Abstract. We examined the actin-nucleating activity in polymorphonuclear leukocyte lysates prepared at various times after chemotactic peptide addition. The actin nucleation increases two-to threefold within 15 s after peptide addition, decays to basal levels within 90 s, and is largely independent of cytoplasmic calcium fluxes. The peptide-induced nucleation sites behave as free barbed ends and therefore may increase the level of polymerized actin in vivo. The new nucleation sites may also determine the cellular sites of actin polymerization. This localization of actin polymerization could be important for the directional extension of lamellipodia during chemotaxis.D1r~G chemotactic peptides to polymorphonuclear leukocytes (PMNs) ~ induces actin polymerization (F-actin) (9,11,12,23,27,35). Concomitantly, PMNs spread and extend ruffles over their surface. Both responses peak within 30 s. At later times some of the ruffles withdraw and the amount of F-actin decreases to an intermediate level. Cytochalasin D (CD), which blocks the peptide-induced actin polymerization, inhibits the cell shape change (35,38). Consequently, actin polymerization appears to be required for the protrusion of surface ruffles in PMNs. The extension of filopodia on platelets, of the acrosomal process on thyone sperm, and of microvilli on fertilized sea urchin eggs have been previously shown to correlate with actin polymerization in similar studies (8,10,32).The cell must have mechanisms for preventing complete actin polymerization. Actin, in the ionic conditions and at the concentrations present within a cell, would always be 99 % polymerized in the absence of regulation (13, 30). However, by a number of criteria (actin sedimentation, inhibition of DNAase activity, nitrobenzoxadiazole phallacidin staining), 50-70% of the actin is unpolymerized in an unstimulated PMN (9, 23). This state could be maintained by regulatory proteins that bind free actin monomers with high affinity, thus preventing their polymerization, and by proteins that cap filament ends, thus blocking polymerization.Proteins with these properties, able to regulate actin polymerization, have been isolated from PMNs (13, 30). Profilin binds monomeric actin, preventing monomer addition to the nonpreferred (pointed) end of actin filaments but not to the preferred (barbed) end (5, 33, 34). Profilin could maintain a large pool of unpolymerized actin in PMNs in conjunction with a capping protein such as gelsolin, which blocks polymerization on the barbed end of an actin filament, (15, 39). Chemotactic peptides, therefore, could increase the fraction of actin polymerized by releasing actin monomers, by uncapping pre-existing filaments, or by creating new barbed end polymerization sites.Studies of isolated actin regulatory proteins suggest that none of the receptor-generated signals would yield the appropriate regulation of actin polymerization. The binding of chemotactic peptide to its receptor causes a transient decrease in the intracellular concentration of phosphafidylinositol 4...

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Polarized bundles of actin filaments within microvilli of fertilized sea urchin eggs.

Cited by 202 publications

References 13 publications

Identification of an Actin Binding Region and a Protein Kinase C Phosphorylation Site on Human Fascin

Identification of an Actin Binding Region and a Protein Kinase C Phosphorylation Site on Human Fascin

Partial reconstruction of the microvillus core bundle: characterization of villin as a Ca(++)-dependent, actin-bundling/depolymerizing protein

An actin-nucleating activity in polymorphonuclear leukocytes is modulated by chemotactic peptides.

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