scholarly journals Ultrastructural analysis of the dynactin complex: an actin-related protein is a component of a filament that resembles F-actin.

1994 ◽  
Vol 126 (2) ◽  
pp. 403-412 ◽  
Author(s):  
D A Schafer ◽  
S R Gill ◽  
J A Cooper ◽  
J E Heuser ◽  
T A Schroer
Keyword(s):  
Electron Microscopy ◽  
Binding Protein ◽  
Terminal Region ◽  
Ultrastructural Analysis ◽  
Actin Binding ◽  
Related Protein ◽  
Actin Binding Protein ◽  
Capping Protein ◽  
Short Filament ◽  
Dynactin Complex

The dynactin complex visualized by deepetch electron microscopy appears as a short filament 37-nm in length, which resembles F-actin, plus a thinner, laterally oriented filament that terminates in two globular heads. The locations of several of the constituent polypeptides were identified on this structure by applying antibodies to decorate the dynactin complex before processing for electron microscopy. Antibodies to the actin-related protein Arp1 (previously referred to as actin-RPV), bound at various sites along the filament, demonstrating that this protein assembles in a polymer similar to conventional actin. Antibodies to the barbed-end actin-binding protein, capping protein, bound to one end of the filament. Thus, an actin-binding protein that binds conventional actin may also bind to Arp1 to regulate its polymerization. Antibodies to the 62-kD component of the dynactin complex also bound to one end of the filament. An antibody that binds the COOH-terminal region of the 160/150-kD dynactin polypeptides bound to the globular domains at the end of the thin lateral filament, suggesting that the dynactin polypeptide comprises at least part of the sidearm structure.

scholarly journals Heterodimeric Capping Protein from Arabidopsis Is a Membrane-Associated, Actin-Binding Protein

2014 ◽  
Vol 166 (3) ◽  
pp. 1312-1328 ◽  
Author(s):  
J. C. Jimenez-Lopez ◽  
X. Wang ◽  
S. O. Kotchoni ◽  
S. Huang ◽  
D. B. Szymanski ◽  
...  
Keyword(s):  
Binding Protein ◽  
Actin Binding ◽  
Actin Binding Protein ◽  
Capping Protein

scholarly journals The architecture of actin filaments and the ultrastructural location of actin-binding protein in the periphery of lung macrophages.

1986 ◽  
Vol 103 (3) ◽  
pp. 1007-1020 ◽  
Author(s):  
J H Hartwig ◽  
P Shevlin
Keyword(s):  
Electron Microscopy ◽  
Critical Point ◽  
Binding Protein ◽  
The Body ◽  
Actin Binding ◽  
Actin Network ◽  
Actin Binding Protein ◽  
Freeze Dried ◽  
Critical Point Drying ◽  
Actin Fiber

A highly branched filament network is the principal structure in the periphery of detergent-extracted cytoskeletons of macrophages that have been spread on a surface and either freeze or critical point dried, and then rotary shadowed with platinum-carbon. This array of filaments completely fills lamellae extended from the cell and bifurcates to form 0.2-0.5 micron thick layers on the top and bottom of the cell body. Reaction of the macrophage cytoskeletons with anti-actin IgG and with anti-IgG bound to colloidal gold produces dense staining of these filaments, and incubation with myosin subfragment 1 uniformly decorates these filaments, identifying them as actin. 45% of the total cellular actin and approximately 70% of actin-binding protein remains in the detergent-insoluble cell residue. The soluble actin is not filamentous as determined by sedimentation analysis, the DNAase I inhibition assay, and electron microscopy, indicating that the cytoskeleton is not fragmented by detergent extraction. The spacing between the ramifications of the actin network is 94 +/- 47 nm and 118 +/- 72 nm in cytoskeletons prepared for electron microscopy by freeze drying and critical point drying, respectively. Free filament ends are rare, except for a few which project upward from the body of the network or which extend down to the substrate. Filaments of the network intersect predominantly at right angles to form either T-shaped and X-shaped overlaps having striking perpendicularity or else Y-shaped intersections composed of filaments intersecting at 120-130 degrees angles. The actin filament concentration in the lamellae is high, with an average value of 12.5 mg/ml. The concentration was much more uniform in freeze-dried preparations than in critical point-dried specimens, indicating that there is less collapse associated with the freezing technique. The orthogonal actin network of the macrophage cortical cytoplasm resembles actin gels made with actin-binding protein. Reaction of cell cytoskeletons and of an actin gel made with actin-binding protein with anti-actin-binding protein IgG and anti-IgG-coated gold beads resulted in the deposition of clusters of gold at points where filaments intersect and at the ends of filaments that may have been in contact with the membrane before its removal with detergent. In the actin gel made with actin-binding protein, 75% of actin-fiber intersections labeled, and the filament spacing between intersections is consistent with that predicted on theoretical grounds if each added actin-binding protein molecule cross-links two filaments to form an intersection in the gel.(ABSTRACT TRUNCATED AT 400 WORDS)

scholarly journals Homotypic dimerization of the actin-binding protein p57/coronin-1 mediated by a leucine zipper motif in the C-terminal region

2005 ◽  
Vol 387 (2) ◽  
pp. 325-331 ◽  
Author(s):  
Teruaki OKU ◽  
Saotomo ITOH ◽  
Rie ISHII ◽  
Kensuke SUZUKI ◽  
William M. NAUSEEF ◽  
...  
Keyword(s):  
Leucine Zipper ◽  
Fluorescent Protein ◽  
Binding Protein ◽  
Coiled Coil ◽  
Terminal Region ◽  
Full Length ◽  
Actin Binding ◽  
Leucine Zipper Motif ◽  
Actin Binding Protein ◽  
Coiled Coil Domain

The actin-binding protein p57/coronin-1, a member of the coronin protein family, is selectively expressed in immune cells, and has been implicated in leucocyte migration and phagocytosis by virtue of its interaction with F-actin (filamentous actin). We previously identified two sites in the N-terminal region of p57/coronin-1 by which it binds actin, and in the present study we examine the role of the leucine zipper motif located in the C-terminal coiled-coil domain in mediating the homotypic association of p57/coronin-1. Recombinant p57/coronin-1 protein in solution formed a homodimer, as analysed by Superose 12 column chromatography and by sucrose density gradient centrifugation. In vivo, a truncated form consisting of the C-terminal coiled-coil domain co-precipitated with full-length p57/coronin-1 when both were co-expressed in COS-1 cells. A chimaeric construct composed of the C-terminal domain of p57/coronin-1 (which lacks the actin-binding sites) fused with green fluorescent protein co-localized with cortical F-actin-rich regions in COS-1 cells only when full-length p57/coronin-1 was expressed simultaneously in the cells, suggesting that the C-terminal region is required for the homotypic association of p57/coronin-1. Furthermore, p57LZ, a polypeptide consisting of the C-terminal 90 amino acid residues of p57/coronin-1, was sufficient for dimerization. When two leucine residues out of the four that constitute the leucine zipper structure in p57LZ or full-length p57 were replaced with alanine residues, the mutants failed to form homodimers. Taken together, these results demonstrate that p57/coronin-1 forms homodimers, that the association is mediated by the leucine zipper structure in the C-terminal region, and that it plays a role in the cross-linking of F-actin in the cell.

scholarly journals Structural comparison of several actin-binding macromolecules.

1980 ◽  
Vol 85 (2) ◽  
pp. 489-495 ◽  
Author(s):  
J M Tyler ◽  
J M Anderson ◽  
D Branton
Keyword(s):  
Electron Microscopy ◽  
Binding Protein ◽  
Actin Binding ◽  
Erythrocyte Membranes ◽  
Structural Comparison ◽  
Flexible Molecules ◽  
Actin Binding Protein ◽  
Rotary Shadowing

The cytoskeletal components, macrophage actin-binding protein and filamin, were dried from glycerol and examined by low-angle rotary shadowing electron microscopy. Both are elongate, flexible molecules whose general morphologi is similar to that of erythrocyte spectrin. Neither actin-binding protein nor filamin binds to spectrin-depleted erythrocyte membranes.

scholarly journals Neurabin: A Novel Neural Tissue–specific Actin Filament–binding Protein Involved in Neurite Formation

1997 ◽  
Vol 139 (4) ◽  
pp. 951-961 ◽  
Author(s):  
Hiroyuki Nakanishi ◽  
Hiroshi Obaishi ◽  
Ayako Satoh ◽  
Manabu Wada ◽  
Kenji Mandai ◽  
...  
Keyword(s):  
Rat Brain ◽  
Actin Filament ◽  
Binding Protein ◽  
Neural Tissue ◽  
Terminal Region ◽  
Actin Binding ◽  
Calculated Molecular Mass ◽  
Tissue Specific ◽  
Actin Binding Protein ◽  
Neurite Formation

We purified from rat brain a novel actin filament (F-actin)–binding protein of ∼180 kD (p180), which was specifically expressed in neural tissue. We named p180 neurabin (neural tissue–specific F-actin– binding protein). We moreover cloned the cDNA of neurabin from a rat brain cDNA library and characterized native and recombinant proteins. Neurabin was a protein of 1,095 amino acids with a calculated molecular mass of 122,729. Neurabin had one F-actin–binding domain at the NH2-terminal region, one PSD-95, DlgA, ZO-1–like domain at the middle region, a domain known to interact with transmembrane proteins, and domains predicted to form coiled-coil structures at the COOH-terminal region. Neurabin bound along the sides of F-actin and showed F-actin–cross-linking activity. Immunofluorescence microscopic analysis revealed that neurabin was highly concentrated in the synapse of the developed neurons. Neurabin was also concentrated in the lamellipodia of the growth cone during the development of neurons. Moreover, a study on suppression of endogenous neurabin in primary cultured rat hippocampal neurons by treatment with an antisense oligonucleotide showed that neurabin was involved in the neurite formation. Neurabin is a candidate for key molecules in the synapse formation and function.

scholarly journals A calcium- and pH-regulated protein from Dictyostelium discoideum that cross-links actin filaments.

1982 ◽  
Vol 94 (2) ◽  
pp. 466-471 ◽  
Author(s):  
J Condeelis ◽  
M Vahey
Keyword(s):  
Electron Microscopy ◽  
Dictyostelium Discoideum ◽  
Actin Filaments ◽  
Binding Protein ◽  
Actin Binding ◽  
Cross Linking ◽  
Optimal Conditions ◽  
Maximal Inhibition ◽  
Actin Binding Protein ◽  
Cross Links

We have purified an actin binding protein from amebas of Dictyostelium discoideum which we call 95,000-dalton protein (95K). This protein is rod shaped, approximately 40 nm long in the electron microscope, contains two subunits measuring 95,000 daltons each, and cross-links actin filaments. Cross-linking activity was demonstrated by using falling-ball viscometry, Ostwald viscometry, and electron microscopy. Cross-linking activity is optimal at 0.1 microM Ca++ and pH 6.8, but is progressively inhibited at higher Ca++ and pH levels over a physiological range. Half-maximal inhibition occurs at 1.6 microM free Ca++ and pH 7.3, respectively. Sedimentation experiments demonstrate that elevated Ca++ and pH inhibit the binding of 95K to F-actin which explains the loss of cross-linking activity. Electron microscopy demonstrates that under optimal conditions for cross-linking, 95K protein bundles actin filaments and that this bundling is inhibited by microM Ca++. Severing of actin filaments by 95K was not observed in any of the various assays under any of the solution conditions used. Hence, 95K protein is a rod-shaped, dimeric, Ca++- and pH-regulated actin binding protein that cross-links but does not sever actin filaments.

Clathrin Hub Expression Dissociates the Actin-Binding Protein Hip1R from Coated Pits and Disrupts Their Alignment with the Actin Cytoskeleton

Traffic ◽  
2001 ◽  
Vol 2 (11) ◽  
pp. 851-858 ◽  
Author(s):  
Elizabeth M. Bennett ◽  
Chih-Ying Chen ◽  
Asa E. Y. Engqvist-Goldstein ◽  
David G. Drubin ◽  
Frances M. Brodsky
Keyword(s):  
Actin Cytoskeleton ◽  
Binding Protein ◽  
Actin Binding ◽  
Coated Pits ◽  
Actin Binding Protein
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