scholarly journals The polymerization of actin: II. how nonfilamentous actin becomes nonrandomly distributed in sperm: evidence for the association of this actin with membranes

1976 ◽  
Vol 69 (1) ◽  
pp. 51-72 ◽  
Author(s):  
LG Tilney
Keyword(s):  
Plasma Membrane ◽  
Nuclear Envelope ◽  
Thin Section ◽  
Actin Filaments ◽  
Early Stage ◽  
Mature Sperm ◽  
Vacuole Membrane ◽  
Other Substances ◽  
Associated Proteins ◽  
Basal Half

At an early stage in spermiogenesis the acrosomal vacuole and other organelles including ribosomes are located at the basal end of the cell. From here actin must be transported to its future location at the anterior end of the cell. At no stage in the accumulation of actin in the periacrosomal region is the actin sequestered in a membrane-bounded compartment such as a vacuole or vesicle. Since filaments are not present in the periacrosomal region during the accumulation of the actin even though the fixation of these cells is sufficiently good to distinguish actin filaments in thin section, the actin must accumulate in the nonfilamentous state. The membranes in the periacrosomal region, specifically a portion of the nuclear envelope and the basal half of the acrosomal vacuole membrane, become specialized morphologically in advance of the accumulation of actin in this region. My working hypothesis is that the actin in combination with other substances binds to these specialized membranes and to itself and thus can accumulate in the periacrosmoal region by being trapped on these specialized membranes. Diffusion would then be sufficient to move these substances to this region. In support of this hypothesis are experiments in which I treated mature sperm with detergents, glycols, and hypotonic media, which solubilize or lift away the plasma membrane. The actin and its associated proteins remain attached to these specialized membranes. Thus actin can be nonrandomly distributed in cells in a nonfilamentous state presumably by its association with specialized membranes.

scholarly journals Phagocytosis of bacteria by polymorphonuclear leukocytes: a freeze-fracture, scanning electron microscope, and thin-section investigation of membrane structure

1978 ◽  
Vol 76 (1) ◽  
pp. 158-174 ◽  
Author(s):  
PL Moore ◽  
HL Bank ◽  
NT Brissie ◽  
SS Spicer
Keyword(s):  
Plasma Membrane ◽  
Electron Microscope ◽  
Membrane Fusion ◽  
Thin Section ◽  
Membrane Structure ◽  
Freeze Fracture ◽  
Vacuole Membrane ◽  
Attachment Sites ◽  
Pmn Leukocytes ◽  
Scanning Electron

The changes in membrane structure of rabbit polymorphonuclear (PMN) leukocytes during bacterial phagocytosis was investigated with scanning electron microscope (SEM), thin-section, and freeze-fracture techniques. SEM observations of bacterial attachment sites showed the involvement of limited areas of PMN membrane surface (0.01-0.25μm(2)). Frequently, these areas of attachment were located on membrane extensions. The membrane extensions were present before, during, and after the engulfment of bacteria, but were diminished in size after bacterial engulfment. In general, the results obtained with SEM and thin-section techniques aided in the interpretation of the three-dimensional freeze-fracture replicas. Freeze-fracture results revealed the PMN leukocytes had two fracture faces as determined by the relative density of intramembranous particles (IMP). Membranous extensions of the plasma membrane, lysosomes, and phagocytic vacuoles contained IMP's with a distribution and density similar to those of the plasma membrane. During phagocytosis, IMPs within the plasma membrane did not undergo a massive aggregation. In fact, structural changes within the membranes were infrequent and localized to regions such as the attachment sites of bacteria, the fusion sites on the plasma membrane, and small scale changes in the phagocytic vacuole membrane during membrane fusion. During the formation of the phagocytic vacuole, the IMPs of the plasma membrane appeared to move in with the lipid bilayer while maintaining a distribution and density of IMPs similar to those of the plasma membranes. Occasionally, IMPs were aligned to linear arrays within phagocytic vacuole membranes. This alignment might be due to an interaction with linearly arranged motile structures on the side of the phagocytic vacuole membranes. IMP-free regions were observed after fusion of lysosomes with the phagocytic vacuoles or plasma membrane. These IMP-free areas probably represent sites where membrane fusion occurred between lysosomal membrane and phagocytic vacuole membrane or plasma membrane. Highly symmetrical patterns of IMPs were not observed during lysosomal membrane fusion.

The molecular mechanisms of the mammalian exocyst complex in exocytosis

2006 ◽  
Vol 34 (5) ◽  
pp. 687-690 ◽  
Author(s):  
S. Wang ◽  
S.C. Hsu
Keyword(s):  
Plasma Membrane ◽  
Molecular Mechanisms ◽  
Early Stage ◽  
Secretory Vesicles ◽  
Attachment Protein ◽  
Trafficking Pathway ◽  
Exocyst Complex ◽  
Protein Receptor ◽  
Associated Proteins ◽  
Protein Attachment

Exocytosis is a highly ordered vesicle trafficking pathway that targets proteins to the plasma membrane for membrane addition or secretion. Research over the years has discovered many proteins that participate at various stages in the mammalian exocytotic pathway. At the early stage of exocytosis, co-atomer proteins and their respective adaptors and GTPases have been shown to play a role in the sorting and incorporation of proteins into secretory vesicles. At the final stage of exocytosis, SNAREs (soluble N-ethylmaleimide-sensitive fusion protein attachment protein receptor) and SNARE-associated proteins are believed to mediate the fusion of secretory vesicles at the plasma membrane. There are multiple events that may occur between the budding of secretory vesicles from the Golgi and the fusion of these vesicles at the plasma membrane. The most obvious and best-known event is the transport of secretory vesicles from Golgi to the vicinity of the plasma membrane via microtubules and their associated motors. At the vicinity of the plasma membrane, however, it is not clear how vesicles finally dock and fuse with the plasma membrane. Identification of proteins involved in these events should provide important insights into the mechanisms of this little known stage of the exocytotic pathway. Currently, a protein complex, known as the sec6/8 or the exocyst complex, has been implicated to play a role at this late stage of exocytosis.

scholarly journals Fodrin and band 4.1 in a plasma membrane-associated fraction of human neutrophils

Blood ◽  
1989 ◽  
Vol 74 (6) ◽  
pp. 2136-2143 ◽  
Author(s):  
KB Stevenson ◽  
RA Clark ◽  
WM Nauseef
Keyword(s):  
Plasma Membrane ◽  
Actin Filaments ◽  
Polymorphonuclear Leukocytes ◽  
Membrane Vesicles ◽  
Human Neutrophils ◽  
Plasma Membrane Vesicles ◽  
Associated Proteins ◽  
Human Pmns ◽  
Discontinuous Percoll Gradient ◽  
Human Polymorphonuclear Leukocytes

Abstract Erythrocytes possess a well-characterized submembranous filamentous network which interacts with transmembrane glycoproteins and is composed primarily of spectrin, ankyrin, band 4.1, and short actin filaments. An analogous structure was recently described in platelets. Human polymorphonuclear leukocytes (PMNs) were examined for the presence and plasma membrane association of similar proteins. Isolated PMNs, free of contamination with erythrocytes or platelets, were disrupted by nitrogen cavitation and separated into subcellular organelles on a discontinuous Percoll gradient. Detergent lysates of plasma membrane vesicles, but not azurophilic or specific granules, contained insoluble actin filaments and associated proteins. Immunoblots of detergent-insoluble plasma membrane fractions contained proteins recognized by antibodies to brain fodrin and erythrocyte band 4.1, whereas blots probed with antibodies to erythrocyte spectrin and ankyrin were negative. Fodrin and band 4.1 were not detected in granule fractions, but some fodrin was present in the cytosol. The association of proteins related to fodrin and band 4.1 with the plasma membrane suggests that PMNs contain a submembranous skeleton structurally analogous to that of erythrocytes and platelets. The specific function of these proteins and their structural organization in human PMNs await further study.

scholarly journals Actin filaments elongate from their membrane-associated ends

1981 ◽  
Vol 90 (2) ◽  
pp. 485-494 ◽  
Author(s):  
LG Tilney ◽  
EM Bonder ◽  
DJ DeRosier
Keyword(s):  
Plasma Membrane ◽  
Actin Filament ◽  
Actin Filaments ◽  
The Other ◽  
Dense Material ◽  
Thin Sections ◽  
Vacuole Membrane ◽  
Filament Bundles ◽  
Filament Bundle

In limulus sperm an actin filament bundle 55 mum in length extends from the acrosomal vacuole membrane through a canal in the nucleus and then coils in a regular fashion around the base of the nucleus. The bundle expands systematically from 15 filaments near the acrosomal vacuole to 85 filaments at the basal end. Thin sections of sperm fixed during stages in spermatid maturation reveal that the filament bundle begins to assemble on dense material attached to the acrosomal vacuole membrane. In micrographs fo these early stages in maturation, short bundles are seen extending posteriorly from the dense material. The significance is that these short, developing bundles have about 85 filaments, suggesting that the 85-filament end of the bundle is assembled first. By using filament bundles isolated and incubated in vitro with G actin from muscle, we can determine the end "preferred" for addition of actin monomers during polymerization. The end that would be associated with the acrosomal vacuole membrane, a membrane destined to be continuous with the plasma membrane, is preferred about 10 times over the other, thicker end. Decoration of the newly polymerized portions of the filament bundle with subfragment 1 of myosin reveals that the arrowheads point away from the acrosomal vacuole membrane, as is true of other actin filament bundles attached to membranes. From these observations we conclude that the bundle is nucleated from the dense material associated with the acrosomal vacuole and that monomers are added to the membrane-associated end. As monomers are added at the dense material, the thick first-made end of the filament bundle is pushed down through the nucleus where, upon reaching the base of the nucleus, it coils up. Tapering is brought about by the capping of the peripheral filaments in the bundle.

scholarly journals Fodrin and band 4.1 in a plasma membrane-associated fraction of human neutrophils

Blood ◽  
1989 ◽  
Vol 74 (6) ◽  
pp. 2136-2143
Author(s):  
KB Stevenson ◽  
RA Clark ◽  
WM Nauseef
Keyword(s):  
Plasma Membrane ◽  
Actin Filaments ◽  
Polymorphonuclear Leukocytes ◽  
Membrane Vesicles ◽  
Human Neutrophils ◽  
Plasma Membrane Vesicles ◽  
Associated Proteins ◽  
Human Pmns ◽  
Discontinuous Percoll Gradient ◽  
Human Polymorphonuclear Leukocytes

Erythrocytes possess a well-characterized submembranous filamentous network which interacts with transmembrane glycoproteins and is composed primarily of spectrin, ankyrin, band 4.1, and short actin filaments. An analogous structure was recently described in platelets. Human polymorphonuclear leukocytes (PMNs) were examined for the presence and plasma membrane association of similar proteins. Isolated PMNs, free of contamination with erythrocytes or platelets, were disrupted by nitrogen cavitation and separated into subcellular organelles on a discontinuous Percoll gradient. Detergent lysates of plasma membrane vesicles, but not azurophilic or specific granules, contained insoluble actin filaments and associated proteins. Immunoblots of detergent-insoluble plasma membrane fractions contained proteins recognized by antibodies to brain fodrin and erythrocyte band 4.1, whereas blots probed with antibodies to erythrocyte spectrin and ankyrin were negative. Fodrin and band 4.1 were not detected in granule fractions, but some fodrin was present in the cytosol. The association of proteins related to fodrin and band 4.1 with the plasma membrane suggests that PMNs contain a submembranous skeleton structurally analogous to that of erythrocytes and platelets. The specific function of these proteins and their structural organization in human PMNs await further study.

Ultrastructure and gold-immunolabelling of cell-substratum adhesions (podosomes) in RSV-transformed BHK cells

1989 ◽  
Vol 94 (1) ◽  
pp. 85-99
Author(s):  
I. Gavazzi ◽  
M.V. Nermut ◽  
P.C. Marchisio
Keyword(s):  
Plasma Membrane ◽  
Actin Filaments ◽  
Rous Sarcoma Virus ◽  
Ultrastructural Level ◽  
Bhk Cells ◽  
Rous Sarcoma ◽  
Adhesion Sites ◽  
Associated Proteins ◽  
Sarcoma Virus ◽  
Dense Aggregation

Rous sarcoma virus-transformed BHK (RSV/B4-BHK) cells develop peculiar dot-like adhesions, that have been named podosomes, which, in the presence of serum, aggregate into ring- or crescent-shaped adhesion sites, the rosettes of podosomes. We have used the lysis-squirting technique and gold-immunolabelling to study the 3D-organisation of podosomes and the location of vinculin, gelsolin, phosphotyrosine-containing proteins and pp60src at an ultrastructural level. Podosomes appear to be conical bodies, 0.1-0.5 micron high, made by a dense aggregation of actin oligomers and several associated proteins, connected in the rosette by actin filaments. Gelsolin and some phosphotyrosine-containing proteins are found within the podosomes, often associated with the actin filaments, while vinculin is found predominantly at the podosome periphery, associated with microfilaments, and pp60src is located on the adjacent plasma membrane.

Deformation of Yeast Plasma Membrane by Ethidium Bromide

1988 ◽  
Vol 46 ◽  
pp. 254-255
Author(s):  
E. Keyhani
Keyword(s):  
Plasma Membrane ◽  
Thin Section ◽  
Ethidium Bromide ◽  
Freeze Fracture ◽  
Mutagenic Effect ◽  
Yeast Cells ◽  
Hydrophobic Region ◽  
Thin Section Electron Microscopy ◽  
Section Electron ◽  
Deep Pits

The mutagenic effect of ethidium bromide on the mitochondrial DNA is well established. Using thin section electron microscopy, it was shown that when yeast cells were grown in the presence of ethidium bromide, besides alterations in the mitochondria, the plasma membrane also showed alterations consisting of 75 to 110 nm-deep pits. Furthermore, ethidium bromide induced an increase in the length and number of endoplasmic reticulum and in the number of intracytoplasmic vesicles.Freeze-fracture, by splitting the hydrophobic region of the membrane, allows the visualization of the surface view of the membrane, and consequently, any alteration induced by ethidium bromide on the membrane can be better examined by this method than by the thin section method.Yeast cells, Candida utilis. were grown in the presence of 35 μM ethidium bromide. Cells were harvested and freeze-fractured according to the procedure previously described.

Exterior and Interior Events in Early Stage of Thrombin-induced Platelet Aggregation

1977 ◽  
Vol 38 (03) ◽  
pp. 0630-0639 ◽  
Author(s):  
Shuichi Hashimoto ◽  
Sachiko Shibata ◽  
Bonro Kobayashi
Keyword(s):  
Molecular Weight ◽  
Enzyme Activity ◽  
Plasma Membrane ◽  
Platelet Aggregation ◽  
Early Stage ◽  
Adenyl Cyclase ◽  
Cellular Component ◽  
Primary Event ◽  
Rabbit Platelets ◽  
Membrane Bound

SummaryTreatment of washed rabbit platelets with 1 u/ml of thrombin at 37° C resulted in a disappearance from platelets of a protein with 250,000 dalton molecular weight which was shown to be originated from plasma membrane. Parallel loss of adenyl cyclase was noted, and both reactions were complete within 30 sec. From the patterns of disc electrophoretograms, the importance of quick suppression of thrombin action in demonstrating the primary event was stressed.Thrombin induced an apparent activation of membrane bound phosphodiesterase. This reaction was also complete within 30 sec. The cellular component which contained the enzyme activity was distinct from plasma membrane. Soluble phosphodiesterase was not influenced by thrombin at all.These reactions required intact platelet cells to react with thrombin, and no reaction was detected when subcellular preparations were treated with thrombin.Possibility of collaboration of changes in externally located synthetic enzyme with those in internally located degrading enzyme in the early phase of thrombin action on platelets was suggested.

Biotinylation and Purification of Plasma Membrane-associated Proteins from Rodent Cultured Neurons

BIO-PROTOCOL ◽  
2016 ◽  
Vol 6 (10) ◽  
Author(s):  
Margarida Caldeira ◽  
Joana Ferreira ◽  
Ana Carvalho ◽  
Carlos Duarte
Keyword(s):  
Plasma Membrane ◽  
Cultured Neurons ◽  
Associated Proteins
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