scholarly journals Fimbrin, a new microfilament-associated protein present in microvilli and other cell surface structures.

1980 ◽  
Vol 86 (1) ◽  
pp. 335-340 ◽  
Author(s):  
A Bretscher ◽  
K Weber
Keyword(s):  
Cell Surface ◽  
Indirect Immunofluorescence ◽  
Immunofluorescence Microscopy ◽  
Cultured Cells ◽  
Intestinal Epithelial Cells ◽  
Surface Structures ◽  
Intestinal Epithelial ◽  
Indirect Immunofluorescence Microscopy ◽  
Cell Surface Structures ◽  
Microfilament Bundles

A 68,000 mol wt polypeptide has been identified as one of the few major proteins in the microfilament bundles of the microvilli present on intestinal epithelial cells. Antibodies against the purified protein have been used in indirect immunofluorescence microscopy on several cultured cells. The protein have been used in indirect immunofluorescence microscopy on several cultured cells. The protein is found particularly prominent in membrane ruffles, microspikes, and microvilli.

scholarly journals Localization of actin and microfilament-associated proteins in the microvilli and terminal web of the intestinal brush border by immunofluorescence microscopy.

1978 ◽  
Vol 79 (3) ◽  
pp. 839-845 ◽  
Author(s):  
A Bretscher ◽  
K Weber
Keyword(s):  
Epithelial Cells ◽  
Indirect Immunofluorescence ◽  
Brush Border ◽  
Immunofluorescence Microscopy ◽  
Intestinal Epithelial Cells ◽  
Intestinal Epithelial ◽  
Indirect Immunofluorescence Microscopy ◽  
Terminal Web ◽  
Intestinal Brush Border ◽  
Associated Proteins

Indirect immunofluorescence microscopy was used to localize microfilament-associated proteins in the brush border of mouse intestinal epithelial cells. As expected, antibodies to actin decorated the microfilaments of the microvilli, giving rise to a very intense fluorescence. By contrast, antibodies to myosin, tropomyosin, filamin, and alpha-actinin did not decorate the microvilli. All these antibodies, however, decorated the terminal web region of the brush border. Myosin, tropomyosin, and alpha-actinin, although present throughout the terminal web, were found to be preferentially located around the periphery of the organelle. Therefore, two classes of microfilamentous structures can be documented in the brush border. First, the highly ordered microfilaments which make up the cores of the microvilli apparently lack the associated proteins. Second, seemingly less-ordered microfilaments are found in the terminal web, in which region the myosin, tropomyosin, filamin and alpha-actinin are located.

scholarly journals Purification of an 80,000-dalton protein that is a component of the isolated microvillus cytoskeleton, and its localization in nonmuscle cells.

1983 ◽  
Vol 97 (2) ◽  
pp. 425-432 ◽  
Author(s):  
A Bretscher
Keyword(s):  
Indirect Immunofluorescence ◽  
Immunofluorescence Microscopy ◽  
Cultured Cells ◽  
Major Protein ◽  
Intestinal Epithelial ◽  
Indirect Immunofluorescence Microscopy ◽  
Core Proteins ◽  
Nonmuscle Cells ◽  
Protein Components

The microvillus cytoskeleton, isolated from chicken intestinal epithelial cell brush borders, is known to contain five major protein components, the 110,000-dalton polypeptide, villin (95,000 daltons), fimbrin (68,000 daltons), actin (43,000 daltons), and calmodulin (17,000 daltons). In this paper we describe our first step in studying the minor components of the isolated core. We have so far identified and purified an 80,000-dalton polypeptide that was present in the isolated structure in approximately 0.7% the molar abundance of actin. Antibodies to the 80,000-dalton component did not react with other microvillus core proteins, and, when used in indirect immunofluorescence microscopy, they stained the microvilli of intestinal epithelial cells fixed in situ. The 80,000-dalton component therefore appears to be a newly-identified, authentic component of intestinal microvilli in vivo and of isolated microvillus cores. Immunological studies demonstrate that the 80,000-dalton component is widely distributed in nonmuscle cells. Indirect immunofluorescence microscopy reveals that it is particularly enriched in surface structures, such as blebs, microvilli, and retraction fibers of cultured cells.

Syncytioskeletons in choriocarcinoma in culture

1984 ◽  
Vol 66 (1) ◽  
pp. 1-20
Author(s):  
C.D. Ockleford ◽  
L. Dearden ◽  
R.A. Badley
Keyword(s):  
Indirect Immunofluorescence ◽  
Immunofluorescence Microscopy ◽  
Cultured Cells ◽  
Cortical Layers ◽  
Dense Population ◽  
Bewo Cells ◽  
Indirect Immunofluorescence Microscopy ◽  
Microtubule Organizing Centres ◽  
Transmission Electron ◽  
Cytoskeletal Elements

Indirect immunofluorescence microscopy using anti-actin serum has been used to investigate the distribution of actin-containing polymers in BeWo cells. This cell line, derived from a human choriocarcinoma, contains tissue that, like its tissue of origin, is partly syncytial. The syncytial nature has been inferred from study of Nomarski optical sections and from transmission electron microscopy. The multinucleated plaques of tissue possess a syncytioskeleton with a number of actin-containing features characteristic of cultured cells. These include stress fibres, cortical layers and ruffled membranes. Other actin-containing structures are more typical of the related non-pathological syncytiotrophoblast. These include a dense population of microvilli. The overall organization of the actin syncytioskeletons bears no obvious relationship to the number or position of nuclei in the syncytium. Indirect immunofluorescence microscopy has also been employed to localize the protein tubulin in BeWo cells. The microtubules do not appear to be spatially organized by a particular nucleus. Rather, there are numerous microtubule-organizing centres (MTOCs) that exist in the cytoplasm and do not have the expected numerical and positional relationship to nuclei. From these data it appears that polymeric cytoskeletal elements in these syncytia are organized in a manner not immediately subordinate to syncytial nuclei.

Cell-to-substratum contacts in living cells: a direct correlation between interference-reflexion and indirect-immunofluorescence microscopy using antibodies against actin and alpha-actinin

1979 ◽  
Vol 37 (1) ◽  
pp. 257-273
Author(s):  
J. Wehland ◽  
M. Osborn ◽  
K. Weber
Keyword(s):  
Indirect Immunofluorescence ◽  
Immunofluorescence Microscopy ◽  
Living Cells ◽  
Mammary Cells ◽  
Indirect Immunofluorescence Microscopy ◽  
Focal Contacts ◽  
Microfilament Bundles

Rat mammary cells growing on glass coverslips were photographed first using interference-reflexion microscopy and then after processing for indirect-immunofluorescence microscopy with antibodies to actin or to alpha-actinin. A comparison of the images of the same cell given by the 2 microscopical procedures indicates that the focal contacts between the cell and the substratum correspond to distal ends of microfilament bundles, and the these bundles are only in limited areas close to the substratum. The focal contracts are rich in alpha-actinin which has been proposed as a membrane-anchorage protein for microfilament bundles. Use of stereo immunofluorescence microscopy allows a direct comparison between the interference-reflexion image, and the underside of the cell after staining with antibodies to actin or alpha-actinin.

scholarly journals Binding of deoxyribonuclease I to actin: a new way to visualize microfilament bundles in nonmuscle cells.

1978 ◽  
Vol 26 (9) ◽  
pp. 745-749 ◽  
Author(s):  
E Wang ◽  
A R Goldberg
Keyword(s):  
Indirect Immunofluorescence ◽  
Immunofluorescence Microscopy ◽  
Staining Pattern ◽  
Fluorescent Microscopy ◽  
Dnase I ◽  
Deoxyribonuclease I ◽  
Indirect Immunofluorescence Microscopy ◽  
Microfilament Bundles ◽  
Nonmuscle Cells ◽  
In Cells

Intracellular actin-containing fibers can be visualized by indirect immunofluorescence microscopy when they are stained with antibody directed against DNase I. The location of actin-containing fibers in cells appears to be similar to the staining pattern of antibody to actin. Actin fibers were also visualized by direct fluorescent microscopy with rhodamine-conjugated DNase I.

Subcellular localization of Trypanosoma cruzi glycoprotein Gp72

1996 ◽  
Vol 109 (13) ◽  
pp. 2979-2988 ◽  
Author(s):  
P.A. Haynes ◽  
D.G. Russell ◽  
G.A. Cross
Keyword(s):  
Cell Surface ◽  
Trypanosoma Cruzi ◽  
Cell Body ◽  
Indirect Immunofluorescence ◽  
Immunofluorescence Microscopy ◽  
Subcellular Location ◽  
Surface Glycoprotein ◽  
Null Mutant ◽  
Western Blots ◽  
Indirect Immunofluorescence Microscopy

We have investigated the subcellular location of the Trypanosoma cruzi surface glycoprotein, Gp72, by introducing epitope-tagged copies of gp72 null-mutant cells. A tagged Gp72, containing three tandemly repeated copies of a human influenza hemagglutinin nonapeptide (HA) adjacent to the mature Gp72 amino terminus, was able to complement the null mutant phenotype, as well as being recognized in Western blots by both anti-HA antibody and the carbohydrate-specific monoclonal antibody WIC29.26. Integration of this epitope-tagged gp72 into the chromosomal gp72 locus produced a clonal cell line, 72HAN3.1G7, which was used for studies of the subcellular location of the epitope-tagged Gp72. Indirect immunofluorescence microscopy of fixed 72HAN3.1G7 epimastigotes showed that GP72 was evenly distributed over the cell body and somewhat concentrated in the proximal region of the flagellum. No fluorescence could be detected in the distal tip of the flagellum. Immunoelectron microscopy of fixed 72HAN3.1G7 epimastigotes revealed that Gp72 was predominantly membrane-associated and located on the cell surface. Indirect immunofluorescence microscopy of live 72HAN3.1G7 epimastigote cells showed a similar pattern of fluorescence on the flagellum, but no fluorescence was detected on the cell body, which was attributed to masking by other cell-surface components. Indirect immunofluorescence microscopy of fixed 72HAN3.1G7 amastigotes revealed that Gp72, which has long been considered to be expressed only in epimastigotes and metacyclic trypomastigotes, can be expressed in amastigotes, but it no longer contains the WIC29.26 carbohydrate epitope.

scholarly journals Moesin, like ezrin, colocalizes with actin in the cortical cytoskeleton in cultured cells, but its expression is more variable

1993 ◽  
Vol 105 (1) ◽  
pp. 219-231 ◽  
Author(s):  
Z. Franck ◽  
R. Gary ◽  
A. Bretscher
Keyword(s):  
Cell Surface ◽  
Cultured Cells ◽  
Adherens Junctions ◽  
Membrane Binding ◽  
Binding Domain ◽  
Terminal Region ◽  
Surface Structures ◽  
Amino Terminal ◽  
Sequence Identity ◽  
Cell Surface Structures

The band 4.1 superfamily of proteins show approx. 30% sequence identity in their amino-terminal region to the membrane binding domain of erythrocyte band 4.1. Within this superfamily are three members, ezrin, radixin and moesin, that show approx. 75% overall sequence identity. A comparison of the domain structure and intracellular localization of ezrin and moesin in cultured cells is reported here. Limited proteolytic digestion of ezrin or moesin yields a relatively stable 32 kDa domain derived from the amino-terminal region that is homologous to the protease-resistant membrane binding domain of erythrocyte band 4.1. The remaining regions of the two proteins give rise to very different fragments, suggesting that the secondary/tertiary structures of the two proteins are different in these regions. We have generated polyclonal antibodies that discriminate between ezrin and moesin, and do not react with radixin. All cultured cell lines investigated contain ezrin, whereas moesin is variably expressed. Cells that contain both ezrin and moesin show a very similar pattern: both proteins are enriched and colocalize with actin in cell surface structures. Ezrin is also detected in the cytoplasm. In cells with few or no surface structures, both proteins show a patchy distribution in regions of the cell that contain fine networks of actin filaments. No staining of focal contacts or adherens junctions was observed. These results, together with those of others, lead to the conclusion that, of the members of this protein family, only radixin is an authentic component of adherens junctions and focal contacts. Ezrin and moesin are both found in cell surface structures after treatment of human A431 cells with epidermal growth factor, and ezrin, but not moesin, becomes phosphorylated on tyrosine. This study shows that ezrin and moesin have a similar subcellular distribution in cultured cells, yet are distinguishable in their expression, structure and ability to serve as a kinase substrate.

The Role of Cytokines in the Modulation of Cell Surface Antigens of Human Melanoma

1993 ◽  
Vol 8 (3) ◽  
pp. 151-154 ◽  
Author(s):  
A. Anichini ◽  
R. Mortarini ◽  
G. Parmiani
Keyword(s):  
Cell Surface ◽  
Surface Antigens ◽  
Surface Structures ◽  
Biological Behavior ◽  
Cell Surface Antigens ◽  
Membrane Expression ◽  
Neoplastic Cells ◽  
Antigenic Modulation ◽  
Human Malignant Melanoma ◽  
Cell Surface Structures

A number of different cytokines, including IL-1α. and ß, IL-2, IL-3, IL-4, IL-6, IL-7, IL-8, IFN-α, -ß and γ, TNF-α -ß, and TGF-ß1, can modulate the expression of distinct cell surface antigens of normal and neoplastic cells. Both induction/increase of expression and reduction of expression can be achieved depending on the antigen and on the cytokine. Antigens subjected to the modulating activity of cytokines include distinct families of cell surface structures such as the molecules coded by the major histocompatibility complex (MHC), the superfamily of adhesion receptors that regulate cell-cell and cell-matrix interaction, receptors for cytokines and growth factors and tumor-associated antigens. The modulating activity of cytokines is a consequence of their influence on gene expression, protein synthesis, membrane expression and shedding of antigens from the cell surface. The changes of phenotype due to the action of cytokines can influence the signalling pathways dependent on the expression and function of cell surf ace structures. Therefore, the antigen modulating activity of cytokines can thoroughly affect the biological behavior of normal and neoplastic cells. As described here, most of the modulating effects of cytokines on different cell surface structures and the functional consequences of antigenic modulation can be verified in human malignant melanoma cells.

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