scholarly journals The Organization and Animal–Vegetal Asymmetry of Cytokeratin Filaments in Stage VIXenopusOocytes Is Dependent upon F-Actin and Microtubules

1997 ◽  
Vol 184 (1) ◽  
pp. 95-114 ◽  
Author(s):  
David L. Gard ◽  
Byeong Jik Cha ◽  
Edward King
Keyword(s):  
Cytokeratin Filaments

A monoclonal antibody against mouse oocyte cytoskeleton recognizing cytokeratin-type filaments

Development ◽  
1985 ◽  
Vol 90 (1) ◽  
pp. 197-209
Author(s):  
E. Lehtonen
Keyword(s):  
Epithelial Cells ◽  
Immunofluorescence Microscopy ◽  
Staining Pattern ◽  
Blastocyst Stage ◽  
Tissue Sections ◽  
Mouse Oocytes ◽  
Thick Filaments ◽  
Trophectoderm Cells ◽  
Early Embryos ◽  
Cytokeratin Filaments

Monoclonal antibodies were raised to detergent-extracted cytoskeleton preparations of mouse oocytes. In immunofluorescence microscopy, one of the antibodies, OCS-1, localizes exclusively to epithelial cells in frozen tissue sections, including various simple and stratified epithelia. The antibody decorates a keratin-type of fibrillax, vinblastine-resistant network in various cultured, epithelial-type cells, but not in myoid or fibroblastoid cells. In mouse oocytes and cleavage-stage embryos, the OCS-1 antibody gives a diffuse, spotty staining pattern. In blastocyst-stage embryos, the antibody reveals a keratin-type filamentous organization in the trophectoderm cells. In immunoelectron microscopy, the OCS-1 antibody decorates 10 nm-thick filaments, often identifiable as desmosome-attached tonofilaments, in detergent-treated trophectoderm cells. The antigen(s) recognized by the OCS-1 antibody is apparently present in, or closely associated with, cytokeratin filaments. In addition to mouse oocytes and early embryos, a wide variety of epithelial cells in various species seem to share this antigen(s). The present results suggest that at the early stages, the cytokeratin-related antigen(s) defined by the OCS-1 antibody are stored in a non-fibrillar form which is then converted into a fibrillar network at the blastocyst stage. A pre-existing supply of cytokeratin-related protein may be essential for the development of the blastocyst.

Desmosomes: interconnected calcium-dependent structures of remarkable stability with significant integral membrane protein turnover

2002 ◽  
Vol 115 (8) ◽  
pp. 1717-1732 ◽  
Author(s):  
Reinhard Windoffer ◽  
Monika Borchert-Stuhlträger ◽  
Rudolf E. Leube
Keyword(s):  
Plasma Membrane ◽  
Fluorescence Microscopy ◽  
Structural Integrity ◽  
Time Lapse ◽  
Fluorescence Recovery ◽  
Adhesion Sites ◽  
Time Space ◽  
Molecular Stability ◽  
Filament Bundles ◽  
Cytokeratin Filaments

Desmosomes are prominent cell adhesion structures that are major stabilizing elements, together with the attached cytoskeletal intermediate filament network, of the cytokeratin type in epithelial tissues. To examine desmosome dynamics in tightly coupled cells and in situations of decreased adhesion, fluorescent desmosomal cadherin desmocollin 2a (Dsc2a) chimeras were stably expressed in human hepatocellular carcinoma-derived PLC cells (clone PDc-13) and in Madin-Darby canine kidney cells (clone MDc-2) for the continuous monitoring of desmosomes in living cells. The hybrid polypeptides integrated specifically and without disturbance into normal-appearing desmosomes that occurred in association with typical cytokeratin filament bundles. Tracking of labeled adhesion sites throughout the cell cycle by time-lapse fluorescence microscopy revealed that they were immobile and that they maintained their structural integrity for long periods of time. Time-space diagrams further showed that desmosomal positioning was tightly controlled, even during pronounced cell shape changes, although the desmosomal arrays extended and contracted, suggesting that they were interconnected by a flexible system with intrinsic elasticity. Double-fluorescence microscopy detecting Dsc2a chimeras together with fluorescent cytokeratin 18 chimeras revealed the association and synchronous movement of labeled desmosomes and fluorescent cytokeratin filaments. Only a minor destabilization of desmosomes was observed during mitosis, demonstrated by increased diffuse plasma membrane fluorescence and the fusion of desmosomes into larger structures. Desmosomes did not disappear completely at any time in any cell, and residual cytokeratin filaments remained in association with adhesion sites throughout cell division. On the other hand, a rapid loss of desmosomes was observed upon calcium depletion, with irreversible uptake of some desmosomal particles. Simultaneously, diffusely distributed desmosomal cadherins were detected in the plasma membrane that retained the competence to nucleate the reformation of desmosomes after the cells were returned to a standard calcium-containing medium. To examine the molecular stability of desmosomes, exchange rates of fluorescent chimeras were determined by fluorescence recovery after photobleaching, thereby identifying considerable Dsc2a turnover with different rates of fluorescence recovery for PDc-13 cells (36±17% recovery after 30 minutes) and MDc-2 cells (60±20% recovery after 30 minutes). Taken together, our observations suggest that desmosomes are pliable structures capable of fine adjustment to functional demands despite their overall structural stability and relative immobility.

Selective rearrangement of cytokeratin filaments in cultured liver epithelial cells induced by nickel

1987 ◽  
Vol 5 (3) ◽  
pp. 344-354 ◽  
Author(s):  
Y. Katsuma ◽  
S.H.H. Swierenga ◽  
N. Marceau ◽  
S.W. French
Keyword(s):  
Epithelial Cells ◽  
Liver Epithelial Cells ◽  
Cytokeratin Filaments

scholarly journals Change of cytokeratin filament organization during the cell cycle: selective masking of an immunologic determinant in interphase PtK2 cells.

1983 ◽  
Vol 97 (4) ◽  
pp. 1255-1260 ◽  
Author(s):  
W W Franke ◽  
E Schmid ◽  
J Wellsteed ◽  
C Grund ◽  
O Gigi ◽  
...  
Keyword(s):  
Cell Cycle ◽  
Monoclonal Antibody ◽  
Cell Cultures ◽  
Immunofluorescence Microscopy ◽  
Specific Reaction ◽  
Low Concentrations ◽  
Interphase Cells ◽  
Triton X ◽  
Cytokeratin Filaments ◽  
Striking Contrast

The organization of intermediate-sized filaments (IF) of the cytokeratin type was studied in cultures of PtK2 cells in which typical IF structures are maintained during mitosis, using a monoclonal antibody (KG 8.13). This antibody reacts, in immunoblotting experiments, with the larger of the two major cytokeratin polypeptides present in these cells but, using standard immunofluorescence microscopy procedures, does not react with the cytokeratin filaments abundant in interphase cells, in striking contrast to various antisera and other monoclonal cytokeratin antibodies. In the same cell cultures, however, the antibody does react with cytokeratin filaments of mitotic and early postmitotic cells. The specific reaction with cytokeratin filaments of mitotic cells only is due to the exposure of the specific immunologic determinant in mitosis and its masking in interphase cells. Treatment of interphase cells with both Triton X-100 as well as with methanol and acetone alters the cytokeratin filaments and allows them to react with this monoclonal antibody. A similar unmasking was noted after treatment with buffer containing 2 M urea or low concentrations of trypsin. We conclude that the organization of cytokeratin, albeit still arranged in typical IF, is altered during mitosis of PtK2 cells.

scholarly journals Evidence for the involvement of microtubules, ER, and kinesin in the cortical rotation of fertilized frog eggs.

1991 ◽  
Vol 114 (5) ◽  
pp. 1017-1028 ◽  
Author(s):  
E Houliston ◽  
R P Elinson
Keyword(s):  
Cell Cycle ◽  
Plasma Membrane ◽  
Sea Urchin ◽  
Rana Pipiens ◽  
Cortical Microtubules ◽  
Cell Biol ◽  
Vegetal Cortex ◽  
Almost All ◽  
Cytoplasmic Side ◽  
Cytokeratin Filaments

During the first cell cycle, the vegetal cortex of the fertilized frog egg is translocated over the cytoplasm. This process of cortical rotation creates regional cytoplasmic differences important in later development, and appears to involve an array of aligned microtubules that forms transiently beneath the vegetal cortex. We have investigated how these microtubules might be involved in generating movement by analyzing isolated cortices and sections of Xenopus laevis and Rana pipiens eggs. First, the polarity of the cortical microtubules was determined using the "hook" assay. Almost all microtubules had their plus ends pointing in the direction of cortical rotation. Secondly, the association of microtubules with other cytoplasmic elements was examined. Immunofluorescence revealed that cytokeratin filaments coalign with the microtubules. The timing of their appearance and their position on the cytoplasmic side of the microtubules suggested that they are not involved directly in generating movement. ER was visualized with the dye DiIC16(3) and by immunofluorescence with anti-BiP (Bole, D. G., L. M. Hendershot, and J. F. Kearney, 1986. J. Cell Biol. 102:1558-1566). One layer of ER was found closely underlying the plasma membrane at all times. An additional, deeper layer formed in association with the microtubules of the array. Antibodies to sea urchin kinesin (Ingold, A. L., S. A. Cohn, and J. M. Scholey. 1988. J. Cell Biol. 107:2657-2667) detected antigens associated with both the ER and microtubules. On immunoblots they recognized microtubule associated polypeptide(s) of approximately 115 kD from Xenopus eggs. These observations are consistent with a role for kinesin in creating movement between the microtubules and ER, which leads in turn to the cortical rotation.

Cytokeratin filament assembly in the preimplantation mouse embryo

Development ◽  
1987 ◽  
Vol 101 (3) ◽  
pp. 565-582 ◽  
Author(s):  
J.C. Chisholm ◽  
E. Houliston
Keyword(s):  
Mouse Embryo ◽  
Inner Cell Mass ◽  
Cell Mass ◽  
Blastocyst Stage ◽  
Lineage Tracing ◽  
Cell Stage ◽  
Filament Assembly ◽  
Filament Networks ◽  
Tracing Techniques ◽  
Cytokeratin Filaments

The timing, spatial distribution and control of cytokeratin assembly during mouse early development has been studied using a monoclonal antibody, TROMA-1, which recognizes a 55,000 Mr trophectodermal cytokeratin (ENDO A). This protein was first detected in immunoblots at the 4-cell stage, and became more abundant at the 16-cell stage and later. Immunofluorescence analysis revealed assembled cytokeratin filaments in some 8-cell blastomeres, but not at earlier stages. At the 16-cell stage, filaments were found in both polarized (presumptive trophectoderm; TE) and apolar (presumptive inner cell mass; ICM) cells in similar proportions, although polarized cells possessed more filaments than apolar cells. By the late 32-cell, early blastocyst, stage, all polarized (TE) cells contained extensive filament networks whereas cells positioned inside the embryo tended to have lost their filaments. The presence of filaments in inside cells at the 16-cell stage and in ICM cells was confirmed by immunoelectron microscopy. Lineage tracing techniques demonstrated that those cells in the ICM of early blastocysts which did possess filaments were almost exclusively the progeny of polar 16-cell blastomeres, suggesting that these filaments were directly inherited from outside cells at the 16- to 32-cell transition. Inhibitor studies revealed that proximate protein synthesis but not mRNA synthesis is required for filament assembly at the 8-cell stage. These results demonstrate that there are quantitative rather than qualitative differences in the expression of cytokeratin filaments in the inner cell mass and trophectoderm cells of the mouse embryo.

Polar asymmetry in the organization of the cortical cytokeratin system of Xenopus laevis oocytes and embryos

Development ◽  
1987 ◽  
Vol 100 (3) ◽  
pp. 543-557 ◽  
Author(s):  
M.W. Klymkowsky ◽  
L.A. Maynell ◽  
A.G. Polson
Keyword(s):  
Xenopus Laevis ◽  
Mature Stage ◽  
Xenopus Laevis Oocytes ◽  
Blastula Stage ◽  
Early Embryos ◽  
Large Fibre ◽  
Filament Bundles ◽  
Punctate Pattern ◽  
Cortical Cytoskeleton ◽  
Cytokeratin Filaments

We have used whole-mount immunofluorescence microscopy of late-stage Xenopus laevis oocytes and early embryos to examine the organization of their cortical cytokeratin systems. In both mature oocytes and early embryos, there is a distinct animal-vegetal polarity in cytokeratin organization. In mature (stage-VI) oocytes, the cytokeratin filaments of the vegetal region form a unique, almost geodesic network; in the animal region, cytokeratin organization appears much more variable and irregular. In unfertilized, postgerminal vesicle breakdown eggs, the cortical cytokeratin system is disorganized throughout both animal and vegetal hemispheres. After fertilization, cytokeratin organization reappears first in a punctate pattern that is transformed into an array of oriented filaments. These cytokeratin filaments appear first in the vegetal hemisphere and are initially thin. Subsequently, they form bundles that grow thicker through the period of first to second cleavage, at which point large cytokeratin filament bundles form a loose, fishnet-like system that encompasses the vegetal portion of each blastomere. In the animal region, cytokeratin filaments do not appear to form large fibre networks, but rather appear to be organized into a system of fine filaments. The animal-vegetal polarity in cytokeratin organization persists until early blastula (stage 5); in later-stage embryos, both animal and vegetal blastomeres possess qualitatively similar cytokeratin filament systems. The entire process of cytokeratin reorganization in the egg is initiated by prick activation. These observations indicate that the cortical cytoskeleton of Xenopus oocytes and early embryos is both dynamic and asymmetric.

Differentiation in vitro of human-mouse teratocarcinoma hybrids

1983 ◽  
Vol 3 (12) ◽  
pp. 2259-2270
Author(s):  
F J Benham ◽  
M V Wiles ◽  
P N Goodfellow
Keyword(s):  
Retinoic Acid ◽  
Embryonal Carcinoma ◽  
Developmental Regulation ◽  
Genetic Material ◽  
Cell Types ◽  
Type Iv ◽  
Parietal Endoderm ◽  
Mouse Teratocarcinoma ◽  
Cytokeratin Filaments

The mouse embryonal carcinoma (EC) line, PCC4, was used to construct a series of somatic cell hybrids which contain a single or a few human chromosomes. The hybrids all retained the EC phenotype as determined by morphology, expression of SSEA-1, lack of cell surface H-2 antigen and cytokeratin filaments, high alkaline phosphatase levels, the ability to form EC tumors ectopically in nude mice, and the ability to differentiate in response to retinoic acid. Constitutively differentiated cloned lines were derived from retinoic acid-treated hybrid cultures. Several derived lines had a phenotype indistinguishable from that of parietal endoderm cells, which includes synthesis of large amounts of laminin, type IV procollagen, and plasminogen activator. One differentiated line showed a fibroblast-like morphology. The differentiated lines derived from two of the hybrids, MCP6 and GEOC4, stably maintained the sole human chromosomal component present in the EC progenitors. These EC hybrids therefore provide a system to study developmental regulation of the introduced and stably maintained human genetic material derived from a variety of cell types.

Disruption of the cytokeratin filament network in the preimplantation mouse embryo

Development ◽  
1988 ◽  
Vol 104 (2) ◽  
pp. 219-234
Author(s):  
J.A. Emerson
Keyword(s):  
Mouse Embryo ◽  
Blastocyst Stage ◽  
Cell Stage ◽  
Trophectoderm Cells ◽  
Preimplantation Mouse Embryos ◽  
Preimplantation Mouse Embryo ◽  
Preimplantation Mouse ◽  
Morula Stage ◽  
Cytokeratin Filaments

The distribution of the cytokeratin network in the intact preimplantation mouse embryo and the role of cytokeratin filaments in trophectoderm differentiation were investigated by means of whole-mount indirect immunofluorescence microscopy and microinjection of anti-cytokeratin antibody. Assembled cytokeratin filaments were detected in some blastomeres as early as the compacted 8-cell stage. The incidence and organization of cytokeratin filaments increased during the morula stage, although individual blastomeres varied in their content of assembled filaments. At the blastocyst stage, each trophectoderm cell contained an intricate network of cytokeratin filaments, and examination of sectioned blastocysts confirmed that extensive arrays of cytokeratin filaments were restricted to cells of the trophectoderm. Microinjection of anticytokeratin antibody into individual mural trophectoderm cells of expanded blastocysts resulted in a dramatic rearrangement of the cytokeratin network in these cells. Moreover, antibody injection into 2-cell embryos inhibited assembly of the cytokeratin network during the next two days of development. Despite this disruption of cytokeratin assembly, the injected embryos compacted and developed into blastocysts with normal morphology and nuclear numbers. These results suggest that formation of an elaborate cytokeratin network in preimplantation mouse embryos is unnecessary for the initial stages of trophectoderm differentiation resulting in blastocyst formation.

Antibodies to Cytokeratin Filaments in Patients with Alcoholic Liver Disease

1984 ◽  
Vol 8 (2) ◽  
pp. 212-215 ◽  
Author(s):  
Pekka Kurki ◽  
Ismo Virtanen ◽  
Veli-Pekka Lehto ◽  
Olof Atfthan ◽  
Mikko Salaspuro
Keyword(s):  
Liver Disease ◽  
Alcoholic Liver Disease ◽  
Cytokeratin Filaments
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