Rapid Purification of Tubulin from Tissue and Tissue Culture Cells Using Solid-Phase Ion Exchange

The phosphorylation-dependent anchorage of retinoblastoma protein Rb in the nucleus is essential for its function. We show that its pocket C domain is both necessary and sufficient for nuclear anchorage by transiently expressing green fluorescent protein (GFP) chimeras of Rb fragments in tissue culture cells and by extracting the cells with hypotonic solutions. Solid phase binding assays using glutathioneS-transferase-fusion of Rb pockets A, B, and C revealed a direct association of lamin C exclusively to pocket C. Lamina-associated polypeptide (LAP) 2α, a binding partner of lamins A/C, bound strongly to pocket C and weakly to pocket B. When LAP2α was immunoprecipitated from soluble nuclear fractions, lamins A/C and hypophosphorylated Rb were coprecipitated efficiently. Similarly, immunoprecipitation of expressed GFP-Rb fragments by using anti-GFP antibodies coprecipitated LAP2α, provided that pocket C was present in the GFP chimeras. On redistribution of endogenous lamin A/C and LAP2α into nuclear aggregates by overexpressing dominant negative lamin mutants in tissue culture cells, Rb was also sequestered into these aggregates. In primary skin fibroblasts, LAP2α is expressed in a growth-dependent manner. Anchorage of hypophosphorylated Rb in the nucleus was weakened significantly in the absence of LAP2α. Together, these data suggest that hypophosphorylated Rb is anchored in the nucleus by the interaction of pocket C with LAP2α–lamin A/C complexes.

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Cytoplasmic Tubules in Cells Infected with Laryngotracheitis Virus

Proceedings, annual meeting, Electron Microscopy Society of America ◽

10.1017/s0424820100063792 ◽

1969 ◽

Vol 27 ◽

pp. 376-377

Author(s):

A. M. Watrach

Keyword(s):

Tissue Culture ◽

Small Proportion ◽

Chicken Embryo ◽

Culture Cells ◽

Tissue Culture Cells ◽

Morphologic Characteristics ◽

Infectious Laryngotracheitis ◽

Laryngotracheitis Virus ◽

In Cells

During a study of the development of infectious laryngotracheitis (LT) virus in tissue culture cells, unusual tubular formations were found in the cytoplasm of a small proportion of the affected cells. It is the purpose of this report to describe the morphologic characteristics of the tubules and to discuss their possible association with the development of virus.The source and maintenance of the strain of LT virus have been described. Prior to this study, the virus was passed several times in chicken embryo kidney (CEK) tissue culture cells.

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Sulfhydryl bond formation is a prerequisite for proper cycling of centrosomes and chromosomes in mammalian tissue culture cells

Proceedings, annual meeting, Electron Microscopy Society of America ◽

10.1017/s0424820100147557 ◽

1993 ◽

Vol 51 ◽

pp. 342-343

Author(s):

Heide Schatten ◽

Neidhard Paweletz ◽

Ron Balczon

Keyword(s):

Tissue Culture ◽

Cell Cycle Progression ◽

Group Formation ◽

Sulfhydryl Group ◽

Mammalian Tissue ◽

Mitotic Chromosomes ◽

Microtubule Network ◽

Culture Cells ◽

Tissue Culture Cells ◽

Transmission Electron

To study the role of sulfhydryl group formation during cell cycle progression, mammalian tissue culture cells (PTK2) were exposed to 100¼M 2-mercaptoethanol for 2 to 6 h during their exponential phase of growth. The effects of 2-mercaptoethanol on centrosomes, chromosomes, microtubules, membranes and intermediate filaments were analyzed by transmission electron microscopy (TEM) and by immunofluorescence microscopy (IFM) methods using a human autoimmune antibody directed against centrosomes (SPJ), and a mouse monoclonal antibody directed against tubulin (E7). Chromosomes were affected most by this treatment: premature chromosome condensation was detected in interphase nuclei, and the structure in mitotic chromosomes was altered compared to control cells. This would support previous findings in dividing sea urchin cells in which chromosomes are arrested at metaphase while the centrosome splitting cycle continues. It might also support findings that certairt-sulfhydryl-blocking agents block cyclin destruction. The organization of the microtubule network was scattered probably due to a looser organization of centrosomal material at the interphase centers and at the mitotic poles.

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Trypsin action on the growth of Sendai virus in tissue culture cells. IV. Evidence for activation of sendai virus by cleavage of a glycoprotein.

Journal of Virology ◽

10.1128/jvi.18.3.1147-1150.1976 ◽

1976 ◽

Vol 18 (3) ◽

pp. 1147-1150 ◽

Cited By ~ 21

Author(s):

M Ohuchi ◽

M Homma

Keyword(s):

Tissue Culture ◽

Sendai Virus ◽

Culture Cells ◽

Tissue Culture Cells

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UDP-D-glucose 4-epimerase activity of the tissue culture of white poplars (Populus alba L. var. pyramidalis)

Collection of Czechoslovak Chemical Communications ◽

10.1135/cccc19821530 ◽

1982 ◽

Vol 47 (5) ◽

pp. 1530-1536 ◽

Cited By ~ 1

Author(s):

Ladislav Bilisics ◽

Štefan Karácsonyi ◽

Marta Kubačková

Keyword(s):

Tissue Culture ◽

Cell Walls ◽

Enzyme Preparation ◽

Uridine Diphosphate ◽

Populus Alba ◽

Activity Change ◽

White Poplar ◽

Culture Cells ◽

Tissue Culture Cells ◽

Galactose Content

The presence of UDP-D-glucose 4-epimerase (EC 5.1.3.2) in the culture tissue of white poplar was evidenced. As found, the partially purified enzyme preparation contained UDP-D-glucose glucosyltransferase, UDP-D-galactose galactosyltransferase and non-specific enzymes able to cleave the uridine-diphosphate saccharides into the appropriate hexose monophosphates. The activity change of UDP-D-glucose 4-epimerase in tissue culture cells during the growth was in accord with changes in D-galactose content in cell walls and indicated the possibility to regulate the formation of polysaccharides containing D-galactose at the level of production of UDP-D-galactose in cells.

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