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  • 11
    Electronic Resource
    Electronic Resource
    The cenpB gene is not essential in mice (1998)
    Springer
    Chromosoma 107 (1998), S. 570-576 
    Details
    ISSN: 1432-0886
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Medicine
    Notes: Abstract.  Centromere protein B (CENP-B) is a centromeric DNA-binding protein that binds to α-satellite DNA at the 17 bp CENP-B box sequence. The binding of CENP-B, along with other proteins, to α-satellite DNA sequences at the centromere, is thought to package the DNA into heterochromatin subjacent to the kinetochore of mitotic chromosomes. To determine the importance of CENP-B to kinetochore assembly and function, we generated a mouse null for the cenpB gene. The deletion removed part of the promoter and the entire coding sequence except for the carboxyl-terminal 35 amino acids of the CENP-B polypeptide. Mice heterozygous or homozygous for the cenpB null mutation are viable and healthy, with no apparent defect in growth and morphology. We have established mouse embryo fibroblasts from heterozygous and homozygous cenpB null littermates. Microscopic analysis, using immunofluorescence and electron microscopy of the cultured cells, indicated that the centromere-kinetochore complex was intact and identical to control cells. Mitosis was identical in fibroblasts derived from cenpB wild-type, heterozygous and null animals. Our studies demonstrate that CENP-B is not required for the assembly of heterochromatin or the kinetochore, or for completion of mitosis.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1007/s004120050343
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  • 12
    Electronic Resource
    Electronic Resource
    The fine structure of the kinetochore of a mammalian cell in vitro (1966)
    Springer
    Chromosoma 19 (1966), S. 28-43 
    Details
    ISSN: 1432-0886
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Medicine
    Notes: Abstract The chromosomes of Chinese hamster cells were examined with the electron microscope and the following observations were made concerning the structure and organization of the kinetochore. — The kinetochore consists of a dense core 200–300 Å in diameter surrounded hy a less dense zone 200–600 Å wide. The dense core consists of a pair of axial fibrils 50–80 Å in diameter which may be coiled together in a cohelical manner. The less dense zone about the axial elements is composed of numerous microfibrils which loop out at right angles to the axial fibrils. Together the structures comprise a lampbrush-like filament which extends along the surface of each chromatid. Some sections suggested that two such filaments may be present on each chromatid. The fine structure of kinetochores associated with spindle filaments was essentially the same as those free of filaments. The structure and organization of the kinetochore of these mammalian cells was compared to that of lampbrush chromosomes of certain amphibian oöcytes, dipteran polytene chromosome puffs, and the synaptinemal complex seen during meiotic prophase.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1007/BF00332792
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  • 13
    Electronic Resource
    Electronic Resource
    Mitosis-specific phosphorylation of histone H3 initiates primarily within pericentromeric heterochromatin during G2 and spreads in an ordered fashion coincident with mitotic chromosome condensation (1997)
    Springer
    Chromosoma 106 (1997), S. 348-360 
    Details
    ISSN: 1432-0886
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Medicine
    Notes: Abstract. We have generated and characterized a novel site-specific antibody highly specific for the phosphorylated form of the amino-terminus of histone H3 (Ser10). In this study, we used this antibody to examine in detail the relationship between H3 phosphorylation and mitotic chromosome condensation in mammalian cells. Our results extend previous biochemical studies by demonstrating that mitotic phosphorylation of H3 initiates nonrandomly in pericentromeric heterochromatin in late G2 interphase cells. Following initiation, H3 phosphorylation appears to spread throughout the condensing chromatin and is complete in most cell lines just prior to the formation of prophase chromosomes, in which a phosphorylated, but nonmitotic, chromosomal organization is observed. In general, there is a precise spatial and temporal correlation between H3 phosphorylation and initial stages of chromatin condensation. Dephosphorylation of H3 begins in anaphase and is complete immediately prior to detectable chromosome decondensation in telophase cells. We propose that the singular phosphorylation of the amino-terminus of histone H3 may be involved in facilitating two key functions during mitosis: (1) regulate protein-protein interactions to promote binding of trans-acting factors that “drive” chromatin condensation as cells enter M-phase and (2) coordinate chromatin decondensation associated with M-phase.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1007/s004120050256
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  • 14
    Electronic Resource
    Electronic Resource
    Compound kinetochores of the Indian muntjac (1984)
    Springer
    Chromosoma 91 (1984), S. 1-11 
    Details
    ISSN: 1432-0886
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Medicine
    Notes: Abstract The chromosomes of the Indian muntjac (Muntiacus muntjak vaginalis) are unique among mammals due to their low diploid number (2N=6♀, 7♂) and large size. It has been proposed that the karyotype of this small Asiatic deer evolved from a related deer the Chinese muntjac (Muntiacus reevesi) with a diploid chromosome number of 2n= 46 consisting of small telocentric chromosomes. In this study we utilized a kinetochore-specific antiserum derived from human patients with the autoimmune disease scleroderma CREST as an immunofluorescent probe to examine kinetochores of the two muntjac species. Since CREST antiserum binds to kinetochores of mitotic chromosomes as well as prekinetochores in interphase nuclei, it was possible to identify and compare kinetochore morphology throughout the cell cycle. Our observations indicated that the kinetochores of the Indian muntjac are composed of a linear beadlike array of smaller subunits that become revealed during interphase. The kinetochores of the Chinese muntjac consisted of minute fluorescent dots located at the tips of the 46 telocentric chromosomes. During interphase, however, the kinetochores of the Chinese muntjac clustered into small aggregates reminiscent of the beadlike arrays seen in the Indian muntjac. Morphometric measurements of fluorescence indicated an equivalent amount of stained material in the two species. Our observations indicate that the kinetochores of the Indian muntjac are compound structures composed of linear arrays of smaller units the size of the individual kinetochores seen on metaphase chromosomes of the Chinese muntjac. Our study supports the notion that the kinetochores of the Indian muntjac evolved by linear fusion of unit kinetochores of the Chinese muntjac. Moreover, it is concluded that the evolution of compound kinetochores may have been facilitated by the nonrandom aggregation of interphase kinetochores in the nuclei of the ancestral species.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1007/BF00286479
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  • 15
    Electronic Resource
    Electronic Resource
    Arrangements of kinetochores in mouse cells during meiosis and spermiogenesis (1986)
    Springer
    Chromosoma 94 (1986), S. 309-317 
    Details
    ISSN: 1432-0886
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Medicine
    Notes: Abstract Antibodies from the serum of patients with the autoimmune disease scleroderma CREST were used to investigate the association and distribution of kinetochores in mouse cells during meiosis and spermiogenesis. The pattern of indirect immunofluorescent staining in pachytene nuclei indicated that each autosomal bivalent contains one fluorescent spot. Throughout pachytene, the kinetochores were arranged non-randomly into several clusters and distributed around the periphery of the nucleus. In subsequent stages of meiotic prophase I, distribution was random and the number of fluorescent spots increased from 21 to 40 corresponding to the diploid chromosome number and the number of halfbivalents oriented to the spindle poles at the metaphase I. Twenty pairs of kinetochores were observed at metaphase II. During spermiogenesis, the number of kinetochores correlated with the haploid chromosome number in early spermatids but tandem association of centromeres and clustering into a conspicuous chromocenter corresponded to a significant reduction in the number of fluorescent foci in mid-spermatid nuclei. The number of stained sites per nucleus continued to decrease during sperm maturation and total absence of staining was apparent in mature spermatozoa. Immunoblotting of proteins extracted from mature sperm however, indicated that a kinetochore antigen of Mr 80,000 was still present. Therefore, the absence of kinetochore staining in mature spermatozoa is probably due to the blockage of epitopes during chromatin condensation.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1007/BF00290861
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  • 16
    Electronic Resource
    Electronic Resource
    Localization of the centriole and keratin intermediate filaments in PtK1 cells by double immunofluorescence (1984)
    New York, NY : Wiley-Blackwell
    Cell Motility and the Cytoskeleton 4 (1984), S. 241-247 
    Details
    ISSN: 0886-1544
    Keywords: cytoskeleton ; centrosome ; tonofilaments ; Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: We present observations on the relative location of the centriole and keratin filament cap in motile PtK1 cells. Subconfluent cells were double labeled with anticentriole and antikeratin sera. These preparations revealed that the centriole is separate from, but neighboring, the keratin filament cap. Serial ultrathin sections confirm this observation. These observations are consistent with the idea that the microtubule organizing center and intermediate filament distribution center are not identical or concentric in PtK1 cells.
    Additional Material: 3 Ill.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1002/cm.970040403
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  • 17
    Electronic Resource
    Electronic Resource
    Localization of NuMA protein isoforms in the nuclear matrix of mammalian cells (1994)
    New York, NY : Wiley-Blackwell
    Cell Motility and the Cytoskeleton 29 (1994), S. 167-176 
    Details
    ISSN: 0886-1544
    Keywords: NuMA ; spindle ; nuclear matrix ; core filaments ; Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: Using a monoclonal antibody 2D3 generated against a kinetochore-enriched human chromosome preparation, we identified a high molecular mass protein with nuclear staining in interphase and polar staining of the pericentriolar region in the mitotic spindle. Initially termed centrophilin, this protein associates with the minus-ends of spindle microtubules (MT) and appears to be important in spindle organization [Tousson et al., 1991: J. Cell Biol. 112:427-440]. Comparison of a partial cDNA sequence obtained for centrophilin with the full length cDNA sequence of nuclear mitotic apparatus protein (NuMA) [Compton et al., 1992: J. Cell Biol. 116:1395-1408; Yang et al., 1992: J. Cell Biol. 116:1303-1317] has indicated that NuMA and centrophilin are the same protein. Using a polyclonal NuMA antibody, we have provided further evidence that NuMA exists as iso-forms as shown by peptide mapping and immunoblots. Sequential fractionation experiments along with immunofluorescence, immunoblotting, and EM immunogold labeling have demonstrated that NuMA isoforms are novel components of nuclear core filaments. Thus, NuMA, a long coiled-coil protein, appears to have dual functions in interphase and mitosis during the cell cycle. In interphase, NuMA likely plays a structural role in the nucleoskeleton that may be important in nuclear organization and functions, whereas in mitosis, NuMA appears to be associated with spindle MT organization and chromosome positioning. © 1994 Wiley-Liss, Inc.
    Additional Material: 6 Ill.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1002/cm.970290208
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  • 18
    Electronic Resource
    Electronic Resource
    Tubulin nucleation and assembly in mitotic cells: Evidence for nucleic acids in kinetochores and centrosomes (1980)
    New York, NY : Wiley-Blackwell
    Cell Motility and the Cytoskeleton 1 (1980), S. 1-15 
    Details
    ISSN: 0886-1544
    Keywords: centrosomes ; kinetochores ; microtubule initiation ; nuclease enzymes ; electron microscopy ; Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: A lysed cell system was used to study the organelle structure and nucleation of exogenous tubulin at kinetochores and centrosomes in mitotic PtK2 cells. We have used this lysed cell system in conjunction with nuclease digestion experiments to determine which specific nucleic acids (DNA or RNA) are involved in either the structure and/or microtubule-initiating capacity of kinetochores and centrosomes. The results indicate that DNase I specifically decondenses the kinetochore plate structure, with the eventual loss in the ability of the chromosomes to nucleate microtubule assembly. DNase I had no effect on either the structure or nucleating capacity of centrosomes. Both RNase T1 and RNase A specifically attacked the amorphous pericentriolar material of the centrosomes, with a concomitant loss in the ability of this material to nucleate microtubule formation. Neither RNase appeared to affect the structure or nucleating capacity of the kinetochore. Therefore, the two types of nucleases appear to exert preferential effects on the different types of microtubule initiation sites in mitotic mammalian cells. The results suggest that DNA is a major component of the kinetochore, while RNA is a major component of the amorphous pericentriolar material. These findings support the concept that microtubule initiation sites in mitotic cells contain nucleic acids which are essential for the structural and functional integrity of the sites.
    Additional Material: 45 Ill.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1002/cm.970010102
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  • 19
    Electronic Resource
    Electronic Resource
    Mitotic inhibition and chromosome displacement induced by estradiol in chinese hamster cells (1987)
    New York, NY : Wiley-Blackwell
    Cell Motility and the Cytoskeleton 7 (1987), S. 235-247 
    Details
    ISSN: 0886-1544
    Keywords: diethylstilbestrol ; estradiol ; microtubules ; mitotic apparatus ; cytoplasmic microtubule complex ; indirect immunofluorescence ; electron microscopy ; Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: We tested diethylstilbestrol (DES) and 17 β-estradiol as mitotic arrestants to determine their effects on chromosome distribution, spindle microtubules, and the cytoplasmic microtubule complex (CMTC) in the Chinese hamster strain Don. Cytological experiments assessed micronuclei induction, chromosome displacement, and anaphase recovery Indirect immunofluorescence microscopy with antibody to tubulin and electron microscopy were used to illustrate effects on microtubules. Both DES and estradiol were potent inhibitors of mitosis when applied to cells in vitro. Estradiol induced micronuclei at a greater frequency than did DES. Estradiol-arrested metaphases often contained misaligned chromosomes despite the presence of a bipolar spindle and an equatorial plate. Equatorial plates were not observed in DES-arrested cells. Cells recovered quickly from estradiol exposure upon removal of the steroid. The frequency of abnormal metaphases and abnormal anaphases declined as the recovery period increased. Microtubule experiments showed that DES inhibited spindle assembly and disassembled the CMTC, whereas estradiol, at similar concentrations, arrested mitosis in a manner that allowed spindle assembly. A definite effect on the CMTC by estradiol could not be determined. However, changes in cell morphology were observed. In the presence of estradiol, centrosomes organized microtubules that joined with kinet-ochores of chromosomes at the equatorial plate as well as with those of misaligned chromosomes. Misaligned chromosomes appeared predominantly at polar regions of mitotic cells. Following drug removal, the pole-oriented chromosomes reoriented at the equatorial plate. The unique arresting properties of estradiol may prove useful in studies of chromosome migration and segregation during mitosis.
    Additional Material: 10 Ill.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1002/cm.970070306
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  • 20
    Electronic Resource
    Electronic Resource
    Toward a structural and molecular definition of the kinetochore (1990)
    New York, NY : Wiley-Blackwell
    Cell Motility and the Cytoskeleton 16 (1990), S. 104-109 
    Details
    ISSN: 0886-1544
    Keywords: Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Additional Material: 1 Ill.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1002/cm.970160204
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