Metaphase

About: Metaphase is a research topic. Over the lifetime, 6925 publications have been published within this topic receiving 291590 citations. The topic is also known as: GO:0007091 & mitotic metaphase/anaphase transition.

Functional autonomy of monopolar spindle and evidence for oscillatory movement in mitosis.

Abstract: The oscillations of chromosomes associated with a single spindle pole in monocentric and bipolar spindles were analysed by time-lapse cinematography in mitosis of primary cultures of lung epithelium from the newt Taricha granulosa. Chromosomes oscillate toward and away from the pole in all stages of mitosis including anaphase. The duration, velocity, and amplitude of such oscillations are the same in all stages of mitosis. The movement away from the pole in monocentric spindle is rapid enough to suggest the existence of a previously unrecognized active component in chromosome movement, presumably resulting from a pushing action of the kinetochore fiber. During prometaphase oscillations, chromosomes may approach the pole even more closely than at the end of anaphase. Together, these observations demonstrate that a monopolar spindle is sufficient to generate the forces for chromosome transport, both toward and away from the pole. The coordination of the aster/centrosome migration in prophase with the development of the kinetochore fibers determines the course of mitosis. After the breaking of the nuclear envelope in normal mitosis, aster/centrosome separation is normally followed by the rapid formation of bipolar chromosomal fibers. There are two aberrant extremes that may result from a failure in coordination between these processes: (a) A monocentric spindle will arise when aster separation does not occur, and (b) an anaphaselike prometaphase will result if the aster/centrosomal complexes are already well-separated and bipolar chromosomal fibers do not form. In the latter case, the two monopolar prometaphase half-spindles migrate apart, each containing a random number of two chromatid (metaphase) monopolar-oriented chromosomes. This random segregation of prometaphase chromosome displays many features of a standard anaphase and may be followed by a false cleavage. The process of polar separation during prometaphase occurs without any visible interzonal structures. Aster/centrosomes and monopolar spindles migrate autonomously by an unknown mechanism. There are, however, firm but transitory connections between the aster center and the kinetochores as demonstrated by the occasional synchrony of centrosome-kinetochore movement. The data suggest that aster motility is important in the progress of both prometaphase and anaphase in normal mitosis.

Antibodies to defined histone epitopes reveal variations in chromatin conformation and underacetylation of centric heterochromatin in human metaphase chromosomes.

Abstract: Unfixed metaphase chromosome preparations from human lymphocyte cultures were immunofluorescently labelled using antibodies to defined histone epitopes. Both mouse monoclonal antibody HBC-7, raised against the N-terminal region of H2B, and rabbit serum R5/12, which recognizes H4 acetylated at Lys-12, gave non-uniform labelling patterns, whereas control antibodies against total histone fractions H4 and H1 produced homogeneous fluorescence. HBC-7 bound approximately uniformly to the bulk of the chromosomes, but the major heterochromatic domains of chromosomes 1, 9, 15, 16 and the Y showed significantly brighter fluorescence. Serum R5/12 indicated an overall reduction in acetylation of H4 in metaphase chromosomes compared with interphase nuclei, although some specific chromosomal locations had considerably elevated acetylation levels. Acetylation levels in the major heterochromatic domains appeared extremely low. To investigate further the differences noted in heterochromatin labelling, metaphases from cultures grown in the presence of various agents known to induce undercondensation of the major heterochromatic domains were similarly immunolabelled. Decondensed heterochromatin no longer exhibited higher than normal immunofluorescence levels with HBC-7. The higher resolution afforded by "stretching" the centromeric heterochromatin of chromosomes 1, 9 and 16 confirmed the low level of H4 acetylation in these domains. We consider the implications of these observations in relation to chromatin conformation and activity.

Cell cycle behavior of human HP1 subtypes: distinct molecular domains of HP1 are required for their centromeric localization during interphase and metaphase

Abstract: Heterochromatin protein 1 (HP1) plays an important role in heterochromatin formation. Three subtypes of HP1, namely HP1α, β, and γ, have been identified in humans. In this study, using yellow fluorescent protein (YFP) fusion constructs, we examined the intracellular localization of human HP1 subtypes during the cell cycle. During interphase, all three HP1 subtypes were localized to centromeric heterochromatin and to promyelocytic leukemia (PML) nuclear bodies. Different preferences, however, were observed among the subtypes: during interphase HP1β localized most preferentially to centromeric heterochromatin, whereas HP1α and γ were more preferentially localized to PML nuclear bodies. During metaphase, only HP1α, was localized to the centromere. We thus determined which molecular domains of HP1 were necessary for their intracellular localization. Our results showed that the C-terminal fragment (amino acid residues 101-180) of HP1α was necessary for localization to the metaphase centromere and the Nterminal fragment (amino acid residues 1-76) of HP1β was necessary for localization to the interphase centromere. Interestingly, simultaneous observations of residues 101180 of HP1α and residues 1-76 of HP1β in living HeLa cells revealed that during late prophase, the HP1β fragment dissociated from centromeric regions and the HP1α fragment accumulated in centromeric regions. These results indicate that different specific regions of human HP1α and HP1β mediate localization to metaphase and interphase centromeric regions resulting in association of different subtypes of HP1 with the centromere at different times during the cell cycle. Summary

The effects of cooling human oocytes.

Abstract: Preovulatory human oocytes were cooled to 0 degrees C at 1 degree C/min, with or without the cryoprotectant dimethyl sulphoxide (DMSO), to assess the effects of cooling on the meiotic spindles and on oocyte structure. Batches of oocytes, cultured for 3-9 h, were held at 0 degrees C for 20 or 60 min and then fixed for transmission electron microscopy (TEM) either at 0 or 8 degrees C. Control oocytes were not cooled and were fixed at 22 or 37 degrees C for comparison. TEM revealed that 80% of the oocytes were at metaphase II, while 20% were at metaphase I and most had resumed meiosis recently. Control oocytes had more or less barrel-shaped meiotic spindles composed of microtubules (MT), some associated with chromosomes at kinetochores. Both metaphase I and II spindles were disassembled when cooled and fixed at 0 degrees C, with or without DMSO, due to extensive depolymerization of MT. The few MT that survived were found at the poles or were bundled together or were associated with chromosomes. Kinetochores were not prominent. Some oocytes cooled with DMSO and fixed at 0 or 8 degrees C showed evidence of MT, but the spindles were still disorganized and were abnormal in structure. Chromosomes tended to clump together or were dislocated in the cortical ooplasm in cooled oocytes, but widespread scattering was not observed. This was particularly evident in the absence of DMSO. Elements of the endoplasmic reticulum, Golgi, mitochondria and the cytosol were also adversely affected in some of the cooled oocytes and their surrounding cumulus cells. The results show that meiotic spindles are very sensitive to simple cooling and that DMSO does not provide substantial stabilization of the meiotic spindle even at 0 degrees C. The findings are discussed with reference to recent work on frozen human and mouse oocytes.