An ultrastructural and radioautographic study of the chromocentric interphase nucleus in plant meristematic cells (Raphanus sativus)

1976 ◽  
Vol 21 (1) ◽  
pp. 193-207
Author(s):  
A. Lord ◽  
J.G. Lafontaine

In Raphanus sativus, the mitotic chromosomes are quite short and, on reaching the cell poles, soon undergo extensive unravelling. By late telophase and early interphase, only a few chromosome segments, believed to correspond to the centromeric regions, are still visible in the form of chromocentres closely associated with the nuclear envelope. Although interphase nuclei show little internal structural differentiation, high-resolution radioautography has permitted us to establish which of them have reached the early, mid and late S periods. In early S nuclei, only the nucleolus and the euchromatin which pervades the nuclear cavity become labelled. By the mid S-period, the diffuse chromatin and nucleolus incorporate less thymidine and DNA synthesis is initiated within the peripheral chromocentres. Subsequently, the radioautographic grains become restricted to the chromocentres. The finding that certain late S nuclei exhibit loosely organized chromocentres strongly suggests that these heterochromatic chromosome segments undergo important conformational modifications during DNA replication. Finally, the presence of radioautographic grains over the lacunar regions of the nucleolus in early and mid S nuclei demonstrates that intranucleolar DNA replicates during the earlier portion of the S-period.

1974 ◽  
Vol 14 (2) ◽  
pp. 263-287
Author(s):  
J. G. LAFONTAINE ◽  
A. LORD

Radioautography under both light and electron microscopy was exploited to investigate the structural changes of the chromatin reticulum which characterizes the interphase nucleus of a number of plants. Allium porrum meristematic plant cells were used for this purpose. In this species, the telophase chromosomes uncoil into dense strands which, during the G1 period, gradually give rise to a coarse reticulum. There then follows an extensive unravelling of portions of these strands, and high-resolution radioautography reveals that labelling with tritiated thymidine predominantly occurs over zones of the nucleus consisting of diffuse fine fibrillar material. As the S-period progresses, a chromatin reticulum reappears throughout the nuclear cavity, the tortuous strands being approximately 0.25 µm in diameter. Most of the radioautographic grains still remain over the light nucleoplasmic areas but a number of these are now located on the outermost portion of the dense chromatin profiles. By the end of the S-period, the chromatin strands are slightly thicker (ca. 0.3 µm) and form a looser reticulum. Labelling has decreased noticeably in nuclei of that period, the radioautographic grains being grouped into clusters resting over more or less spherical regions of the chromatin reticulum. Judging from their localization at the surface of the nucleolus or close to the nuclear envelope, these structures correspond to chromocentres. The additional interesting finding that such nuclear structures appear much less compactly organized strongly suggests that chromocentres undergo important conformational modifications during duplication of their DNA.


1949 ◽  
Vol 32 (4) ◽  
pp. 489-502 ◽  
Author(s):  
Hans Ris ◽  
A. E. Mirsky

In the living interphase nucleus no chromosomal structures are visible. Yet in the injured cell and after treatment with most histological fixatives chromatin structures become apparent. Under certain conditions this appearance of structure in the living interphase nucleus is reversible. We have found that this change in the interphase nucleus is the result of a change in the state of the chromosomes. In the living nucleus the chromosomes are in a greatly extended state, filling the entire nucleus. Upon injury the chromosomes condense and therefore become visible. At the same time the nuclear volume decreases. This behavior of the chromosomes is connected with their content of desoxyribonucleic acid (DNA). This view is based on the following observations: (a) Distribution of DNA in the Nucleus.—(1) The living interphase nucleus of uninjured cells absorbs diffusely at 2537 Å. No chromosomal structures are visible in ultraviolet photographs unless they are also distinct in ordinary light. If the chromosomes are made to condense they become visible and the absorption at 2537 Å is now localized in these structures. (2) After fixation with formalin and osmic acid interphase nuclei stain diffusely with Feulgen. These fixatives preserve the extended state of the chromosomes. (3) If nuclei are teased out in non-electrolytes (sucrose, glycerin) the chromosomes are extended. Such nuclei stain homogeneously with methyl green. On adding salts the chromosomes condense and the methyl green is now restricted to the visible structures. (b) Extension and Condensation of Isolated Chromosomes.—When chromosomes isolated from interphase nuclei of calf thymus are suspended in sucrose, their volume is four to five times larger than in saline, but they retain their characteristic shapes. Chromosomes from which DNA and histone have been removed do not show this reversible extension and condensation, neither do lampbrush chromosomes of frog oocytes which contain very little DNA. During mitosis a partial condensation of the DNA occurs in prophase, so that the mitotic chromosomes now occupy a much smaller volume of the nucleus. At telophase the chromosomes swell again to fill the entire nucleus.


1983 ◽  
Vol 25 (5) ◽  
pp. 478-486 ◽  
Author(s):  
Catharine P. Fussell

The arrangement of telomeres in eight types of differentiated Allium cepa L. interphase cells was studied to find whether the distribution pattern varies with differentiation as centromere distribution appears to in differentiated cells of mice and Triturus vulgaris L. The results show that telomere positions and groupings in A. cepa are essentially the same in differentiated and meristematic interphase nuclei; telomeres, which are roughly paired, are arranged in a telophase configuration along one side of the nucleus. Thus telomeres appear to maintain the same basic arrangement in differentiated and in meristematic cells. Comparison of chromosome arm lengths and interphase telomere positions suggests that interphase telomere arrangements are a function of chromosome arm lengths at the time the nuclear envelope forms. Such an arrangement would place homologous telomeres in the same latitudinal band of the interphase nucleus.


1997 ◽  
Vol 110 (17) ◽  
pp. 1979-1988 ◽  
Author(s):  
H. Bousbaa ◽  
L. Correia ◽  
G.J. Gorbsky ◽  
C.E. Sunkel

The progression of cells from metaphase to anaphase is thought to be regulated by a checkpoint that delays entry into anaphase until all chromosomes reach a stable bi-polar attachment at the metaphase plate. Previous work has suggested that the 3F3/2 kinetochore phosphoepitopes are involved in this checkpoint system. We show that the 3F3/2 centromere phosphoepitopes are present in Kc cells, third instar larval neuroblasts and isolated chromosomes of Drosophila melanogaster. In tissue culture cells and neuroblasts isolated from third instar larvae, centromere labelling is detected from early prophase to the metaphase-anaphase transition but absent once cells center anaphase. During anaphase, the antibody stains the spindle mid zone and during telophase the midbody is labelled until cells separate. In both cell types, the 3F3/2 antibody stains the centrosome from prophase to late telophase. The 3F3/2 staining is retained in Kc cells and third instar larval neuroblasts arrested at the prometaphase state with microtubule inhibitors. Also, two mitotic mutants that show abnormal spindle morphology retain the centromere labelling in a metaphase-like configuration, suggesting that they activate the metaphase-anaphase checkpoint. Finally, mitotic chromosomes isolated in the presence of a phosphatase inhibitor show phosphoepitopes at the primary constriction on the surface of each chromatid, however, chromosomes isolated in the absence of a phosphatase inhibitor do not. Incubation of these chromosomes with ATP causes the rephosphorylation of the phosphoepitopes at the centromere.


2002 ◽  
Vol 50 (12) ◽  
pp. 1697-1698 ◽  
Author(s):  
Ulf Steinhaeuser ◽  
Heike Starke ◽  
Angela Nietzel ◽  
Joerg Lindenau ◽  
Peter Ullmann ◽  
...  

We describe a versatile method for performing fluorescence in situ hybridization (FISH) in suspension instead of on a slide as usually done. This so-called suspension-FISH (S-FISH) opens new possibilities for the analysis of shape and functions of the human interphase nucleus. The procedure is described and the first results using this approach are presented.


1997 ◽  
Vol 110 (4) ◽  
pp. 417-420 ◽  
Author(s):  
Carlos Roberto de Carvalho ◽  
Luiz Sergio Saraiva

1974 ◽  
Vol 62 (1) ◽  
pp. 104-113 ◽  
Author(s):  
Yoshitaka Obara ◽  
Lee S. Chai ◽  
Herbert Weinfeld ◽  
Avery A. Sandberg

Fusing human HeLa metaphase cells with HeLa interphase cells resulted within 30 min in either of two phenomena in the resultant binucleate cell: either prophasing of the interphase nucleus or formation of a normal-appearing nuclear envelope around the metaphase chromosomes. The frequency of either occurrence was strongly dependent on environmental pH. At pH's of 6.6–8.0, prophasing predominated; at pH 8.5 nuclear envelope formation predominated. Additionally, the frequencies of the two events in multinucleate cells depended on the metaphase/interphase ratio. When the ratio was 0.33 nuclear envelope formation predominated; when it was 2.0 prophasing predominated. In their general features, the results with fused HeLa cells resembled those reported earlier with fused Chinese hamster Don cells. However, the results provided an indication that between pH 6.6 and 8.0 the HeLa metaphase cells possessed a much greater capacity than the Don metaphase cells to induce prophasing. Fusion of Don metaphase cells with HeLa interphase cells or of Don interphase cells with HeLa metaphase cells at pH 8.0 resulted in nuclear envelope formation or prophasing in each kind of heterokaryon. As in the homokaryons, the frequencies of the two events in the heterokaryons depended on the metaphase/interphase ratio. The statistics of prophasing and nuclear envelope formation in the homo- and heterokaryon populations were consistent with the notion that disruption or formation of the nuclear envelope depends on the balance attained between disruptive and formative processes.


1979 ◽  
Vol 39 (1) ◽  
pp. 13-27
Author(s):  
J.G. Lafontaine ◽  
B.T. Luck ◽  
D. Dontigny

Loose, fibrillar, spherical structures have been observed during recent years in interphase nuclei of both animal and plant cells. These nuclear formations have been referred to as karyosomes, fibrillar bodies, micropuffs and centromeres. In order to gain further information on the nature of these structures, a cytochemical and radioautographic investigation was undertaken using plant meristematic cells (Allium porrum). For that purpose roots were fixed with either formaldehyde or glutaraldehyde in order to carry out cytochemical tests for DNA, RNA and proteins. Certain of the preparations were also first digested with DNase, RNase or proteinase K and then stained according to different procedures. Other specimens were labelled with thymidine for high-resolution radioautographic observations. Staining with diaminobenzidine (DAB) revealed that these nuclear puff-like formations consisted partly of a loose fibrillar meshwork containing nucleic acids. Part of this fine fibrillar reticulum persisted whether the preparations were digested with DNase or RNase before staining with DAB, thus indicating that these nuclear structures contained both DNA and RNA. The fact that these formations incorporate thymidine furnished additional support for the view that they correspond to specific chromosome segments. Staining with ethanolic phosphotungstic acid or digestion of specimens with proteinase K showed that these loose fibrillar structures also consisted of proteins. Judging from their ultrastructure, their association with the chromatin reticulum as well as from their cytochemical characteristics, these nuclear formations most likely correspond to centromeres. In view of the presence of DNA within these structures, it is possible to distinguish them from other equally spherical nuclear formations, observed in certain plant species, that have generally been referred to as karyosomes or micronucleoli and that appear to consist of ribonucleoproteins.


1976 ◽  
Vol 70 (3) ◽  
pp. 592-607 ◽  
Author(s):  
I B Heath ◽  
M C Heath

Aspects of the ultrastructure of mitotic nuclei of the fungus Uromyces phaseoli var. vignae are described from both intercellular hyphae in the cowpea host and infection structures induced to differentiate in vitro. The interphase nucleus-associated organelle (NAO) consists of two trilamellar acircular disks connceted by an osmiophilic bar. The intranuclear spindle develops between these disks when they separate. The spindle contains pole to pole, interdigitating, chromosomal, and fragmentary microtubules arranged to form a central bundle along the surface of which lie the metaphase chromosomes. No metaphase plate is found. There are up to three microtubules per kinetochore and approximately 14 chromosomes on the haploid spindle. Telophase elongation appears to involve extension of pole to pole microtubules with no evidence for the remaining presence of interdigitating microtubules. Concomitantly, numerous cytoplasmic microtubules develop from each NAO disk where few or none are present in other phases. Reformation of the interphase NAO involves the formation of a sausage-shaped intermediate at late telophase. The nuclear envelope remains intact and the nucleolus persists throughtout division. Various aspects of the spindle and NAOs appear to be evolutionary intermediates between Ascomycetes and higher Basidiomycetes, thus supporting the theory of Basidiomycete evolution from the former group and demonstrating an encouraging correlation between mitotic characteristics and other phylogenetic markers.


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