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PubMed1981

The behaviour of silver-positive structures during meiotic prophase of male mice.

Air dried preparations and sections of mouse seminiferous tubules were stained with an ammoniacal silver solution to study the behaviour of silver positive structures in meiotic prophase I nuclei. The presence of RNA was investigated using specific staining techniques and RNase digestion. Pachytene nuclei showed silver precipitation at the paracentromeric region of two to six autosomal bivalents. In spermatogonia at least the chromosome nos. 12, 15, 16, 17 and 18 proved to have transcriptionally active nucleolus organizer regions. From late pachytene the Ag-positive structures migrate towards the sex vesicle. In prediffuse diplotene, when the silver spots reached the sex vesicle, a vacuole-like body appeared near the sex vesicle. At the same time significant amounts of RNA accumulate near to the sex vesicle. Finally, in diffuse diplotene a tripartite structure could be observed, composed of (a) a horseshoe-shaped structure adjacent to the sex vesicle, which contains a great deal of RNA, (b) a vacuole-like body, being enclosed by the horseshoe and (c) an Ag-positive mass, migrated from the nucleolus organizer regions. It is probable that the tripartite structure, or at least a part of it, is a large nucleolus. The significance of the structure is discussed.

Animals↗
PubMed1976

Ultrastructure of cytoplasmic nucleolus-like bodies and nuclear RNP particles in late prophase of tipulid spermatocytes.

Late prophase stages of Pales ferruginea (Tipulidae) spermatocytes were examined by means of conventional electron microscopic section technique, combined with cytochemical methods. The cytoplasm of cells in diakinesis contains nucleolus-like bodies (NLB) 1 mum in diameter which are formed in diplotene at the pores of the nuclear membrane. They are compound structures consisting of fibro-granular RNP material which is associated wth one or two electron-dense gobules. The RNP material has a hollow core which contains polyribosomes. The NLBs possibly indicate rRNA gene amplification. At diakinesis the nucleus contains numerous electron-dense RNP particles scattered throughout the chromatin-free karyoplasm, and associated with the condensed chromosomes. The diameter of the chromation associated particles is markedly higher (mean 630 A) than that of the free particles (mean 540 A). The RNP particles seem to be aggregates of 200 A subunits. They are regarded as transcription products of chromosomal genes.

Animals↗
PubMed1976

The meotic prophase in Bombyx mori females analyzed by three dimensional reconstructions of synaptonemal complexes.

Serial sectioning followed by three dimensional reconstruction of lateral componenets of the synaptoemal complex have been used to follow chromosome pairing during theprophase of the achiasmatic meiotic division in the silkworm. Bombyx mori. During leptotene and early zygotene, the lateral components become attached to the nuclear envelope at a specific region, thus forming a chromosome bouquet. The attachment of lateral componenets to the nuclear envelope precedes the completion of the components between their attachment points. Synapsis and synaptonemal complex formation start during the period of lateral component organization in the individual nucleus. Telomeric movements on the nuclear envelope occur at two stages of the prophase: the chromosome pairing appears to be initiated by an association of unpaired ends of homologus chromosomes, the nature of this primary attraction and recognition being unknown. Secondly, the paired chromosomes become dispersed in the nucleus by shifting of attachment sites of completed synaptonemal complexes at the end of zygotene. This movement is possible related to a membranes flow occuring during this stage. Membrane material is synthesized at the region of synaptonemal complex attachment. Later, the excess membrane material is shifted to the opposite pole where it protudes into the lumen of the nuclei thus forming vacuoles. Two previously undescribed features of chromosomes paring were revealed. In late zygotene, chromosome pairing and synaptonmal complex formation were frequently observed to be delayed or even prevented over s short distance by interlocking two bivalents, both being attached to the nuclear envelope. Such interlocking of bivalents was not found in pachytene...

Animals↗
PubMed1981

Chromosome behaviour during early meiotic prophase of mouse primary spermatocytes.

A correlative light- and electron microscopical study has yielded both a description of chromosome behaviour during early meiotic prophase in mouse spermatocytes, and an accurate timing of the zygotene and early pachytene stage. The light microscopic part of the study consists of an analysis of orcein stained or C-banded, air-dried preparations, and series of separately squashed sections of seminiferous tubules. The mice used for air-dried preparations were treated with hyroxyurea and triaziquone, which reduces the spermatocyte population to a small, well-defined group of cells in meiotic interphase. The development of the restricted spermatocyte population is followed during four days. The electron microscopical study consists of an investigation of synaptonemal complexes in thin sections of seminiferous tubules in various stages of the epithelial cycle. It turned out that synaptic pairing starts about half a day after the end of the DNA replication phase. At most one and a half day later chromosome pairing is completed. In very early pachytene spermatocytes a sex vesicle is not visible. The orientation of the X and Y chromosome into a sex vesicle starts about one day after the beginning of the pachytene stage. The existence of a leptotene stage is discussed. The presence of a pre-leptotene contraction phase in male mice is excluded.

Animals↗
PubMed1975

Nucleolar RNA synthesis of meiotic prophase spermatocytes in the human testis.

Human meiotic prophase spermatocyte nuclei were studied by electron microscope autoradiography after a 3 hours 3H-uridine labeling pulse, followed by postincubation in non-radioactive medium. In autosomes, 3H-uridine nucleolar labeling reaches a peak during early-middle zygotene prior to the peak labeling of chromosmal RNA species at middle pachytene. Transcription activities of sex chromosomes are inconspicuous when compared with that of autosomes. An increasing condensation of nucleolar-associated chromatin in acrocentric bivalents contributes to the formation of basal knobs in human pachytene spermatocytes. Upon completion of knob formation, nucleolar components segregate and the uptake of 3H-uridine decreases. These findings suggest that the template capability of ribosomal DNA cistrons, located next to the basal knob region, is largely associated with a dispersed state of chromatin whereas increased chromatin condensation is correlated with a restriction of ribosomal RNA transcription.

Autoradiography↗
PubMed1982

Ultrastructural characterization of the meiotic prophase. A tool in the assessment of radiation damage in man.

The three-dimensional reconstruction of meiotic nuclei from serial sections micrographed in the electron microscope has provided information about man and several other organisms that is not obtainable by light microscopy or biochemical analysis. At zygotene, the previously unpaired chromosomes align and form synaptonemal complexes between homologous chromosome segments either by progressive initiation from the telomeres or by interstitial recognition. Chromosome and bivalent interlocking at zygotene is a regular phenomenon and occurs at a frequency of 0.7-4.0 per nucleus in samples of meiocytes analyzed from different organisms. This frequency is reduced to 0.1 per nucleus at pachytene. The interlockings are resolved by breakage and precise rejoining of the broken ends. This breakage and rejoining can also occur in the absence of the DNA nicking and repair involved in crossing-over. The synaptonemal complexes combining homologous chromosome segments are stabilized by recombination nodules, after which a second round of synaptonemal complex formation between as yet unpaired or unstably paired chromosome segments occurs, apparently for optimization of bivalent formation. Non-homologous pairing with the synaptonemal complex can take place in this phase of pachytene. Continuity between recombination nodules and chromatin chiasmata has been traced at the ultrastructural level but not all nodules lead to chiasmata. The distributions of recombination nodules among the bivalents and along the bivalents at successive stages of meiotic prophase show that the nodules are placed at random at early-zygotene after which bivalents without nodules have preference for the acquisition of these structures. Chiasma interference appears as a consequence of the limited number of recombination nodules available together with a decreased affinity of a bivalent arm with a nodule for additional ones. The relevance of these observations in the study of genetic damage by radiation is discussed.

Aneuploidy↗
PubMed1976

A novel temperature-sensitive mammalian cell line exhibiting defective prophase progression.

A temperature-sensitive mammalian cell line has been isolated which grows and divides normally at the permissive temperature of 33 degrees C. When incubated at 39 degrees C, the nonpermissive temperature, interphase cells continue to enter a prophase-like state. Chromatin-like material condenses and coalesces into dark-staining clumps rather than into discernible chromosomes. Disappearance of the nuclear boundary is observed, but re-formation of the boundary around the clumps fails to occur. In corporation of labeled precursors reveals a decrease in protein synthesis which is accompanied by a slower decrease in DNA synthesis. Approximately 0.2% of the mutant cells revert in their capability of growth and cell division at 39 degrees C. These "revertants" are found to contain a higher number of chromosomes. The isolation of this mutant is based on the initial observation that the cells become rounded at the nonpermissive temperature. The cell-rounding process characteristic of mitotic cells should serve as a useful marker in the isolation of mitotic mutants.

Animals↗
PubMed1995

High-resolution cytogenetic mapping of the short arm of chromosome 1 with newly isolated 411 cosmid markers by fluorescence in situ hybridization: the precise order of 18 markers on 1p36.1 on prophase chromosomes and "stretched" DNAs.

A high-resolution cytogenetic map of the short arm of chromosome 1 with newly isolated 411 cosmid markers was constructed by fluorescence in situ hybridization (FISH). These markers were scattered throughout chromosome 1p, but they were preferentially concentrated on R-band dominant regions such as 1p36, 1p34, 1p32, 1p22, and 1p13. Among these markers, 197 were localized on chromosome band 1p36, a region frequently deleted in neuroblastoma. Of these, 18 were precisely ordered on 1p36.1 by multicolor FISH of prophase chromosomes and "stretched" DNAs as follows: 1pter-163-41-11-1-226-586-568-614-631-665-451-199-190-561-241-74-1 76-652-1cen. The high-density map of chromosome 1p constructed here can provide useful landmarks for constructing a contig map of the short arm of chromosome 1 with YACs and cosmid clones and will expedite the identification of breakpoints and/or tumor suppressor gene(s) associated with several types of malignant tumors that frequently exhibit chromosomal aberrations or deletions of chromosome 1p.

Animals↗
PubMed2005

Mnd2, an essential antagonist of the anaphase-promoting complex during meiotic prophase.

Meiotic cohesin serves in sister chromatid linkage and DNA repair until its subunit Rec8 is cleaved by separase. Separase is activated when its inhibitor, securin, is polyubiquitinated by the Cdc20 regulated anaphase-promoting complex (APC(Cdc20)) and consequently degraded. Differently regulated APCs (APC(Cdh1), APC(Ama1)) have not been implicated in securin degradation at meiosis I. We show that Mnd2, a factor known to associate with APC components, prevents premature securin degradation in meiosis by APC(Ama1). mnd2Delta cells lack linear chromosome axes and exhibit precocious sister chromatid separation, but deletion of AMA1 suppresses these defects. Besides securin, Sgo1, a protein essential for protection of centromeric cohesion during anaphase I, is also destabilized in mnd2delta cells. Mnd2's disappearance prior to anaphase II may activate APC(Ama1). Human oocytes may spend many years in meiotic prophase before maturation. Inhibitors of meiotic APC variants could prevent loss of chiasmata also in these cells, thereby guarding against aberrant chromosome segregation.

Anaphase-Promoting Complex-Cyclosome↗
PubMed1997

Plant CDC2 is not only targeted to the pre-prophase band, but also co-localizes with the spindle, phragmoplast, and chromosomes.

A polyclonal antiserum against the p34cdc2 homologue of Arabidopsis thaliana, CDC2aAt, was used in parallel with a polyclonal antiserum against the PSTAIRE motif to study the subcellular localization of CDC2 during the cell cycle of isolated root tip cells of Medicago sativa. During interphase, CDC2 was located in the nucleus and in the cytoplasm. The cytoplasmic localization persisted during the complete cell cycle, whereas the nuclear signal disappeared at nuclear envelope breakdown. At the beginning of anaphase, the anti-CDC2aAt antibody transiently co-localized with condensed chromosomes. The chromosomal co-localization disappeared as anaphase continued and remained excluded from the separated chromosomes until cytokinesis, when CDC2 re-located to the newly forming nuclei. We also observed a co-localization of CDC2 with three microtubular structures, the pre-prophase band, the spindle, and the phragmoplast.

Arabidopsis↗
PubMed2002

The dissociation of cohesin from chromosomes in prophase is regulated by Polo-like kinase.

The separation of sister chromatids in anaphase depends on the dissociation of cohesin from chromosomes. In vertebrates, some cohesin is removed from chromosomes at the onset of anaphase by proteolytic cleavage. In contrast, the bulk of cohesin is removed from chromosomes already in prophase and prometaphase by an unknown mechanism that does not involve cohesin cleavage. We show that Polo-like kinase is required for the cleavage-independent dissociation of cohesin from chromosomes in Xenopus. Cohesin phosphorylation depends on Polo-like kinase and reduces the ability of cohesin to bind to chromatin. These results suggest that Polo-like kinase regulates the dissociation of cohesin from chromosomes early in mitosis.

Animals↗
PubMed2003

Prophase destruction of Emi1 by the SCF(betaTrCP/Slimb) ubiquitin ligase activates the anaphase promoting complex to allow progression beyond prometaphase.

Progression through mitosis occurs because cyclin B/Cdc2 activation induces the anaphase promoting complex (APC) to cause cyclin B destruction and mitotic exit. To ensure that cyclin B/Cdc2 does not prematurely activate the APC in early mitosis, there must be a mechanism delaying APC activation. Emi1 is a protein capable of inhibiting the APC in S and G2. We show here that Emi1 is phosphorylated by Cdc2, and on a DSGxxS consensus site, is subsequently recognized by the SCF(betaTrCP/Slimb) ubiquitin ligase and destroyed, thus providing a delay for APC activation. Failure of betaTrCP-dependent Emi1 destruction stabilizes APC substrates and results in mitotic catastrophe including centrosome overduplication, potentially explaining mitotic deficiencies in Drosophila Slimb/betaTrCP mutants. We hypothesize that Emi1 destruction relieves a late prophase checkpoint for APC activation.

Amino Acid Motifs↗
PubMed1967

DNA-dependent formation of the synaptinemal complex at meiotic prophase.

EVALAUTION OF MICROSPOROCYTES CULTURED DURING DISCRETE PERIODS OF MEIOTIC PROPHASE IN THE PRESENCE OF DEOXYADENOSINE, AN INHIBITOR OF DNA SYNTHESIS, INDICATE THAT: (1) late leptonema or early zygonema DNA synthesis is required to initiate the formation of the synaptinemal complex; (2) DNA synthesized during late zygonema is necessary for the disjunction of the paired homologs at diplonema; and (3) DNA synthesis in pachynema is a requisite for normal anaphase II separation of sister chromatids.

Chromosomes↗
PubMed1972

Induction of prophase in interphase nuclei by fusion with metaphase cells.

Fusion of an interphase cell with a metaphase cell results in profound changes in the interphase chromatin that have been called "chromosome pulverization" or "premature chromosome condensation" In addition to the usual light microscopy, the nature of the changes has been investigated in the present study with electron microscopy and biochemical techniques Metaphase and interphase cells were mixed and fused at 37 degrees C by means of ultraviolet-inactivated Sendai virus. After cell fusion, morphological changes in interphase nuclei occurred only in binucleate cells which contained one intact set of metaphase chromosomes Irrespective of the nuclear stage at the time of cell fusion, the morphologic changes that occurred 5-20 min later simulated very closely a sequence of events that characterizes the normal G(2)-prophase transition. Radioautography revealed that, late in the process, substantial amounts of RNA and probably protein were transferred from the interphase nucleus into the cytoplasm of fused cells. Thus, the findings indicate the existence in metaphase cells of factor(s) which are capable of initiating biochemical and morphological events in interphase nuclei intrinsic to the normal mitotic process.

Animals↗
PubMed1974

Transcription sites in spread meiotic prophase chromosomes from mouse spermatocytes.

Mouse spermatocytes at pachytene stage have been examined by whole-mount electron microscope techniques complemented with autoradiography as an approach for visualizing their transcriptive activity. Structural elements of meiotic bivalents, such as synaptonemal complexes and chromatin fibers, have been satisfactorily displayed in the total set of autosomal and sexual bivalents in single spermatocytes. Adequate preservation of the entire set of bivalents has provided a basis for recognition of sites where presumptive preribosomal RNA and heterogeneous nuclear RNA species are being transcribed at different segments of autosomal bivalents. Nucleoli attached to the basal knob region where nucleolar organizer cistrons are assumed to be located and ribonucleoprotein fibrils associated with distinct chromatin loops have been recognized. These structural findings have been correlated with display of [(3)H]uridine incorporation sites in thin-section and whole-mount electron microscopy autoradiographic preparations. A low transcriptive activity of the sexual bivalent contrasted with extensive gene expression in autosomal bivalents. Each sex chromosome shows a double axial core. A short region of pairing with a synaptonemal complex joins the two chromosomes at one end. We conclude that variations in the rate of RNA synthesis throughout meiotic prophase stages in the mouse are expressed as fluctuations in the amount and distribution of distinct RNA species at specific segments of the bivalents.

Animals↗
PubMedDate not supplied

Macronuclear development in conjugants of Tetrahymena thermophila, which were artificially separated at meiotic prophase.

Conjugant pairs of Tetrahymena thermophila were mechanically separated by vigorous pipetting at the early stages of meiotic prophase. The complete sequence of conjugational nuclear events including the appearance of pronuclei, development of the new macronuclei (postzygotic development), and resorption of the old macronuclei was observed in the separated cells, without pronuclear exchange. The pronuclei in the separated cells were recognised by the presence of components of the extranuclear cytoskeleton, which were labelled with anti-tubulin and anti-fenestrin antibodies in the same way as in undisturbed conjugants. The apical region of the separated conjugants (the post-junction area), corresponding to the junction area of conjugants was labelled with anti-fenestrin antibody and maintained the properties required for the nuclear development. The results of the genetic study were consistent with a hypothesis that cytogamy (pronuclear fusion) was induced in the separated conjugants. Therefore, the lasting cell contact is not necessary for the successful completion of conjugational nuclear events.

Animals↗
PubMed1978

Autoradiographic study of RNA synthesis during meiotic prophase in the human oocyte.

The incorporation of 3H-uridine in oogonia and oocytes during meiotic prophase I was studied in three human fetuses 13, 18, and 19 weeks old. Following a 40- or 60-min pulse, intense nuclear and nucleolar labeling was observed in oogonia. During the preleptotene chromosome condensation stage, the heteropycnotic masses were unlabeled, while numerous silver grains were seen on the filaments persisting around these masses. During leptotene, chromosomal and nucleolar RNA synthesis was significant, but less than that in the oogonia. The rate of incorporation declined rapidly during zygotene and fell to a very low level at early pachytene. Throughout pachytene no nucleolar RNA synthesis was observed. Chromosomal RNA synthesis progressively recovered during middle pachytene, was of moderate intensity at late pachytene, and increased again at early diplotene. Nucleolar RNA synthesis was very intense at early diplotene, at the same time as nucleolar size and basophilia increased.

Autoradiography↗
PubMed1979

Nonhomologous associations of C-heterochromatin at human male meitoic prophase.

In human male meiotic prophase, nonhomologous pairings and connections between C-heterochromatic regions were demonstrated by C-banding and G-11 staining. Approximately 40% of the pachytene stages exhibited bivalent associations at regions of C-heterochromatin. Nonhomologous associations were seen between all possible morphological types of bivalents. Evidence is submitted suggesting that nonhomologous associations between regions of C-heterochromatin are a fairly common event in many organisms, including man, and the possible roles of this phenomenon in meiosis are discussed.

Azure Stains↗