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PubMed1984

[Ultrastructural changes in female germ cells during the meiotic prophase in the rat: a special study of membranes after cryofracture].

Ultrastructural changes in the nuclear and cytoplasmic elements in the germ cells of female rats were followed before meiotic prophase (15.50 days post-coïtum and 17.25 days post-coïtum) and during it (17.75 days post-coïtum to birth). We observed: modifications in the nuclear envelope which was thick during the oogonial stage, becoming thinner when the chromosomes entered preleptotene stage. The thinning of the envelope was due to the disappearance of the chromatin material lining it; variations in the number and distribution of germ cell nuclear pores according to stage; the pores were first scattered in small clusters of 6 to 8 over the entire nuclear membrane. From the preleptotene to zygotene stage, these clusters enriched in pores to form large areas. Finally, in the pachytene and diplotene stages, clusters of more than 100 pores were seen; nucleolar fragmentation from the preleptotene stage, followed by the formation of a new active nucleole in the diplotene; polarization of the mitochondria in the oldest oogonia just before the beginning of meiotic prophase. This polarization disappeared after the onset of the meiotic processes, then appeared again near the developing Golgi apparatus at the end of the pachytene stage; the formation of large gap junctions and numerous bands of tight junctions between the somatic cells; these formations contrasted with small gap junctions, and the tight junctions became scarce just before the meiotic process began. These observations, as well as those concerning nuclear pore distribution were made using the cryofracture technique.

Animals↗
PubMedDate not supplied

Relationship between G band number and chromosome length from prophase to metaphase.

The statistical analysis of the distribution of human chromosomes length served as a basis for their classification according to the suggestions of the Denver Conference (1960). Standards of metaphasal chromosome length were used for programmes of computer-assisted classification. The introduction of banded chromosomes in the study provided a new measurable feature which was used for classification standards accepted at the Paris Conference (1971). The measurements of these two features have been used also in the mathematical models of computer analysis and classification of chromosomes. In the present study the relationship between the number of bands and the length of chromosomes was studied in the period of cell division from prophase to metaphase in the cultures of leucocytes of healthy people. This relationship is strongly correlated and can be approximated in description to simple regression in the period from prophase to metaphase. The relationship will facilitate the construction of dynamic patterns for the needs of computer-assisted classification of chromosomes and detection of structural aberrations.

Cells, Cultured↗
PubMed1980

[Dynamics of chromosome rearrangements in the meiotic prophase in human oogenesis].

A quantitative analysis of germ cell population development in human female embryonic gonads has been carried out, with special attention being paid to chromosomal changes in the prophase of the first meiotic division. 97 embryos at the age of 6 to 40 weeks were investigated. The morphology of nuclei at different prophase stages is described, temporal dynamics of all the stages is presented, dynamics of germ cell degeneration is outlined.

Chromosomes, Human↗
PubMed1994

Cryopreservation of human prophase I oocytes collected from unstimulated follicles.

OBJECTIVE: To evaluate the cryopreservation of immature human oocytes obtained from unstimulated ovarian tissue. DESIGN: Immature prophase I oocytes were obtained from unstimulated follicles and were either cryopreserved or cultured as controls. Cryopreservation was performed in a programmable freezing machine using one of two protocols. Method I (n = 133) used a one-step addition of cryoprotectant followed by a slow freeze and thaw protocol. With method II (n = 95), the cryoprotectant was added in a stepwise manner with cryopreservation performed in the presence of 0.2 M sucrose followed by rapid freezing and thawing. SETTING: Basic research center at a medical school. PATIENTS: Patients undergoing oophorectomy for nonovarian pathology. MAIN OUTCOME MEASURES: Rates of survival and maturation to metaphase II were compared between control oocytes and oocytes cryopreserved with methods I and II. RESULTS: With method I, a survival rate of 15.6% was obtained with 58.3% of surviving oocytes reaching metaphase II after culture compared with 50.0% of nonfrozen control oocytes. Method II produced a survival rate of 43.3% with 27.3% maturing to metaphase II. Maturation of control oocytes for method II was 46.4%. Although the survival rate with method II was significantly higher than with method I, the rate of in vitro maturation to metaphase II showed no difference. CONCLUSIONS: These results demonstrate that human prophase I oocytes obtained from unstimulated antral follicles are capable of meiotic maturation after cryopreservation.

Cell Survival↗
PubMed1992

A coiled-coil related protein specific for synapsed regions of meiotic prophase chromosomes.

Synaptonemal complexes (SCs) are structures that are formed between homologous chromosomes during meiotic prophase. They are probably involved in chromosome pairing and recombination. Using a monoclonal anti-SC antibody we isolated cDNAs encoding a major component of SCs which is localized specifically in synapsed segments of meiotic prophase chromosomes. The protein predicted from the nucleotide sequence of a full-length cDNA, named SCP1, consists of 946 amino acid residues and has a molecular weight of 111 kDa. It shares several features with nuclear lamins and some recently identified nuclear matrix proteins. The major part of SCP1 consists of long stretches capable of forming amphipathic alpha-helices. This region shows amino acid sequence similarity to the coiled-coil region of myosin heavy chain. A leucine zipper is included in this region. The carboxy-terminus has two small basic domains and several S/T-P-X-X motifs, which are characteristic of DNA-binding proteins. One of these motifs is a potential target site for p34cdc2 protein kinase. The amino-terminus is acidic and relatively proline-rich, but does not contain the S/T-P-X-X motif. The transcription of the gene encoding SCP1 is restricted to zygotene-diplotene spermatocytes. A polyclonal antiserum raised against the fusion protein of one of the cDNA clones recognizes a single protein on Western blots of isolated SCs, with an electrophoretic mobility identical to that of the antigen recognized by the original monoclonal antibody (mAb), IX5B2. From a detailed comparison of the immunogold labelling of rat SCs by mAb IX5B2 and the polyclonal anti-fusion protein antiserum respectively, we tentatively infer that the carboxy-terminus of SCP1 is orientated towards the lateral elements and that the other domains of the protein extend towards the central region between the lateral elements. We conclude that SCP1 is the major component of the transverse filaments of SCs, and speculate that it has evolved by specialization of a nuclear matrix protein.

Amino Acid Sequence↗
PubMed1979

Nucleolar organizers in human oocytes at meiotic prophase I, studied by the silver-NOR method and electron microscopy.

Use of the silver-NOR method to study the nucleolar organizers in human oocytes demonstrates that topographic and quantitative variations occur during meiotic prophase. In the oogonia nucleolus the nucleolar organizers are dispersed, whereas beginning at leptotene and throughout the remaining stages of meiotic prophase they occupy a marginal position in the nucleolus. At leptotene, a modal number of seven nucleolar organizers can be observed, whereas this number falls to 2.5 at pachytene and rises to ten at diplotene, thus showing that there is intense rRNA synthesis during the latter stage of meiosis. During pachytene, one end of the bivalents containing the ribosomal cistrons is always associated with the Ag-positive zone of the nucleolus. Observation of pachytene in the electron microscope shows that the secondary constriction region of D and G bivalents is constantly associated with the fibrillar center of the nucleolus. Comparison of these two methods of investigation reveals that the silver-stained regions of the nucleolus correspond to the fibrillar centers. The latter are surrounded by a layer of electron-dense fibrils corresponding to the zone of rDNA transcription. This electron-dense layer is absent during pachytene when the nucleolus displays spontaneous segregation of its components; this absence is related to temporary arrest of rDNA transcription. The affinity of the fibrillar centers for silver-NOR staining confirms that these structures contain ribosomal cistrons. During the diplotene stage, numerous micronucleoli are formed outside the nucleolar organizers of D and G chromosomes. Most of these micronucleoli present an Ag-positive granule on one of their margins, thus indicating that they contain an actively transcribed sequence of rDNA. This observation confirms the existence of amplification of ribosomal genes in the human oocyte.

Cell Nucleolus↗
PubMed1989

Nucleolar structures in chromosome and SC preparations from human oocytes at first meiotic prophase.

We describe a comparative study of the behavior of nucleolar structures and their relationship with nucleolar chromosomes and synaptonemal complexes at first meiotic prophase of human oocytes in an attempt to elucidate the nature of this cellular organization and to learn more about maternal nondisjunction. The number of main nucleoli varies along the different stages of prophase I and is usually low. It shows an increase from leptotene to pachytene and a decrease from pachytene to diplotene related to a decrease and an increase of main nucleoli volume, respectively. The methodology employed has enabled us to analyze in detail dark bodies, round bodies, dense bodies, and main nucleoli in chromosome or synaptonemal complex spreads. The relationship between nucleolar chromosomes or synaptonemal complexes and the nucleoli implies the existence, in a very reduced space, of chromosomal regions that contain homologous sequences and that are often unpaired. This situation may facilitate the production of heterologous pairing and chromosomal exchanges between nonhomologous chromosomes and finally result in aneuploidy. Thus, the situation explained above together with the differences between the oocyte and spermatocyte NOR cycles could be one of the reasons for the higher incidence of aneuploidies of maternal origin at meiosis I.

Cell Nucleolus↗
PubMed1979

The grasshopper X chromosome. I. States of condensation and the nuclear envelope at G1, S and G2 of premeiotic interphase and at early meiotic prophase.

The sub-stages of spermatocyte interphase (G1, S and G2) have been identified in the grasshopper Brachystola magna using E.M. autoradiography and serial thin sectioning techniques. The X chromosome occupies a nuclear envelope bound compartment separate from an autosome compartment during G1 and S. At G2 the X compartment is resolved by coalescence of the membranes enveloping the X chromosome and autosome compartments.--At G1 and S, the compartmentalized X chromatin is laced with nuclear membrane material. This X chromatin associated membrane decreases in amount as the cell passes through G2 and enters early meiotic prophase. There are at least 2 and possibly 3 states of condensation of the heterochromatic X during premeiotic interphase and early meiotic prophase correlated with the presence or absence of membrane material associated with the chromatin.--The X chromatin replicates asynchronously with autosomal euchromatin and synchronously with autosomal heterochromatin associated with nucleoli. The X chromatin replication appears to be associated with the nuclear membrane.--The observations indicate that the nuclear membrane is involved with X chromosome condensation and may be implicated in asynchronous X chromosome replication as well.

Animals↗
PubMed1976

Arrangement of chromosome ends and axial core formation during early meiotic prophase in the male grasshopper Brachystola magna by 3D, E.M. reconstruction.

Evidence is presented that chromosome ends are attached to the nuclear envelope prior to the formation of axial cores during early meiotic prophase in the grasshopper Brachystola magna. The attachment sites of distal and proximal chromosome ends are clustered in a small region of the inner nuclear envelope resulting in a classical bouquet arrangement of the chromosomes. Proximal ends are tightly clustered due to the presence of chromocenters. Distal chromosome ends are more widely scattered throughout the base of the bouquet.--Axial core formation can be initiated at chromosome ends or at internal chromosome sites. However, there is a preference for axial cores to form in distal chromosome regions rather than proximal regions during early meiotic prophase.--Virtually all of the nuclear pore complexes are located in the general vicinity of the chromosome attachment sites but each specific attachment site is surrounded by a small area of nuclear envelope which is devoid of pore complexes.

Animals↗
PubMed1980

Chromatin organization during meiotic prophase of Bombyx mori.

Chromatin organization during the early stages of male meiotic prophase in Bombyx mori was investigated by electron microscopy. The analysis of nuclei prepared by the Miller spreading procedure, suggests that chromatin fibers which are 200-300 A in diameter undergo an orderly folding coincident with the formation of the synaptonemal complex. In very early stages the chromatin is released in linear arrays typical of interphase chromatin material. With time loops containing 5-25 mu of B conformation DNA, initially visualized at the periphery of early meiotic prophase nuclei, aggregate into discrete foci. These foci coalesce to form the longitudinal axis of the chromosome in conjunction with the initial appearance of the axial elements of the synaptonemal complex. At pachytene, the loops are evenly distributed along the length of the chromosome and extend radially so that in well spread preparations the chromosome has a brush-like appearance. Throughout this period nascent RNP-fibers were visualized along some of the loops.

Animals↗
PubMed1997

Colchicine effects on meiosis in the male mouse. I. Meiotic prophase: synaptic arrest, univalents, loss of damaged spermatocytes and a possible checkpoint at pachytene.

Antimitotic agents administered at the time of synapsis (leptotene/zygotene) have been shown to induce synaptic abnormalities visible during pachytene in the male mouse. The object of this study was to test the hypothesis that cells with relatively large amounts of colchicine-induced damage to the synaptonemal complex (SC) are eliminated from prophase whereas cells with relatively small amounts of SC damage proceed through to the end of prophase. Male mice were injected with tritiated thymidine to mark a cohort of spermatocytes at premeiotic S-phase for tracking through pachytene. Forty-eight hours later, when those cells were at leptotene/zygotene, colchicine was administered intratesticularly. Whole-mount SC spreads were made from animals sacrificed at various times following colchicine administration, and prepared for autoradiography. The marked cells were examined by light and electron microscopy and the kind and number of synaptic abnormalities were scored throughout pachytene. Colchicine-induced SC damage included single axial elements (univalents), together with partially synapsed and nonhomologously synapsed SCs. The amount of SC damage (amount and type per cell and frequency of cells with damage) scored at early pachytene exceeded by three- to fivefold the amount at late pachytene. This is consistent with spermatogenic cell loss from the seminiferous tubule via colchicine-induced destruction of Sertoli cell microtubules. The presence of spermatocytes with no more than four autosomal univalents at late pachytene indicates that some cells with low amounts of synaptic damage progress to the end of pachytene. The loss of the most severely damaged cells may represent a meiotic checkpoint at early pachytene in the male mouse.

Animals↗
PubMed1997

A novel fission yeast gene, kms1+, is required for the formation of meiotic prophase-specific nuclear architecture.

In the meiotic prophase nucleus of the fission yeast Schizosaccharomyces pombe, chromosomes are arranged in an oriented manner: telomeres cluster in close proximity to the spindle pole body (SPB), while centromeres form another cluster at some distance from the SPB. We have isolated a mutant, kms1, in which the structure of the meiotic prophase nucleus appears to be distorted. Using specific probes to localize the SPB and telomeres, multiple signals were observed in the mutant nuclei, in contrast to the case in wild-type. Genetic analysis showed that in the mutant, meiotic recombination frequency was reduced to about one-quarter of the wild-type level and meiotic segregation was impaired. This phenotype strongly suggests that the telomere-led rearrangement of chromosomal distribution that normally occurs in the fission yeast meiotic nucleus is an important prerequisite for the efficient pairing of homologous chromosomes. The kms1 mutant was also impaired in karyogamy, suggesting that the kms1+ gene is involved in SPB function. However, the kms1+ gene is dispensable for mitotic growth. The predicted amino acid sequence of the gene product shows no significant similarity to known proteins.

Amino Acid Sequence↗
PubMed1988

Fanconi's anemia lymphocytes: effect of caffeine, adenosine and niacinamide during G2 prophase.

In this investigation peripheral blood lymphocytes from 3 Fanconi's anemia (FA) patients, 2 FA heterozygotes and 4 normal subjects were treated with caffeine and/or adenosine, and/or niacinamide during G2 prophase. Caffeine dramatically increased breakage levels in homozygote and heterozygote cells. Niacinamide and adenosine decreased the amount of chromosomal aberrations detected in FA homozygote and heterozygote lymphocytes treated and untreated with caffeine during G2 prophase. Caffeine sensitivity of heterozygote lymphocytes is proposed as a new clinical test to explore heterozygosis in individuals of FA families.

Adenosine↗
PubMed1983

Multiplication of nucleolar fibrillar centres and absence of rDNA amplification in mouse oocyte during meiotic prophase I.

During meiotic prophase I the nucleolus of the mouse oocyte assumes a reticulate structure of 'nucleolonema' type. This change coincides with the appearance of several secondary fibrillar centres. The number of these centres at diplotene (97-113), largely exceeds that of nucleolar organizers (4c DNA = 20 NORs). The quantitative analysis of autoradiographs after hybridization in situ with 3H-uridine labelled rRNA, enabled us to demonstrate that the multiplication of the fibrillar centres in mouse oocyte nucleolus during meiotic prophase I is not the result of an amplification of the rDNA. The number of silver grains in pachytene and diplotene nuclei was twice that counted for somatic cell and oogonium nuclei (2c DNA).

Animals↗
PubMed2006

Human Wapl is a cohesin-binding protein that promotes sister-chromatid resolution in mitotic prophase.

BACKGROUND: The linkage between duplicated chromosomes (sister chromatids) is established during S phase by the action of cohesin, a multisubunit complex conserved from yeast to humans. Most cohesin dissociates from chromosome arms when the cell enters mitotic prophase, leading to the formation of metaphase chromosomes with two cytologically discernible chromatids. This process is known as sister-chromatid resolution. Although two mitotic kinases have been implicated in this process, it remains unknown exactly how the cohesin-mediated linkage is destabilized at a mechanistic level. RESULTS: The wings apart-like (Wapl) protein was originally identified as a gene product that potentially regulates heterochromatin organization in Drosophila melanogaster. We show that the human ortholog of Wapl is a cohesin-binding protein that facilitates cohesin's timely release from chromosome arms during prophase. Depletion of Wapl from HeLa cells causes transient accumulation of prometaphase-like cells with chromosomes that display poorly resolved sister chromatids with a high level of cohesin. Reduction of cohesin relieves the Wapl-depletion phenotype, and depletion of Wapl rescues premature sister separation observed in Sgo1-depleted or Esco2-depleted cells. Conversely, overexpression of Wapl causes premature separation of sister chromatids. Wapl physically associates with cohesin in HeLa-cell nuclear extracts. Remarkably, in vitro reconstitution experiments demonstrate that Wapl forms a stoichiometric, ternary complex with two regulatory subunits of cohesin, implicating its noncatalytic function in inactivating cohesin's ability to interact with chromatin. CONCLUSIONS: Wapl is a new regulator of sister chromatid resolution and promotes release of cohesin from chromosomes by directly interacting with its regulatory subunits.

Carrier Proteins↗
PubMed1970

Duration of premeiotic deoxyribonucleic acid synthesis and the stages of prophase I in rabbit oocytes.

To estimate the duration of oocyte DNA synthesis 36, 3-day-old female rabbits received 3, 6, 9, 12, 15, or 18 injections of tritiated thymidine (thy-(3)H) at hourly intervals. The ovaries, removed at 1, 10, or 20 days after the first injection, were radioautographed. Counts made of the number of silver grains associated with oocyte nuclei in meiotic Prophase I indicate that the duration of DNA synthesis is between 9 and 12 hr. To determine the length of the stages of meiotic Prophase I, a group of 2-3-day-old rabbits was given a single sub-cutaneous injection of thy-(3)H, and the ovaries were removed at hourly and/or daily intervals after treatment. The minimum duration of leptotene was 3 hr and the maximum duration probably was less than 8 hr. The maximum durations of zygotene, pachytene, and diplotene were estimated to be 44, 216, and 96 hr, respectively. The interval from the end of oogonial DNA synthesis to the beginning ofpremeiotic DNA synthesis (G(2) + Mitosis + G(1)) appeared to be less than 6 hr.

Animals↗
PubMed1970

RNA synthesis in HeLa cells. Pattern in hypertonic medium and its similarity to synthesis during G2-prophase.

Interphase HeLa cells manifest a stepwise shutoff of RNA synthesis when the tonicity of the extracellular medium is gradually increased. Synthesis of heterogeneous nuclear RNA is most sensitive and is selectively inhibited at 1.5 times isotonicity (450 milliosmols/liter), while 45S ribosomal RNA synthesis is not affected significantly below 2.0 times isotonicity. Transfer RNA synthesis is least sensitive to increased osmolarity and is not completely inhibited until the electrolyte concentration of the medium is elevated to 2.8 times isotonicity. Although the transcription and methylation of 45S ribosomal precursor is unaffected at 1.5 times isotonicity, there is pronounced impairment of its processing into 32S and 18S RNA. Using a refined cell synchronization technique, we have been able to compare these effects of hypertonicity with the shutoff of RNA synthesis which occurs during the G(2)-prophase interval of the cell division cycle. In this case, as with random cells in hypertonic medium, a selective inhibition of heterogeneous nuclear RNA synthesis and slowed processing of 45S ribosomal RNA were found, whereas synthesis of 45S and transfer RNA continued unabated throughout G(2)-prophase. While it is known that RNA synthesis essentially ceases during metaphase, we have noted that transfer RNA synthesis continues in metaphase at 10-15% of the interphase rate, which is of particular interest in view of the relative resistance of this species to hypertonicity. The close correlation between the patterns of cessation of RNA synthesis at mitosis and during exposure to hypertonic medium supports our earlier contention that alteration of intracellular electrolyte levels provides a useful model for studying the mechanism of mitosis.

Carbon Isotopes↗