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PubMed1988

Synaptonemal complex damage induced by clastogenic and anti-mitotic chemicals: implications for non-disjunction and aneuploidy.

Mice were treated with mitomycin C, cyclophosphamide, amsacrine, colchicine, or vinblastine sulfate, and meiotic prophase cells analyzed for synaptonemal complex (SC) damage. All test agents caused synaptonemal complex breakage and synapsis irregularities, although propensities for inducing specific types of damage at S-phase or prophase stages varied among the chemicals. The data indicate that SC analysis can reveal chemical-specific alterations to meiotic homologue pairing/synapsis which have not generally been recognized, and which theoretically may be implicated in non-disjunction.

Aneuploidy↗
PubMed1988

Chromosomal preparations of human triploid zygotes and embryos fertilized in vitro.

Forty-eight zygotes with more than two pronuclei were identified after in vitro fertilization, representing 6.1% of all fertilized oocytes. The chromosome preparations from pronuclear stage to the cleaved human embryos were examined. Prophase was found in eight out of ten zygotes. The spreading of chromosomes allowed an adequate counting in only two cases. Six of the eight preparations displayed a late prophase. In this stage each haploid group of chromosomes can be analysed separately. Kariogamy usually occurred 4 to 5 h after the pronuclei had disappeared, and polyploid number of chromosomes were found in well-spread metaphases. The chromosomal preparations were made for eleven human embryos arising from zygotes with three pronuclei. Out of ten preparations, where the chromosomes could be counted, seven embryos (70%) contained hypodiploidic groups of chromosomes. In two of the cases, however, triploid metaphases were found, and in the last one a triploid/diploid mosaicism.

Embryo, Mammalian↗
PubMed1994

Acute effects of rat growth hormone (GH), human GH and prolactin on proliferating rat liver cells in vitro: a study of mitotic behaviour and ultrastructural alterations.

Examination of livers from transgenic mice over expressing human growth hormone (hGH) revealed numerous alterations including a striking incidence of mitotic figures. The reason for increased proliferation is unclear, but could be related to effects of hGH, which is also acting as a lactogen in rodents. In order to identify some of the actions of GH, we have examined the effects of rat and human GH and rat prolactin on proliferation, as well as on morphological differentiation of normal rat liver cells in vitro. These cells, isolated from a 20-day-old rat, proliferate in culture, incorporate BrdU and are thus strikingly different from primary cultures of isolated hepatocytes, which typically are non-proliferating cells. Monolayers of these cells were treated with rat prolactin (rPRL), rat growth hormone (rGH), rPRL and rGH in combination, or hGH, for 24 hr. Subsequently, mitotic figures were counted and the cultures were processed for transmission electron microscopy. The incidence of mitotic figures was significantly increased by rPRL (27.4%) versus control (19%), while rGH (13%) and hGH (9.6%) significantly decreased proliferation. In controls, 2% of the proliferating cells were in prophase, approximately 12% in metaphase and approximately 15% in telophase. In contrast, rPRL caused a significant increase in the number of cells in prophase (14%) and reduced the number of cells in the other mitotic stages. hGH and rGH reduced the overall number of mitotic figures. Unexpectedly, the effects of rGH plus rPRL were different from the effect of hGH. In addition, each treatment caused distinct morphological changes of liver cell organelles.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗
PubMed1993

Multiple chromosomal populations of topoisomerase II detected in vivo by time-lapse, three-dimensional wide-field microscopy.

The localization of topoisomerase II (topo II) in vivo was studied by recording time-lapse, three-dimensional data sets of living Drosophila melanogaster embryos injected with rhodamine-labeled topo II. These images show that topo II is concentrated at specific sites within the interphase nucleus and that this localization is temporally regulated. The enzyme is not restricted to a central chromosome axis, but is distributed uniformly throughout the chromosome. During mitosis, the enzyme present in the early prophase chromosome is lost in two stages, following prophase and following anaphase. Overall, 70% of the enzyme leaves the nucleus and diffuses into the cytoplasm. The localization of the enzyme thus correlates with its role in chromosome condensation and segregation. Rather than being solely a structural protein, topo II appears to localize at the sites on the chromosome where it is required.

Animals↗
PubMed1982

Retinoblastoma and its association with a deletion in chromosome #13: a survey using high-resolution chromosome techniques.

To our knowledge, 20 cases of retinoblastoma associated with a chromosome #13 aberration have been reported. The present study utilized high-resolution prophase banding analysis of 12 additional retinoblastoma patients to determine the occurrence of chromosome aberrations and identify consistently associated clinical abnormalities. Six male and six female patients were studied representing seven cases of bilateral and five cases of unilateral retinoblastoma. One case of unilateral and two cases of bilateral retinoblastoma and detectable cytogenetic abnormalities, all involving an interstitial deletion of 13q14 on the long arm of one chromosome #13. In all five unilateral cases the tumor manifested in the left orbit, and in all seven bilateral cases the left eye was at a more malignantly advanced stage than the right eye. All three cases with a chromosome abnormality had varying degrees of developmental and/or mental retardation, along with at least one other congenital abnormality. In addition to the 12 cases of retinoblastoma, a patient with severe ophthalmologic abnormalities and mild congenital anomalies was studied by the prophase banding technique and found to be partially trisomic for the 13q14 region with gene loci for optic development and indicate that cytogenetic abnormalities may occur even more frequently in retinoblastoma than indicated by the small number of cases reported in the literature.

Child, Preschool↗
PubMed1984

Mitotic modifications and aberrations in human cervical cancer.

Mitotic modifications and aberrations characteristic of human malignant tumors have been analyzed and illustrated in cervical cancer. Most of them can be explained by assuming that the coordination of the centrosomal and chromosomal mechanisms, typical of normal mitosis, is disturbed. When the spindle mechanism is ahead of the chromosomes, the prophase is relatively shortened. This expresses itself in an increase of the ratio of metaphases to prophases (M/P), which in normal tissues is around 1. With M/P values of 4-6, the first tripolar metaphases are formed, and with higher ratios, divisions having more and more poles appear. The spindle and the chromosomes are out of step in the opposite direction in endocycles, in which the spindle is slowed down or absent. The most common of the endocycles is endoreduplication, followed by endomitosis, which is found in more than half of the cervical cancers. Mitotic abnormalities include lagging chromosomes in metaphase and anaphase and bridges in anaphase, which, when numerous, may lead to restitution. More sporadically occurring are C-mitosis and other abnormalities, including cell and nuclear fusions. There is a wide range of variation in the occurrence and frequency of chromocenters within a tumor, and an even greater variation between tumors. About one-fifth of cervical cancers lack X chromatin bodies. The abnormal chromosome constitutions in cancer are the result of various mitotic modifications and aberrations, as well as chromosome rearrangements. New chromosome combinations are constantly created and selection promotes the fastest dividing ones, which, in turn, become new stem lines of the tumor.

Chromatin↗
PubMed1991

Clastogenic effects of hydroquinone: induction of chromosomal aberrations in mouse germ cells.

The clastogenic activity of hydroquinone (HQ) in germ cells of male mice was evaluated by analysis of chromosomal aberrations in primary spermatocytes and differentiating spermatogonia. In the first experiment with treated spermatocytes the most sensitive stage of meiotic prophase to aberration induction by HQ was determined. Testicular material was sampled for microscopic analysis of cells in diakinesis-metaphase I at 1, 5, 9, 11, and 12 days after treatment with 80 mg/kg of HQ, corresponding to treated diplotene, pachytene, zygotene, leptotene and preleptotene. The frequencies of cells with structural chromosome aberrations peaked at 12 days after treatment (p less than 0.01). This indicates that the preleptotene when DNA synthesis occurred was the most sensitive stage of meiotic prophase. In the second experiment the dose response was determined 12 days post treatment by applying 2 additional doses of 40 mg/kg and 120 mg/kg. The clastogenic effects induced by 40 and 80 mg/kg were significantly different from the controls (p less than or equal to 0.01) and higher than the results obtained with 120 mg/kg of HQ. A humped dose-effect relationship was observed. In a third experiment the same doses were used to analyse chromosomal aberrations in dividing spermatogonia of mice 24 h after treatment with HQ. All the administered doses gave results statistically different from the control values (p less than or equal to 0.01) and the data were fitted to a linear equation. HQ was found to be clastogenic in male mouse germ cells. It is concluded that the clastogenic effect in male germ cells is of the same order of magnitude as in mouse bone marrow cells.

Animals↗
PubMed1992

Kinetochore-staining of spermatid micronuclei: studies of mice treated with X-radiation or acrylamide.

The rodent spermatid micronucleus (MN) assay was used in conjunction with immunofluorescent techniques to distinguish kinetochores in MN following exposure of mice to X-radiation or acrylamide. After either treatment, modest increases in kinetochore-positive MN were observed. Spermatids which had been exposed during meiotic prophase to X-rays (400 cGy) had approximately 10-fold increases in MN compared to controls; up to 15% of the MN observed were kinetochore-positive. Following acrylamide treatment of meiotic prophase cells, there was a doubling of spermatid MN over baseline levels, approximately one-third of which were kinetochore-positive.

Acrylamide↗
PubMed1992

Polyphosphoinositide synthesis and protein phosphorylation in the plasma membrane from full-grown Bufo arenarum oocytes.

1. Polyphosphoinositide content and phosphorylation of lipids and proteins were analyzed in oocytes of the toad Bufo arenarum Hensel. 2. Plasma membrane-enriched fractions obtained from full-grown, prophase-arrested oocytes incorporated 32P into both phospholipids and proteins after incubation with [gamma-32P]ATP in an Mg(2+)-containing medium. Phosphatidylinositol 4-phosphate (PIP), phosphatidate (PA) and phosphatidylinositol-4,5-bisphosphate (PIP2) were the only labelled lipids. The 32P incorporation depended on incubation time, the amount of protein, and the ATP concentration. 3. Autoradiography of polyacrylamide gel electropherograms and scintillation counting showed that the radioactivity was mainly associated with a group of membrane proteins having an M(r) of 87,000. 4. This paper provides evidence for the capacity of prophase-arrested oocytes from Bufo arenarum to synthesize polyphosphoinositides and to phosphorylate distinct membrane proteins.

Adenosine Triphosphate↗
PubMed1987

Antipain microinjection prevents progesterone to inhibit adenyl cyclase in Xenopus oocytes.

Microinjection of antipain, an inhibitor of thiol and Ca2+-dependent proteases, in immature Xenopus oocytes inhibited meiotic maturation induced by progesterone, but not by transfer of cytoplasm taken from maturing oocytes. Oocytes could be released from antipain inhibition by increasing progesterone concentration. alpha-32P-ATP was microinjected to study adenylcyclase in ovo. As already reported, neosynthesis of cAMP was decreased following progesterone application. This decrease was not observed, or it was considerably reduced, in oocytes previously injected with antipain. In amphibian, full-grown ovarian oocytes are arrested at first meiotic prophase, and have a large nucleus known as the germinal vesicle. Progesterone induces the production of a cytoplasmic maturation-promoting factor (MPF), which itself triggers germinal vesicle breakdown (GVBD), and subsequent events of meiotic maturation (Masui and Markert, 1971; Gerhart et al., 1984). A considerable body of evidences support the view that release from prophase block is due to inactivation of a cAMP-dependent protein kinase (reviewed by Maller, 1983). On the other hand, progesterone has been shown to induce a transient decrease in cAMP level (Speaker and Butcher, 1977; Schorderet-Slatkine et al., 1982; Cicirelli et al., 1985), and this initial drop of cAMP, along with a number of studies indicating a decrease in adenylate cyclase activity (Mulner et al., 1979; Baltus et al., 1981; Sadler and Maller, 1981; Finidori-Lepicard et al., 1981; Jordana et al., 1981), provided key support to the theory that an early drop in cAMP led to the dephosphorylation of a hypothetical protein which initiates maturation.(ABSTRACT TRUNCATED AT 250 WORDS)

1-Methyl-3-isobutylxanthine↗
PubMed2006

Characterization of Drosophila Rad51/SpnA protein in DNA binding and embryonic development.

The Rad51 is a highly conserved protein throughout the eukaryotic kingdom and an essential enzyme in DNA repair and recombination. It possesses DNA binding activity and ATPase activity, and interacts with meiotic chromosomes during prophase I of meiosis. Drosophila Rad51, Spindle-A (SpnA) protein has been shown to be involved in repair of DNA damage in somatic cells and meiotic recombination in female germ cells. In this study, DNA binding activity of SpnA is demonstrated by both agarose gel mobility shift assay and restriction enzyme protection assay. SpnA is also shown to interact with meiotic chromosomes during prophase I in the primary spermatocytes of hsp26-spnA transgenic flies. In addition, SpnA is highly expressed in embryos, and the depletion of SpnA by RNA interference (RNAi) leads to embryonic lethality implying that SpnA is involved in early embryonic development. Therefore, these results suggest that Drosophila SpnA protein possesses properties similar to mammalian Rad51 homologs.

Animals↗
PubMed2005

Shugoshin: guardian spirit at the centromere.

A recently emerging protein family, shugoshin, plays a crucial role in the centromeric protection of cohesin, which is responsible for sister chromatid cohesion. This is especially important at the first meiotic division, where cohesin is cleaved by separase only along chromosome arms while the centromeric cohesin must be preserved. In vertebrate cells, arm cohesion is largely lost during prophase and prometaphase in order to facilitate sister chromatid resolution, whereas centromeric cohesion is preserved until the bipolar attachment of sister chromatids is established. Vertebrate shugoshin plays an essential role in protecting centromeric cohesin from prophase dissociation. In yeast, shugoshin also has a crucial role in sensing the loss of tension at kinetochores and in generating the spindle checkpoint signal.

Animals↗
PubMed2003

The Sgs1 helicase regulates chromosome synapsis and meiotic crossing over.

BACKGROUND: In budding yeast, Sgs1 is the sole member of the RecQ family of DNA helicases. Like the human Bloom syndrome helicase (BLM), Sgs1 functions during both vegetative growth and meiosis. The sgs1 null mutant sporulates poorly and displays reduced spore viability. RESULTS: We have identified novel functions for Sgs1 in meiosis. Loss of Sgs1 increases the number of axial associations, which are connections between homologous chromosomes that serve as initiation sites for synaptonemal complex formation. In addition, mutation of SGS1 increases the number of synapsis initiation complexes and increases the rate of chromosome synapsis. Loss of Sgs1 also increases the number of meiotic crossovers without changing the frequency of gene conversion. The sgs1 defect in sporulation is due to checkpoint-induced arrest/delay at the pachytene stage of meiotic prophase. A non-null allele of SGS1 that specifically deletes the helicase domain is defective in the newly described meiotic functions of Sgs1, but wild-type for most vegetative functions and for spore formation. CONCLUSIONS: We have shown that the helicase domain of Sgs1 serves as a negative regulator of meiotic interchromosomal interactions. The activity of the wild-type Sgs1 protein reduces the numbers of axial associations, synapsis initiation complexes, and crossovers, and decreases the rate of chromosome synapsis. Our data argue strongly that axial associations marked by synapsis initiation complexes correspond to sites of reciprocal exchange. We propose that the Sgs1 helicase prevents a subset of recombination intermediates from becoming crossovers, and this distinction is made at an early stage in meiotic prophase.

Chromosome Pairing↗
PubMed2006

PP2A is required for centromeric localization of Sgo1 and proper chromosome segregation.

Loss of sister-chromatid cohesion triggers chromosome segregation in mitosis and occurs through two mechanisms in vertebrate cells: (1) phosphorylation and removal of cohesin from chromosome arms by mitotic kinases, including Plk1, during prophase, and (2) cleavage of centromeric cohesin by separase at the metaphase-anaphase transition. Bub1 and the MEI-S332/Shugoshin (Sgo1) family of proteins protect centromeric cohesin from mitotic kinases during prophase. We show that human Sgo1 binds to protein phosphatase 2A (PP2A). PP2A localizes to centromeres in a Bub1-dependent manner. The Sgo1-PP2A interaction is required for centromeric localization of Sgo1 and proper chromosome segregation in human cells. Depletion of Plk1 by RNA interference (RNAi) restores centromeric localization of Sgo1 and prevents chromosome missegregation in cells depleted of PP2A_Aalpha. Our findings suggest that Bub1 targets PP2A to centromeres, which in turn maintains Sgo1 at centromeres by counteracting Plk1-mediated chromosome removal of Sgo1.

Cell Cycle Proteins↗
PubMed2007

Effects of luteinizing hormone treatment on oogenesis in ovarian germ cells of the chick (Gallus domesticus).

The effect of the luteinizing hormone (LH) on the oogenesis of ovaries from newly-hatched chicks treated in vivo on days 13, 15, and 17 of embryonic development was analyzed. Changes in oogonial proliferation, meiotic prophase, degeneration of germ cells, and primordial follicular organization were determined. Results indicate that the total number of germ cells was not affected by the LH treatment, but significant differences existed in the number of oogonia and oocytes between the ovaries of control and LH-treated chicks. LH treatment increased the percentage of oocytes and diminished the percentage of oogonia. The mitotic activity of oogonia and degeneration of germ cells decreased, but the number of follicles during development increased in LH-treated ovaries. These findings suggest that LH treatment might trigger a cascade of endocrine events, resulting in inhibition of oogonial proliferation and induction of the meiotic prophase and follicle formation.

Animals↗
PubMed2006

Entamoeba histolytica: ultrastructure of the chromosomes and the mitotic spindle.

We have analyzed by transmission electron microscopy the mitotic process of Entamoeba histolytica trophozoites in an asynchronous population of axenically cultured parasites. Our observations showed that nuclear microtubules, initially located at random in the karyosome during prophase, formed in subsequent stages a mitotic spindle closely related to the nuclear membrane at the polar regions of dividing nuclei. In late prophase and in anaphase, chromosomes appeared as dense bodies 0.1-0.5 microm. At least 15 chromosomes appeared in favorable planes of section, arranged as an incomplete elliptical circle, in close contact with microtubules. There was no morphological evidence of structures resembling the kinetochores of higher eukaryotes. When cut in cross-section, the mitotic spindle was made of 28-35 microtubular rosette assemblies. The latter probably correspond to a similar number of chromosomes, as has been shown by others with pulse-field electrophoresis and fluorescence microscopy of trophozoite spreads. In turn, each microtubular rosette was constituted by 7-12 parallel microtubules. In later stages of the metaphase, two sets of chromosomes were disposed forming a pair of elliptical circles. An additional finding in the dividing nuclei of E. histolytica trophozoites was the presence of compact conglomerates of numerous particles 50 nm in diameter, of similar electron density, shape, and size, probably corresponding to RNA episomes.

Animals↗
PubMed2006

Abnormal progression through meiosis in men with nonobstructive azoospermia.

OBJECTIVE: To study meiotic abnormalities in men with nonobstructive azoospermia. DESIGN: Analysis of synaptonemal complex and recombination in testicular tissue. SETTING: Research laboratory. PATIENT(S): Twenty-nine men with nonobstructive azoospermia and 12 men with normal spermatogenesis. INTERVENTION(S): Testicular tissues were processed with immunofluorescent staining using antibodies against proteins associated with synaptonemal complex and recombination events. MAIN OUTCOME MEASURE(S): Synaptonemal complex configuration and recombination in meiosis I. RESULT(S): In patients with nonobstructive azoospermia, a marked heterogeneity in spermatogenesis was found: nearly half of them had a complete absence of meiotic cells, one case had germ cells arrested at the zygotene stage of meiotic prophase, and, in general, the rest had impaired fidelity of chromosome synapsis and recombination in pachytene cells. Compared with controls, these patients had significantly more cells in leptotene/zygotene and higher frequencies of unpaired chromosome regions in pachytene. Significantly reduced recombination, an increased frequency of achiasmate autosome bivalents, and sex univalents in pachytene were also observed in these patients with nonobstructive azoospermia. CONCLUSION(S): Defects in chromosome synapsis and decreased recombination during meiotic prophase may have led to spermatogenesis arrest and contributed in part to the unexplained infertility in these patients.

Adaptor Proteins, Signal Transducing↗
PubMed2005

Shugoshin, a guardian for sister chromatid segregation.

To ensure sister chromatids to be equally transmitted to daughter cells, it is imperative that physical association of sister chromatids is maintained during S, G2, and early mitosis until the onset of anaphase. Cohesion of sister chromatids in eukaryotes is largely achieved by the cohesin complex. In vertebrates, cohesin molecules are dissociated from chromosome arms but not from centromeres during prophase by the so-called prophase pathway. Although it remains unclear what is the molecular basis by which centromeric cohesin is retained, a flurry of recent studies have shed light on a family of proteins named Shugoshin (Sgo) that are evolutionarily conserved across eukaryotes. Sgo1 functions as a protector of centromeric cohesin during meiosis in yeast and during mitosis in high eukaryotes. Suppression of Sgo1 function results in premature separation of sister chromatids in both meiosis and mitosis. The discovery of members of the Sgo family may help to explain how centromeric cohesin is protected from dissociation from DNA until the onset of anaphase. Given the importance of chromosome cohesion in the maintenance of genomic stability, further characterization of Sgo1 and related molecules may also open up new avenues of research for developing new strategies for cancer intervention.

Animals↗