Pachytene Nuclei Research Articles

The nuclear Argonaute HRDE-1 directs target gene re-localization and shuttles to nuage to promote small RNA-mediated inherited silencing

Argonaute/small RNA pathways and heterochromatin work together to propagate transgenerational gene silencing, but the mechanisms behind their interaction are not well understood. Here, we show that induction of heterochromatin silencing in C.elegans by RNAi or by artificially tethering pathway components to target RNA causes co-localization of targetalleles in pachytene nuclei. Tethering the nuclear Argonaute WAGO-9/HRDE-1 induces heterochromatin formation and independently induces small RNA amplification. Consistent with this finding, HRDE-1, while predominantly nuclear, also localizes to peri-nuclear nuage domains, where amplification is thought to occur. Tethering a heterochromatin-silencing factor, NRDE-2, induces heterochromatin formation, which subsequently causes de novo synthesis of HRDE-1 guide RNAs. HRDE-1 then acts to further amplify small RNAs that load on downstream Argonautes. These findings suggest that HRDE-1 plays a dual role, acting upstream to initiate heterochromatin silencing and downstream to stimulate a new cycle of small RNA amplification, thus establishing a self-enforcing mechanism that propagates gene silencing to future generations.

Cytogenetic Analysis of the Bimodal Karyotype of the Common European Adder, Vipera berus (Viperidae).

Vipera berus is the species with the largest range of snakes on Earth and one of the largest among reptiles in general. It is also the only snake species found in the Arctic Circle. Vipera berus is the most involved species of the genus Vipera in the process of interspecific hybridization in nature. The taxonomy of the genus Vipera is based on molecular markers and morphology and requires clarification using SC-karyotyping. This work is a detailed comparative study of the somatic and meiotic karyotypes of V. berus, with special attention to DNA and protein markers associated with synaptonemal complexes. The karyotype of V. berus is a remarkable example of a bimodal karyotype containing both 16 large macrochromosomes and 20 microchromosomes. We traced the stages of the asynchronous assembly of both types of bivalents. The number of crossing-over sites per pachytene nucleus, the localization of the nucleolar organizer, and the unique heterochromatin block on the autosomal bivalent 6-an important marker-were determined. Our results show that the average number of crossing-over sites per pachytene nucleus is 49.5, and the number of MLH1 sites per bivalent 1 reached 11, which is comparable to several species of agamas.

Homogeneously Staining Regions (HSR) in Chromosome 1 of the House Mouse: Synapsis and Recombination at Meiosis

Amplified sequences constitute a large part of mammalian genomes. A chromosome 1 containing 2 large (up to 50 Mb) homogeneously staining regions (HSRs) separated by a small inverted euchromatic region is present in many natural populations of the house mouse (Mus musculus musculus). The HSRs are composed of a long-range repeat cluster, Sp100-rs, with a repeat length of 100 kb. In order to understand the organization and function of HSRs in meiotic chromosomes, we examined synapsis and recombination in male mice hetero- and homozygous for the HSR-carrying chromosome using FISH with an HSR-specific DNA probe and immunolocalization of the key meiotic proteins. In all homozygous and heterozygous pachytene nuclei, we observed fully synapsed linear homomorphic bivalents 1 marked by the HSR FISH probe. The synaptic adjustment in the heterozygotes was bilateral: the HSR-carrying homolog was shortened and the wild-type homolog was elongated. The adjustment was reversible: desynapsis at diplotene was accompanied by elongation of the HSRs. Immunolocalization of H3K9me2/3 indicated that the HSRs in the meiotic chromosome retained the epigenetic modification typical for C-heterochromatin in somatic cells. MLH1 foci, marking mature recombination nodules, were detected in the proximal HSR band in heterozygotes and in both HSR bands of homozygotes. Unequal crossing over within the long-range repeat cluster can cause variation in size of the HSRs, which has been detected in the natural populations of the house mouse.

Study of Sertoli cells and spermatogenic cells in rats with experimentally induced metabolic syndrome and after balneophysiotherapy

Introduction. Metabolic syndrome (MS) can cause impaired spermatogenesis and a decrease in sperm counts. However, the details of the effect of MS on developing spermatogenic cells remain unclear. Difficulties in solving this problem, the inconsistency of published clinical data, indicate the advisability of using experimental models to solve this urgent problem of andrology and reproductology.The study objective is to describe to investigate the specifics of the course of meiotic prophase I and the activity of the processes of phagocytosis and autophagy in Sertoli cells of rats with experimentally induced MS and in the course of therapeutic and prophylactic procedures during the development of experimental MS.Materials and methods. The animals were divided into three groups, each of which included four sexually mature male rats: 1st group – males fed a standard diet; 2nd group – males receiving a diet high in fat and fructose for 60 days; 3rd group – males with MS receiving sulphate mineral waters therapy, low-intensity ultrahigh frequency electromagnetic radiation therapy. Testicular cells were examined using light and transmission electron microscopy. For the first time in animals with MS, an immunocytochemical study of the peculiarities of chromosome synapsis in prophase I of meiosis was carried out on the basis of analysis of spread synaptonemal complexes of meiotic chromosomes and immunocytochemical analysis of Sertoli cells and spermatogenic cells in squashed preparations of seminiferous tubules. The parametric Student’s t-test and the nonparametric Mann–Whitney U-test were used for statistical data processing.Results. As a result of a histological study of the structure of the seminiferous tubules of animals of three groups, a statistically significant decrease in the indices of the spermatogenesis index in 2nd and 3rd groups compared to the control was revealed. Immunomorphologically, in the spread nuclei of primary spermatocytes of rats of the 2nd and 3rd groups, violations of the architectonics of nuclei, the formation of synaptonemal complexes fragments and circular synaptonemal complexes, numerous atypical inclusions were found. Signs of pachytene arrest were found in 40–50 % of spermatocyte nuclei. In the study of squashed cells preparations of the seminiferous tubules of rats of the 2nd and 3rd groups, signs of phagocytosed synaptonemal complexes were found in the cytoplasm of Sertoli cells, which were confirmed using antibodies to the SCP3 protein. Thus, evidence for the phagocytosis of degenerating primary spermatocytes by Sertoli cells has been obtained. In Sertoli cells, spermatocytes and spermatids, many autophagosomes are found, using LC3B protein marker. The presence of autophagosomes in Sertoli cells and spermatogenic cells in animals of these two groups was also confirmed by electron microscopy. In male rats of the 2nd group, significant disturbances in the structure of the pachytene nuclei were revealed. In the cytoplasm of Sertoli cells and spermatids of rats of the 2nd group, lipid droplets, numerous phagolysosomes containing cell detritus were revealed. Structural damage and phagocytosis of mitochondria were found in Sertoli cells and spermatocytes. Аutophagy in Sertoli cells were most distinctive in animals of the 3rd group.Conclusion. In male rats with experimental MS, significant disturbances in the structure of the nuclei of meiotic cells, a high content of primary spermatocytes with signs of pachytene arrest were revealed. The results obtained are in good agreement with the data of other authors, who revealed a decrease in the number of spermatozoa in the epididymis of rats and mice when modeling MS. It is assumed that the activation of autophagy is an important factor in supporting the viability of Sertoli cells and supporting the viability of germ cells in stressful situations, including MS. Apparently, autophagy is an adaptive mechanism that removes the remnants of apoptotic spermatogenic cells that are selected as a result of MS development.

Meiotic synapsis of homeologous chromosomes and mismatch repair protein detection in the parthenogenetic rock lizard Darevskia unisexualis.

Parthenogenetic species of Caucasian rock lizards of the genus Darevksia are important evidence for reticulate evolution and speciation by hybridization in vertebrates. Female-only lineages formed through interspecific hybridization have been discovered in many groups. Nevertheless, critical mechanisms of oogenesis and specifics of meiosis that provide long-term stability of parthenogenetic species are still unknown. Here we report cytogenetic characteristics of somatic karyotypes and meiotic prophase I nuclei in the diploid parthenogenetic species Darevskia unisexualis from the new population "Keti" in Armenia which contains an odd number of chromosomes 2n = 37, instead of the usual 2n = 38. We revealed 36 acrocentric chromosomes and a single metacentric autosomal chromosome, resulting from Robertsonian translocation. Comparative genomic hybridization revealed that chromosome fusion occurred between two chromosomes inherited from the maternal species, similar to another parthenogenetic species D. rostombekowi. To trace the chromosome behaviour in meiosis, we performed an immunocytochemical study of primary oocytes' spread nuclei and studied chromosome synapsis during meiotic prophase I in D. unisexualis based on analysis of synaptonemal complexes (SCs). We found meiotic SC-trivalent composed of one metacentric and two acrocentric chromosomes. We confirmed that the SC was assembled between homeologous chromosomes inherited from two parental species. Immunostaining of the pachytene and diplotene nuclei revealed a mismatch repair protein MLH1 loaded to all autosomal SC bivalents. Possible mechanisms of meiotic recombination between homeologous chromosomes are discussed.

Maintenance of Genome Integrity by Mi2 Homologs CHD-3 and LET-418 in Caenorhabditis elegans.

Meiotic recombination depends upon the tightly coordinated regulation of chromosome dynamics and is essential for the production of haploid gametes. Central to this process is the formation and repair of meiotic double-stranded breaks (DSBs), which must take place within the constraints of a specialized chromatin architecture. Here, we demonstrate a role for the nucleosome remodeling and deacetylase (NuRD) complex in orchestrating meiotic chromosome dynamics in Caenorhabditis elegans Our data reveal that the conserved Mi2 homologs Chromodomain helicase DNA-binding protein (CHD-3) and its paralog LET-418 facilitate meiotic progression by ensuring faithful repair of DSBs through homologous recombination. We discovered that loss of either CHD-3 or LET-418 results in elevated p53-dependent germ line apoptosis, which relies on the activation of the conserved checkpoint kinase CHK-1 Consistent with these findings, chd-3 and let-418 mutants produce a reduced number of offspring, indicating a role for Mi2 in forming viable gametes. When Mi2 function is compromised, persisting recombination intermediates are detected in late pachytene nuclei, indicating a failure in the timely repair of DSBs. Intriguingly, our data indicate that in Mi2 mutant germ lines, a subset of DSBs are repaired by nonhomologous end joining, which manifests as chromosomal fusions. We find that meiotic defects are exacerbated in Mi2 mutants lacking CKU-80, as evidenced by increased recombination intermediates, corpses, and defects in chromosomal integrity. Taken together, our findings support a model wherein the C. elegans Mi2 complex maintains genomic integrity through reinforcement of a chromatin landscape suitable for homology-driven repair mechanisms.

Meiotic recombination modulates the structure and dynamics of the synaptonemal complex during C. elegans meiosis.

During meiotic prophase, a structure called the synaptonemal complex (SC) assembles at the interface between aligned pairs of homologous chromosomes, and crossover recombination events occur between their DNA molecules. Here we investigate the inter-relationships between these two hallmark features of the meiotic program in the nematode C. elegans, revealing dynamic properties of the SC that are modulated by recombination. We demonstrate that the SC incorporates new subunits and switches from a more highly dynamic/labile state to a more stable state as germ cells progress through the pachytene stage of meiotic prophase. We further show that the more dynamic state of the SC is prolonged in mutants where meiotic recombination is impaired. Moreover, in meiotic mutants where recombination intermediates are present in limiting numbers, SC central region subunits become preferentially stabilized on the subset of chromosome pairs that harbor a site where pro-crossover factors COSA-1 and MutSγ are concentrated. Polo-like kinase PLK-2 becomes preferentially localized to the SCs of chromosome pairs harboring recombination sites prior to the enrichment of SC central region proteins on such chromosomes, and PLK-2 is required for this enrichment to occur. Further, late pachytene nuclei in a plk-2 mutant exhibit the more highly dynamic SC state. Together our data demonstrate that crossover recombination events elicit chromosome-autonomous stabilizing effects on the SC and implicate PLK-2 in this process. We discuss how this recombination-triggered modulation of SC state might contribute to regulatory mechanisms that operate during meiosis to ensure the formation of crossovers while at the same time limiting their numbers.

Distribution of MLH1 Foci in Horse Male Synaptonemal Complex

Advances in molecular cytogenetics have provided the opportunity to study events during prophase I of meiosis. Immunofluorescent localization of different meiotic protein components were used to characterize the early stages of the first meiotic division in horse spermatocytes. The frequency and distribution of recombination events during prophase I were investigated using the mutL homolog 1 (MLH1) protein that is known to be associated with these events. The frequency and distribution of MLH1 foci were investigated in pachytene nuclei of 6 fertile stallions, and the average relative synaptonemal complex length was found to be highly correlated with the average number of MLH1 foci. The frequency and distribution of MLH1 foci were found to closely correspond to the frequency and distribution of chiasmata on metaphase I chromosomes, and genetic length, calculated from MLH1 foci data, for the whole genome was 2,505.5 cM.

Pro-crossover factors regulate damage-dependent apoptosis in the Caenorhabditis elegans germ line

During meiosis, DNA double-strand breaks (DSBs) are physiologically induced to start the recombination process and promote the formation of interhomologue crossovers (COs), which are required to ensure faithful chromosome segregation into the gametes. The timely repair of DSBs is an essential part of the meiotic programme, as accumulation of unprocessed DSBs during the pachytene stage of meiotic prophase triggers a DNA damage checkpoint response that induces apoptosis of damaged cells. We show that CO-promoting factors MSH-4, MSH-5, and ZHP-3, but not COSA-1, are required for the apoptotic response of the meiotic DNA damage checkpoint. Lack of MSH-4 or MSH-5 suppresses the apoptotic response observed in some DNA repair-defective mutants such as fcd-2 and brc-1 (orthologues of FANCD2 and BRCA1), irrespectively of the amount of DSBs present in pachytene nuclei. Although ionizing radiation fails to induce apoptosis in msh-4/5-mutant backgrounds, it induces transcriptional activation of the apoptosis-activator egl-1, which is controlled by the Caenorhabditis elegans p53 orthologue CEP-1. This finding suggests that MSH-4/5 involvement in the apoptotic response occurs downstream or independently of damage sensing and checkpoint activation. This study establishes a role for pro-CO factors MSH-4/5 and ZHP-3 in the execution of apoptosis at late meiotic prophase following the accumulation of exogenous or endogenous DNA damage.

Quantitative high resolution mapping of HvMLH3 foci in barley pachytene nuclei reveals a strong distal bias and weak interference.

In barley (Hordeum vulgare L.), chiasmata (the physical sites of genetic crossovers) are skewed towards the distal ends of chromosomes, effectively consigning a large proportion of genes to recombination coldspots. This has the effect of limiting potential genetic variability, and of reducing the efficiency of map-based cloning and breeding approaches for this crop. Shifting the sites of recombination to more proximal chromosome regions by forward and reverse genetic means may be profitable in terms of realizing the genetic potential of the species, but is predicated upon a better understanding of the mechanisms governing the sites of these events, and upon the ability to recognize real changes in recombination patterns. The barley MutL Homologue (HvMLH3), a marker for class I interfering crossovers, has been isolated and a specific antibody has been raised. Immunolocalization of HvMLH3 along with the synaptonemal complex transverse filament protein ZYP1, used in conjunction with fluorescence in situ hybridization (FISH) tagging of specific barley chromosomes, has enabled access to the physical recombination landscape of the barley cultivars Morex and Bowman. Consistent distal localization of HvMLH3 foci throughout the genome, and similar patterns of HvMLH3 foci within bivalents 2H and 3H have been observed. A difference in total numbers of HvMLH3 foci between these two cultivars has been quantified, which is interpreted as representing genotypic variation in class I crossover frequency. Discrepancies between the frequencies of HvMLH3 foci and crossover frequencies derived from linkage analysis point to the existence of at least two crossover pathways in barley. It is also shown that interference of HvMLH3 foci is relatively weak compared with other plant species.

High chromosome variability and the presence of multivalent associations in buthid scorpions

In this study, we investigated the mitotic and meiotic chromosomes of 11 Buthidae scorpion species, belonging to three genera (Ananteris, Rhopalurus and Tityus), to obtain detailed knowledge regarding the mechanisms underlying the intraspecific and/or interspecific diversity of chromosome number and the origin of the complex chromosome associations observed during meiosis. The chromosomes of all species did not exhibit a localised centromere region and presented synaptic and achiasmatic behaviour during meiosis I. Spermatogonial and/or oogonial metaphase cells of these buthids showed diploid numbers range from 2n = 6 to 2n = 28. In most species, multivalent chromosome associations were observed in pachytene and postpachytene nuclei. Moreover, intraspecific variability associated with the presence or absence of chromosome chains and the number of chromosomes in the complex meiotic configurations was observed in some species of these three genera. Silver-impregnated cells revealed that the number and location of nucleolar organiser regions (NORs) remained unchanged despite extensive chromosome variation; notably, two NORs located on the terminal or subterminal chromosome regions were commonly observed for all species. C-banded and fluorochrome-stained cells showed that species with conspicuous blocks of heterochromatin exhibited the lowest rate of chromosomal rearrangement. Based on the investigation of mitotic and meiotic cells, we determined that the intraspecific variability occurred as a consequence of fission/fusion-type chromosomal rearrangements in Ananteris and Tityus species and reciprocal translocation in Rhopalurus species. Furthermore, we verified that individuals presenting the same diploid number differ in structural chromosome organisation, giving rise to intraspecific differences of chromosome association in meiotic cells (bivalent-like elements or chromosome chains).

Together yes, but not coupled: new insights into the roles of RAD51 and DMC1 in plant meiotic recombination

The eukaryotic recombinases RAD51 and DMC1 are essential for DNA strand-exchange between homologous chromosomes during meiosis. RAD51 is also expressed during mitosis, and mediates homologous recombination (HR) between sister chromatids. It has been suggested that DMC1 might be involved in the switch from intersister chromatid recombination in somatic cells to interhomolog meiotic recombination. At meiosis, the Arabidopsis Atrad51 null mutant fails to synapse and has extensive chromosome fragmentation. The Atdmc1 null mutant is also asynaptic, but in this case chromosome fragmentation is absent. Thus in plants, AtDMC1 appears to be indispensable for interhomolog homologous recombination, whereas AtRAD51 seems to be more involved in intersister recombination. In this work, we have studied a new AtRAD51 knock-down mutant, Atrad51-2, which expresses only a small quantity of RAD51 protein. Atrad51-2 mutant plants are sterile and hypersensitive to DNA double-strand break induction, but their vegetative development is apparently normal. The meiotic phenotype of the mutant consists of partial synapsis, an elevated frequency of univalents, a low incidence of chromosome fragmentation and multivalent chromosome associations. Surprisingly, non-homologous chromosomes are involved in 51% of bivalents. The depletion of AtDMC1 in the Atrad51-2 background results in the loss of bivalents and in an increase of chromosome fragmentation. Our results suggest that a critical level of AtRAD51 is required to ensure the fidelity of HR during interchromosomal exchanges. Assuming the existence of asymmetrical DNA strand invasion during the initial steps of recombination, we have developed a working model in which the initial step of strand invasion is mediated by AtDMC1, with AtRAD51 required to check the fidelity of this process.

Synaptonemal Complex-Dependent Centromeric Clustering and the Initiation of Synapsis in Drosophila Oocytes

The pairing of homologous chromosomes and the intimate synapsis of the paired homologs by the synaptonemal complex (SC) are essential for subsequent meiotic processes including recombination and chromosome segregation. Here we show that the centromere clustering plays an important role in initiating homolog synapsis during meiosis in Drosophila females. Although centromeres are not clustered prior to the onset of meiosis, all four pairs of centromeres are actively clustered into one or two masses during early meiotic prophase. Within the 16-cell cyst, centromeric clustering appears to define the first step in the initiation of synapsis. Clustering is restricted to the nuclei that form the SC and is dependent on all known SC proteins. Surprisingly, both centromeric clusters and the SC components associated with them persist long after the disassembly of the euchromatic SC at the end of pachytene. The initiation of homologous recombination through the formation of programmed double-strand breaks (DSBs) is not required for either the formation or the maintenance of the centromeric clusters. Our data support a view in which the SC-mediated clustering at the centromeres is the initiating event for meiotic synapsis.

Cohesin proteins load sequentially during prophase I in tomato primary microsporocytes

Proteins of the cohesin complex are essential for sister chromatid cohesion and proper chromosome segregation during both mitosis and meiosis. Cohesin proteins are also components of axial elements/lateral elements (AE/LEs) of synaptonemal complexes (SCs) during meiosis, and cohesins are thought to play an important role in meiotic chromosome morphogenesis and recombination. Here, we have examined the cytological behavior of four cohesin proteins (SMC1, SMC3, SCC3, and REC8/SYN1) during early prophase I in tomato microsporocytes using immunolabeling. All four cohesins are discontinuously distributed along the length of AE/LEs from leptotene through early diplotene. Based on current models for the cohesin complex, the four cohesin proteins should be present at the same time and place in equivalent amounts. However, we observed that cohesins often do not colocalize at the same AE/LE positions, and cohesins differ in when they load onto and dissociate from AE/LEs of early prophase I chromosomes. Cohesin labeling of LEs from pachytene nuclei is similar through euchromatin, pericentric heterochromatin, and kinetochores but is distinctly reduced through the nucleolar organizer region of chromosome 2. These results indicate that the four cohesin proteins may form different complexes and/or perform additional functions during meiosis in plants, which are distinct from their essential function in sister chromatid cohesion.

Analysis of Oocyte-Like Cells Differentiated from Porcine Fetal Skin-Derived Stem Cells

We previously reported the differentiation of cells derived from porcine female fetal skin into cells resembling germ cells and oocytes. A subpopulation of these cells expressed germ cell markers and formed aggregates resembling cumulus-oocyte complexes. Some of these aggregates extruded large oocyte-like cells (OLCs) that expressed markers consistent with those of oocytes. The objective of the current study was to further characterize OLCs differentiated from porcine skin-derived stem cells. Reverse transcriptase (RT)-polymerase chain reaction and Western blot revealed the expression of connexin37 and connexin43, both of which are characteristic of ovarian follicles. The expression of meiosis markers DMC1 and synaptonemal complex protein, but not STRA8 and REC8, was detected in the OLC cultures. Immunofluorescence with an antibody against synaptonemal complex protein on chromosome spreads revealed a very small subpopulation of stained OLCs that had a similar pattern to leptotene, zytotene, or pachytene nuclei during prophase I of meiosis. Sodium bisulfite sequencing of the differentially methylated region of H19 indicated that this region is almost completely demethylated in OLCs, similar to in vivo-derived oocytes. We also investigated the differentiation potential of male skin-derived stem cells in the same differentiation medium. Large cells with oocyte morphology were generated in the male stem cell differentiation cultures. These OLCs expressed oocyte genes such as octamer-binding transcription factor 4 (OCT4), growth differentiation factor-9b (GDF9B), deleted in azoospermia-like (DAZL), VASA, zona pellucida B (ZPB), and zona pellucida C (ZPC). It was concluded that skin-derived stem cells from both male and female porcine fetuses are capable of entering an oocyte differentiation pathway, but the culture system currently in place is inadequate to support the complete development of competent oocytes.

How do chromosomes attach to synaptonemal complexes?

Fluorochrome-labeled oligonucleotides (n = 44) corresponding to mouse genome repetitive sequences were hybridized in situ with pachytene nuclei of mouse spermatocytes. Signals of the repetitive sequences MaLR, MER, and (GT)22 were found to be dispersed through chromatin, and signals of BI 1 repeats and minisatellites were mostly attached to synaptonemal complexes immunostained with anti-SYCP3 antibodies. These results suggest that B 1 repeats and minisatellites are candidates for sequences anchoring chromatin to synaptonemal complexes.

Mapping of pachytene bivalents of female codling moth Cydia pomonella (Linnaeus) (Lep., Tortricidae)

AbstractSpread pachytene nuclei of codling moth Cydia pomonella (Linnaeus) (Lep., Tortricidae) females of a Syrian strain (SY) were used to investigate chromomere patterns of chromosome bivalents and determine their length. The karyotype of female codling moths consists of 28 chromosome bivalents, of which seven are clearly distinguishable using chromosome length and the number and size of the chromomeres in the pachytene stage. One autosome bivalent has two nucleolar organizing regions (NORs) that are located at the opposite ends of the chromosome and appear as distinct structural landmarks. In female codling moths, the WZ sex chromosome bivalent was easily identified in pachytene oocytes according to the heterochromatic thread of the W chromosome. This study contributed to the knowledge and identification of pachytene chromosomes of female codling moths.

The synaptonemal complexes of Caenorhabditis elegans: duplication mutants sDp1 and mnDp1, disjunction regulator regions and aneuploidy

AbstractThe duplication mutants sDp1 and mnDp1 in Caenorhabditis elegans differ in their size and recombination/pairing strategies within the pachytene nucleus. mnDp1 is a duplication of approximately 18% of the X chromosome with the duplicated segment transposed and inserted into linkage group V. sDp1 is a free duplication which covers 30 map units of linkage group I and crossing-over has been determined genetically with its homologue. Analysis of the synaptonemal complexes (SC) and pachytene karyotypes of both duplication mutants reveal that there is an extension of one of the SCs in mnDp1 while the sDp1 free duplication partially pairs with its homologue along a small portion of its length. The remaining region exists as a univalent in the pachytene nucleus. This indicates that there is at least one SC initiation site on the sDp1 free duplication. Only bivalent pairing is permitted and there are no trivalents. To some extent, the autosomes preferentially pair at the exclusion of the sDp1 duplication. Switching of pairing partners was evident between the duplication and the homologue, probably because of the size of the duplication. Thus, the mechanism of chromosome segregation in the two duplications is different. The number of Disjunction Regulator Regions, which are associated with X-chromosome nondisjunction, was three in both mutants compared to six in wild-type. The number of males produced in mnDp1 was 1.0%, in sDp1 it was 2.0%, while in wild-type it is 0.3%. Recombination nodules were not observed in any nuclei. The ultimate goal of these studies is to correlate the physical and genetic maps and in this study linkage group I has been identified in the pachytene nucleus.

Translational Repression of Cyclin E Prevents Precocious Mitosis and Embryonic Gene Activation during C. elegans Meiosis

Germ cells, the cells that give rise to sperm and egg, maintain the potential to recreate all cell types in a new individual. This wide developmental potential, or totipotency, is manifested in unusual tumors called teratomas, in which germ cells undergo somatic differentiation. Although recent studies have implicated RNA regulation, the mechanism that normally prevents the loss of germ cell identity remains unexplained. In C. elegans, a teratoma is induced in the absence of the conserved RNA-binding protein GLD-1. Here, we demonstrate that GLD-1 represses translation of CYE-1/cyclin E during meiotic prophase, which prevents germ cells from re-entering mitosis and inducing embryonic-like transcription. We describe a mechanism that prevents precocious mitosis in germ cells undergoing meiosis, propose that this mechanism maintains germ cell identity by delaying the onset of embryonic gene activation until after fertilization, and provide a paradigm for the possible origin of human teratomas.

Influence of sex on the meiotic segregation of a t(13;17) Robertsonian translocation: a case study in the pig

Comparison of male versus female meiotic segregation patterns for Robertsonian translocation (RT) carriers with similar genetic background has rarely been reported in mammalian species. The aim of this study was to compare the segregation patterns determined for related males and females carrying a 13;17 RT in an animal model (Sus scrofa domestica L.), using dual colour fluorescence in situ hybridization on decondensed sperm nuclei and metaphases II of in vitro-matured oocytes. In males, no association between the trivalent and the XY body was observed in any of the 90 pachytene nuclei studied, and the rate of unbalanced spermatozoa ranged between 2.96% and 3.83%. Female meiotic segregation analyses were carried out on 83 metaphase II oocytes. The rate of unbalanced gametes was higher in females than in males (28.91% versus 3.21%, P < 0.001). This difference was due to higher rates of diploid gametes (12.04% versus 0.05%) and unbalanced gametes produced by the adjacent segregation (16.86% versus 3.16%). This study is a new scientific contribution to the comparison of segregation patterns in related males and females carrying an identical chromosomal rearrangement. It allows a better understanding of the meiotic behaviour of RTs. It also clearly illustrates the relevance of swine as an animal model for such meiotic studies.