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The effects of acrylamide on mouse germ-line and somatic cell chromosomes.

The industrial chemical acrylamide is suspected to induce potentially heritable genetic damage. While several studies in rodents have indicated that this substance can damage spermiogenic cells, resulting in dominant lethals and heritable translocations, cytogenetic assessments of premeiotic and meiotic cells after exposure have produced equivocal results. In the present study, various cytogenetic endpoints in both somatic and germ-line cells from acrylamide-treated mice were evaluated. Sister chromatid exchanges and micronuclei, but not chromosome aberrations, were induced in spleen cells; synaptonemal complex irregularities (asynapsis), but not chromosome aberrations, were induced in germ cells.

Acrylamide↗

Cytological aspects of meiotic recombination.

This article reviews current views on the mechanisms of meiotic homology searching and recombination. It discusses the relationship between molecular events at meiotic prophase and concomitant cytological processes. The role of the synaptonemal complex and other meiosis-specific structures is discussed. Whereas the relationship of crossovers, late recombination nodules, and chiasmata is well established, there is still some controversy about the temporal and causal relationships between double strand breaks, homologue recognition, heteroduplexes, early nodules and presynaptic alignment.

Chromosomes↗

An mre11 mutant of Coprinus cinereus has defects in meiotic chromosome pairing, condensation and synapsis.

The rad11 gene of the basidiomycete Coprinus cinereus is required for the completion of meiosis and for survival after gamma irradiation. We have cloned the rad11 gene and shown that it is a homolog of MRE11, a gene required for meiosis and DNA repair in numerous organisms. The expression of C. cinereus mre11 is induced during prophase I of meiosis and following gamma irradiation. The gene encodes a predicted polypeptide of 731 amino acids, and the mre11-1 (rad11-1) mutation is a single base pair change that results in a stop codon after amino acid 315. The mre11-1 mutant shows enhanced sensitivity to ionizing radiation, but no enhanced sensitivity to UV radiation. It shows a delay in fruitbody formation and a reduction in the number of mushrooms formed per dikaryon. The mre11-1 mutant also has several meiotic defects. Pachytene chromatin condensation is disrupted, and although some meiotic cells appear to achieve metaphase I condensation, no further meiotic progression is observed. The mre11-1 mutant also fails to undergo proper chromosome synapsis; neither axial elements nor mature synaptonemal complexes are complete. Finally, meiotic homolog pairing is reduced in the mre11-1 mutant. Thus, in C. cinereus, Mre11 is required for meiotic DNA metabolism.

Amino Acid Sequence↗

[Cytogenetic study of synapsis and crossing-over in male mice heterozygous for reciprocal translocation T(14; 15)6Ca].

Synapsis and crossing over in male mice heterozygous for reciprocal translocation T (14; 15)6Ca were studied. The translocated multivalent undergoes the synaptic adjustment in the course of meiotic prophase. Translocated distal region of the 14th chromosome forms inproportionally long lateral element of synaptonemal complex. The number of chiasmata in the 14th chromosome increases from 1.02 0.02 in normal karyotype to 1.41 0.03 in heterozygous mice. The density of chiasmata in translocated, distal region is ten times higher than in the other part of the 14th chromosome.

Animals↗

The role of BicD, Egl, Orb and the microtubules in the restriction of meiosis to the Drosophila oocyte.

The oocyte is the only cell in Drosophila that goes through meiosis with meiotic recombination, but several germ cells in a 16-cell cyst enter meiosis and form synaptonemal complexes (SC) before one cell is selected to become the oocyte. Using an antibody that recognises a component of the SC or the synapsed chromosomes, we have analysed how meiosis becomes restricted to one cell, in relation to the other events in oocyte determination. Although BicD and egl mutants both cause the development of cysts with no oocyte, they have opposite effects on the behaviour of the SC: none of the cells in the cyst form SC in BicD null mutants, whereas all of the cells do in egl and orb mutants. Furthermore, unlike all cytoplasmic markers for the oocyte, the SC still becomes restricted to one cell when the microtubules are depolymerised, even though the BicD/Egl complex is not localised. These results lead us to propose a model in which BicD, Egl and Orb control entry into meiosis by regulating translation.

Alleles↗

mei-W68 in Drosophila melanogaster encodes a Spo11 homolog: evidence that the mechanism for initiating meiotic recombination is conserved.

Meiotic recombination requires the action of several gene products in both Saccharomyces cerevisiae and Drosophila melanogaster. Genetic studies in D. melanogaster have shown that the mei-W68 gene is required for all meiotic gene conversion and crossing-over. We cloned mei-W68 using a new genetic mapping method in which P elements are used to promote crossing-over at their insertion sites. This resulted in the high-resolution mapping of mei-W68 to a <18-kb region that contains a homolog of the S. cerevisiae spo11 gene. Molecular analysis of several mutants confirmed that mei-W68 encodes an spo11 homolog. Spo11 and MEI-W68 are members of a family of proteins similar to a novel type II topoisomerase. On the basis of this and other lines of evidence, Spo11 has been proposed to be the enzymatic activity that creates the double-strand breaks needed to initiate meiotic recombination. This raises the possibility that recombination in Drosophila is also initiated by double-strand breaks. Although these homologous genes are required absolutely for recombination in both species, their roles differ in other respects. In contrast to spo11, mei-W68 is not required for synaptonemal complex formation and does have a mitotic role.

Amino Acid Sequence↗

Changes in nuclear structure during eupyrene spermatogenesis in Murex brandaris.

Changes in nuclear structure during eupyrene spermatogenesis of Murex brandaris have been studied using light and electron microscopy. In the first phases, spermatogonia show round nuclei, with several electrodense masses of chromatin and a thin layer of heterochromatin associated with the nuclear membrane. Primary spermatocytes possess larger nucleii, with less condensed chromatin, and the synaptonemal complexes are apparent. During spermiogenesis, chromatin becomes lamellar, and the nucleus twists about its principal axis while it elongates. The nuclear twisting is accompanied by a progressive chromatin condensation, which causes a highly electrodense nucleus at the end of the process.

Animals↗

New data on the synaptic process of Mesocricetus auratus: connecting fibers, telomere association and heterosynapsis.

The progression of the prophase-I stage in Syrian hamster spermatocytes has been studied at different ages, from 12 to 41 days after birth. Two stages, leptotene and diplotene, were identified, which had not previously been described in the Syrian hamster using spreading techniques. The most interesting observations are the presence of heterosynapses and telomere associations in 2.5% of the cells studied, and of nucleolar filaments also in 2.5%. Connecting fibers are structures that establish different types of bridges between two or three synaptonemal complexes (SCs) or between the elements of a single SC. Heterosynapses and telomere associations consist in the partial pairing of the terminal regions of non-homologous lateral elements. These phenomena can be observed both in the autosomes and in the sex chromosomes.

Animals↗

Hypervitaminosis E and gametogenesis in Caenorhabditis elegans.

Expected benefits, i.e. increasing life span and vitality, from ingesting d,1-alpha tocopherol (vitamin E), are not realized using vitamin E acetate (VEA) since the acetate form is only slowly converted to vitamin E in either mammalian or nematode tissues. The resultant accumulation of VEA in the cytosol is toxic, which results in aberrations in nuclear morphology, decreased life span and production of progeny, increased mean reproductive day and general loss of vitality. Incorporation of VEA into membranes results in allosteric changes in membrane structure. Such changes are proportional to increasing concentrations of VEA, thereby inhibiting the attachment of the telomere to the nuclear envelope. Reproductive and chromosomal strategies are compromised such that synaptonemal complexes, normally found during pachytene in oocytes, are rarely present in those nematodes exposed to high concentrations of VEA. The resultant loss of SCs correlates with decreased progeny and the chromosomal aberrations characteristic of hypervitaminosis E.

Animals↗

Protein constitution of the chromosome axis.

Diverse studies indicate that certain nonhistone proteins may be responsible for maintaining the integrity of the eukaryotic chromosomes. In view of their dispersal during nuclear division and their acknowledged role in gene expression, none of the nuclear matrix proteins can be regarded as integral proteins of the chromosome. Topoisomerase II, structural maintenance of chromosome (SMC) proteins and high-mobility-group (HMG) proteins exhibit characteristic enzymatic and mechanochemical roles as well as transitory association with chromosomes. Hence they may be regarded as accessory chromosome proteins. In contrast, certain high molecular weight synaptonemal complex proteins (SCPs) exhibit chemical and physical properties that suggest a structural role as the chromosome axis. It is proposed in an experimentally testable model that, in the minimal structure of a eukaryotic chromosome, the terminals of the loops of DNA-histone solenoids are constitutively affixed to chromosome axis proteins that have elastic properties.

Chromosomes↗

Meiosis, recombination and chromosomes: a review of gene isolation and fluorescent in situ hybridization data in plants.

Evidence is now increasing that many functions and processes of meiotic genes are similar in yeast and higher eukaryotes. However, there are significant differences and, most notably, yeast has considerably higher recombination frequencies than higher eukaryotes, different cross-over interference and possibly more than one pathway for recombination, one late and one early. Other significant events are the timing of double-strand breaks (induced by Spo11) that could be either cause or consequence of homologous chromosome synapsis and SC formation depending on the organisms, yeast plants and mammals versus Drosophila melanogaster and Caenorhabditis elegans. Many plant homologues and heterologues to meiotic genes of yeast and other organisms have now been isolated, in particular in Arabidopsis thaliana, showing that overall recombination genes are very conserved while synaptonemal complex and cohesion proteins are not. In addition to the importance of unravelling the meiotic processes by gene discovery, this review discusses the significance of chromatin packaging, genome organization, and distribution of specific repeated DNA sequences for homologous chromosome cognition and pairing, and the distribution of recombination events along the chromosomes.

Centromere↗

[Comparative genomics and proteomics of Drosophila, Brenner's nematode, and Arabidopsis: identification of functionally similar genes and proteins of meiotic chromosome synapsis].

The published principles of computer analysis of genomes and protein sets in taxonomically distant eukaryotes are expounded. The authors developed a search strategy to identify in genomes of such organisms genes and proteins nonhomologous in primary structure but having similar functions in cells dividing by meiosis. This strategy based on the combined principles of genomics, proteomics, and morphometric analysis of subcellular structures was applied to a computer search for genes encoding the proteins of synaptonemal complexes in genomes of Drosophila melanogaster, the nematode Caenorhabditis elegans, and the plant Arabidopsis thaliana. These proteins proved to be functionally similar to their counterparts in yeast Saccharomyces cerevisiae (protein Zip1p) and mammals (protein SCP1).

Animals↗

Meiotic behavior of rye heterozygous for ditelocentric substitutions.

Meiotic behavior has been investigated in rye plants carrying a heterozygous ditelocentric substitution for chromosomes 1R and 3R. Such a chromosomal rearrangement replaces a biarmed chromosome with two telocentrics, a situation karyotypically comparable to a Robertsonian translocation. As in Robertsonian translocation heterozygotes, delayed pairing characterizes the centromeric region in the rye pachytene heteromorphic trivalents. In addition, telocentric axes appear to be frequently engaged in nonhomologous synapsis (e.g., fold-back loops and two-by-two heterosynapsis) at their centromeric ends. Unlike those in Robertsonian translocation heterozygotes, the synaptic abnormalities detected here cannot be due to lack of partner homology. Rather, the chromosomal rearrangement itself appears to interfere with normal synaptonemal complex (SC) progression. The synaptic irregularities, which often result in complex pachytene multivalents, seem to have no effect on chiasma formation, since the rearranged pairs behave properly at metaphase I. Finally, differences in both SC length and progression patterns between plants and animals could explain the trans configurations of telocentric centromeres in plants, whereas only cis configurations of acrocentric kinetochores have been described in animals.

Chromosome Banding↗

Male pachytene pairing in single and double translocation heterozygotes and spermatogenic impairment in the mouse.

In order to clarify the relationship between meiotic pairing and progress of spermatogenesis, an analysis of male meiotic pairing was carried out in four reciprocal translocation heterozygotes and two double heterozygotes for two semi-identical reciprocal translocations. The reciprocal translocations were chosen to range from fertility (T70H/+) through almost complete sterility (T31H/+) to complete sterility (T32H/+, T42/H+). If meiotic pairing in the translocation multivalent was incomplete, it concerned terminal or probably more often proximal chromosome segments (Chain IV). If both segments failed to pair the multivalent symbol is Chain III + I. Complete pairing is symbolized by Ring IV. To contrast and complement observations of this type, the double heterozygotes were introduced. Males of this type in theory possess two heteromorphic bivalents with a central area of incomplete meiotic pairing (loop formation). Of the T70H/T1Wa double heterozygotes, 36% of the males are capable of inducing at least one decidual reaction in two females whereas for T26H/T2Wa, 79% of the males can do so. For the reciprocal translocations, it was found that proximity of the multivalent to the sex bivalent during pachytene increased in the order Ring IV, Chain IV, Chain III + I. The degree of spermatogenic impairment as measured from cell counts in histological sections and tubular whole mounts, is positively related to the frequency of proximity between the sex chromosomes and the translocation multivalent and thus to lack of meiotic pairing within the multivalent. The meiotic pairing analysis of the double heterozygotes yielded the following findings. For the long heteromorphic bivalents a true loop was never seen in T70H/T1Wa and only rarely observed in T26H/T2Wa. Small marker bivalents of both types were usually recognizable by the following criteria: pairing confined to distal or proximal segments, both distal and proximal segments pairing and loop formation and pairing covering the entire length of both "homologues" but the longer one often with a "thickened" lateral element. The same positive correlation between the absence of pairing (proximal, distal or central) and the proximity of the small marker bivalent synaptonemal complex to the sex bivalent has been found as for unpaired segments within reciprocal translocation multivalents. One unexpected finding was the occurrence of diploid spermatids and spermatozoa especially in T32H/+ males (70-91%) but also in T31H/+ (3-39%).

Animals↗

cDNA cloning and characterization of a novel nucleolar protein.

In an initial study of anti-nuclear antibodies in the chronic inflammatory bladder disease interstitial cystitis, we reported that 7% of interstitial cystitis patients studied had autoantibodies to the nucleolus. We now report that, using an autoimmune serum from a patient with interstitial cystitis, we have identified and partially characterized a novel protein with an M(r) of approximately 55 kDa (hereafter referred to as No55) localized to the granular component of the nucleolus. No55 was initially characterized by diffuse nucleolar immunofluorescence staining in interphase cells and by Western blotting as a 55-kDa doublet on whole-cell extracts. During mitosis, No55 was associated with chromosomes and appeared in prenucleolar bodies during telophase, but it did not colocalize with p80-coilin in coiled bodies. Immunoelectron microscopy revealed that No55 was localized uniformly throughout the granular component of the nucleolus compared with a more peripheral localization of nucleolar granular component protein B23. On segregation of the nucleolus with actinomycin D, No55 remained with the granular component of the segregated nucleolus, whereas protein B23 was found predominantly in the nucleoplasm. Finally, a cDNA expression library was screened with the human autoantibody against No55, and a 2.4-kb insert was isolated, subcloned to homogeneity, and then sequenced. Analysis of this sequence showed an open reading frame of approximately 1.3 kb coding for 437 amino acids with a predicted molecular weight of 50 kDa. A search of the gene sequence database indicated homology with SC65, a rat synaptonemal complex protein. Therefore, on the basis of molecular weight, nucleolar sublocalization, response to actinomycin D, and cDNA sequence determination, No55 is a novel protein of the interphase nucleolus.

Amino Acid Sequence↗

Role of heat shock protein HSP70-2 in spermatogenesis.

The HSP70 heat-shock proteins are molecular chaperones that assist other proteins in their folding, transport and assembly into complexes. Most of these proteins are either constitutively expressed or their expression is induced by heat shock and other stresses. However, two members of the Hsp70 family (HSP70-2 and HSC70T in mice) are regulated developmentally and expressed specifically in spermatogenic cells. The HSP70-2 protein is synthesized during the meiotic phase of spermatogenesis and is abundant in pachytene spermatocytes. The knockout approach was used to determine whether HSP70-2 is a chaperone for proteins involved in meiosis. Male mice lacking HSP70-2 were infertile while females lacking HSP70-2 were fertile. Spermatogenic cell development was arrested in prophase of meiosis I at the G2-M-phase transition and late pachytene spermatocytes were eliminated by apoptosis, resulting in an absence of spermatids. HSP70-2 is required for Cdc2 to form a heterodimer with cyclin B1, suggesting that it is a chaperone necessary for the progression of meiosis in the germ cells of male mice. HSP70-2 is also associated with the synaptonemal complex and desynapsis is disrupted in male mice lacking this protein. Homologues of HSP70-2 are present in the testes of many animals, suggesting that the role of this spermatogenic cell chaperone is conserved across phyla.

Animals↗

Ultrastructural and immunocytochemical analysis of the XY body in rat and Guinea pig.

The formation of the XY body involves the compaction of the extended chromatin to form a mesh of fibrogranular structures. During this process the ribonucleoprotein particles (RNP), which were associated with the chromatin filaments progressively disappear. High resolution immunolocalization indicates that the mature XY body does not contain RNA polymerase II, hnRNPs, or snURNPs. Occasionally chromatin fibrils extend outside of the XY body. These fibrils are frequently associated with nascent RNP fibrils and granules indicating that not all the DNA of the sex chromosomes is transcriptionally inactive. However, transcription is located outside the sex body. The recombination protein Dmc1 is present in nodules associated with the unpaired chromosomal axes of the sex chromosomes located in the XY body. Cytochemical staining methods and in situ hybridization at electron microscopic level show that RNA is present in the unpaired chromosomal axes suggesting that the presence of RNA in the chromosomal axes and in forming synaptonemal complexes is related with the process of final pairing. The sex body and the nucleoli associated with it do not interweave and do not exchange RNA or DNA-containing filaments. These observations indicate that the spatial relation between these structures is just a close proximity, which is, however, very frequent.

Animals↗

Cloning and characterization of MS5 from Arabidopsis: a gene critical in male meiosis.

In this paper, we describe the cloning of the MS5 gene, a gene essential for male fertility in Arabidopsis. We previously defined the MS5 locus by characterizing an EMS-induced allele, ms5-1. We identified a new allele of MS5 (ms5-2) that was T-DNA-generated and used the T-DNA tag to clone the gene. Sequencing of mutant and wild-type alleles together with complementation of the ms5-1 mutant phenotype with a wild-type genomic clone confirmed the identity of the gene. Differences between the phenotypes of the two mutant alleles could be attributed to differences in mutant gene structure. The semi-dominant and dominant negative phenotypes of the ms5-2 mutant probably result from production of a truncated polypeptide. An unknown locus in Landsberg erecta can counteract the dominant negative phenotype of ms5-2. Mutations in MS5 cause the formation 'polyads'--tetrads with more than four pools of chromosomes after male meiosis. Similarities between the MS5 sequence and that of a number of proteins were found; two that may be significant were with a synaptonemal complex protein and with a regulatory subunit of a cyclin-dependent kinase. The MS5 gene is a member of a small gene family highly conserved amongst plant species.

Alleles↗