Programmed phosphorylation of histone H2AX precedes a phase of DNA double-strand break-independent synapsis in mouse meiosis

Accurate homologue synapsis during meiosis is essential for faithful chromosome segregation and formation of viable gametes. The finding ofSpo11-dependent gamma-H2AX (γH2AX) formation during leptotene and data on mutant mice have led to the notion that synapsis in mammals depends on meiotic DNA double-stranded break (DSB) repair. A second wave of ataxia telangiectasia mutated (ATM) and Rad3-related (ATR)-dependent γH2AX formation has been observed inAtm-null mice during zygotene, suggesting that this wave of phosphorylation also occurs in normal mice. Here I aimed to confirm and to analyse in deep this wave of phosphorylation. Immunostaining of spread spermatocytes shows that γH2AX accumulates on the short last axis stretches to pair. This accumulation appears within all the nuclei undergoing a specific step of late zygotene and disappears from every spermatocyte immediately after pairing completion. This γH2AX signal co-localises with ATR, isSpo11-independent and does not co-localise with free DNA 3′-end labelling. I conclude that ATR/γH2AX asynapsis signalling at the end of zygotene belongs to a physiologically programmed pathway operating at a specific meiotic step, and I propose that this pathway is involved in the triggering of a phase of DSB-independent chromosome pairing that leads to synapsis completion in normal mouse meiosis.

The Distribution of Early Recombination Nodules on Zygotene Bivalents From Plants

Early recombination nodules (ENs) are protein complexes ~100 nm in diameter that are associated with forming synaptonemal complexes (SCs) during leptotene and zygotene of meiosis. Although their functions are not yet clear, ENs may have roles in synapsis and recombination. Here we report on the frequency and distribution of ENs in zygotene SC spreads from six plant species that include one lower vascular plant, two dicots, and three monocots. For each species, the number of ENs per unit length is higher for SC segments than for (asynapsed) axial elements (AEs). In addition, EN number is strongly correlated with SC segment length. There are statistically significant differences in EN frequencies on SCs between species, but these differences are not related to genome size, number of chromosomes, or phylogenetic class. There is no difference in the frequency of ENs per unit length of SC from early to late zygotene. The distribution of distances between adjacent ENs on SC segments is random for all six species, but ENs are found at synaptic forks more often than expected for a random distribution of ENs on SCs. From these observations, we conclude that in plants: (1) some ENs bind to AEs prior to synapsis, (2) most ENs bind to forming SCs at synaptic forks, and (3) ENs do not bind to already formed SCs.

Analysis of synaptonemal complexes in the amphidiploid of Lolium multiflorum × Festuca drymeja

Synaptonemal complexes in the amphidiploid Lolium multiflorum (4χ) × Festuca drymeja (4χ) have been examined by the whole mount spreading technique in nuclei with between 49 and 100% pairing. At mid to late zygotene nonhomologous associations are formed, with multivalents involving more than half the total axial element length in some cases. However, they are corrected by pachytene. There is evidence of differences in the timing or rate of pairing between the two sets of chromosomes; the L. multiflorum chromosomes seem more advanced than the F. drymeja chromosomes in their pairing at mid and late zygotene, and it is possible that this asynchrony places a constraint on intergenomic chromosome pairing.Key words: Lolium-Festuca, amphidiploid, synaptonemal complexes, nonhomologous pairing, correction mechanism.

Synaptonemal complex spreading in Allium cepa and Allium fistulosum. III. The F1 hybrid

Synaptonemal complexes (SCs) were analysed, by surface spreading, in the F1 hybrid between Allium cepa and Allium fistulosum. These closely related species have a similar karyomorphology, but the A. cepa genome has 27% more DNA than A. fistulosum. At metaphase I, in the hybrid, heteromorphic bivalents were observed, and the pollen mother cell chiasma frequency in the F1 hybrid was reduced when compared with the parents. Synapsis was incomplete and disturbed to some extent in all the prophase I nuclei observed. The reduced level of synapsis at late zygotene – mid pachytene in the hybrid (mean percent homoeologous synapsis is 60%) corresponds to the percent reduction in chiasma frequency. It is suggested that failure of synapsis is the main cause of reduction of chiasma frequency in the F1 hybrid. The chiasma distribution in the hybrid is most similar to that of the A. cepa parent. Although some proximal chiasmata were observed (10% of total), none of the bivalents observed had only proximally localised chiasmata as found in the A. fistulosum parent. In the F1 hybrid, synapsis of the centromeric regions of the bivalents was invariably disturbed throughout prophase I. It is suggested that there are major DNA differences in these regions preventing regular synapsis or progression of synapsis and possibly proximal chiasma formation. Nonhomoeologous synapsis occurred within axes, giving foldback SCs, and between axes, resulting in multivalent formation. It is proposed that disturbance of synapsis is due to DNA differences. There is no indication of any rearrangement or adjustment of synapsis during prophase I either to give perfectly synapsed bivalents or to resolve multivalents into bivalents.Key words: synaptonemal complex, Allium, incomplete pairing, chiasma, nonhomologous pairing, DNA content.

Chromosomal pairing in deer mice heterozygous for the presence of heterochromatic short arms

The pattern of chromosomal pairing was analyzed in male deer mice (Peromyscus maniculatus and Peromyscus sitkensis) heterozygous for the presence of heterochromatic short arms. G- and C-banding of somatic metaphases indicated that the presence of heterochromatic short arms increased the length of chromosome 4 by 15% in P. sitkensis and that of chromosome 8 by 9% in P. maniculatus. Analysis of silver-stained late zygotene and early pachytene nuclei revealed a low frequency of unequal axial lengths in the synaptonemal complexes corresponding to the heteromorphic bivalents. All mid- and late pachytene nuclei, however, exhibited fully paired synaptonemal complexes with equalized axial lengths. These observations suggest the existence of an adjustment mechanism which functions to equalize the lengths of the two axes of the heteromorphic synaptonemal complex.Key words: synaptonemal complex, Peromyscus, heterochromatin, chromosomal polymorphism, synaptic adjustment.

The Occurrence of Polycomplexes During the Sporogenesis in Psilotum Nudum

The structure of the polycomplexes has been extensively studied in recent years (1,2,3). Theres structures are usually described as stacks or aggregates of the tripartite synaptonemal complexes. Although the presence of the synaptonemal complexes is a consistent feature in the late zygotene or pachytene stages of chlasmate meiosis and Its role in the processes of chromosome synapsis and crossing over has been suggested (4), the function of the polycomplexes remains obscure. Most of our understandings of the polycomplexes are obtained from the observations during the gametogenesis of the insects, and only a few examples of this structure in fungi and higher plants have been reported. The present study examines the occurrence of polycomplexes during the sporogenesis in the primitive vascular plant. Sporangia at different developmental stages were fixed with 3% glutaraldehyde in 0.1 M phosphate buffer, and postflxed in 2% osmium tetroxide. Dehydration was carried out with the ethanol series followed by embedding in Epon 812. Ultrathln sections were stained with uranyl acetate and lead citrate.

The assembly of the synaptinemal complex

The assembly of the synaptinemal complex in the ascomycete Neottiella was studied by three-dimensional reconstruction of a late zygotene nucleus. A single banded lateral component is formed between the two sister chromatids of each homologous chromosome prior to their pairing. The central regions are pre-assembled in organized form in folds of the granular part of the nucleolus and then converted into an amorphous transport form. The latter appears to move through the nucleoplasm to sites between the lateral components of synapsing homologous chromosomes. The central region material is reorganized into blocks with a recognizable central component and attached to one lateral component. The last step in the completion of the synaptinemal complex is the association of the free surface of the organized central region with the corresponding segment of the homologous lateral component. The findings are discussed in relation to mechanisms of chromosome pairing and chiasma formation.