Presynaptic Alignment Research Articles

Collaborative homologous pairing during C. elegans meiosis

In preparation for meiotic chromosome segregation, homologous chromosomes need to pair, synapse (i.e., assemble the synaptonemal complex, SC), and then recombine to generate a physical linkage (i.e., chiasma) between them. In many organisms meiotic pairing capacity distributed along the entire chromosome length supports presynaptic alignment. In contrast, the prevailing model for C. elegans proposes that presynaptic homologous pairing is performed solely by a master pairing-site, the pairing center (PC). In this model, the remaining chromosomal regions (the non-PC regions) are not actively involved in presynaptic pairing, and the SC assembling from the PC aligns the homologous chromosomes along non-PC regions and holds them together. Our recent work, however, demonstrates that C. elegans chromosomes establish presynaptic alignment along the entire chromosome length, suggesting that the non-PC regions are also actively involved in the presynaptic pairing process. Furthermore, we have also discovered that the chromodomain protein MRG-1 facilitates this presynaptic non-PC pairing. The phenotype of the mrg-1 mutant indicates that the PC and the non-PC collaborate in successful pairing and synapsis. Therefore, homologous pairing mechanisms in C. elegans possibly share more similarity with those in other organisms than previously thought. Here, we elaborate on these observations and discuss a hypothetical model for presynaptic pairing in C. elegans based on our novel findings.

The Chromodomain Protein MRG-1 Facilitates SC-Independent Homologous Pairing during Meiosis in Caenorhabditis elegans

Homologous chromosome pairing is a prerequisite to establish physical linkage between homologs, which is critical for faithful chromosome segregation during meiosis I. The establishment of pairing is genetically separable from subsequent synapsis, defined as stabilization of pairing by the synaptonemal complex (SC). The underlying mechanism of presynaptic pairing is poorly understood. In the nematode Caenorhabditis elegans, a unique cis-acting element, the pairing center (PC), is essential for presynaptic pairing; however, it is not known whether and how the remainder of the chromosome contributes to presynaptic pairing. Here we report direct evidence for presynaptic pairing activity intrinsic to non-PC regions, which is facilitated by a conserved chromodomain protein, MRG-1. In mrg-1 loss-of-function mutants, pairing is compromised specifically in non-PC regions, leading to nonhomologous SC assembly. Our data support a model in which presynaptic alignment in non-PC regions collaborates with initial PC pairing to ensure correct homologous synapsis.

MER3 is required for normal meiotic crossover formation, but not for presynaptic alignment in rice

MER3, a ZMM protein, is required for the formation of crossovers in Saccharomyces cerevisiae and Arabidopsis. Here, MER3, the first identified ZMM gene in a monocot, is characterized by map-based cloning in rice (Oryza sativa). The null mutation of MER3 results in complete sterility without any vegetative defects. Cytological analyses show that chiasma frequency is reduced dramatically in mer3 mutants and the remaining chiasmata distribute randomly among different pollen mother cells, implying possible coexistence of two kinds of crossover in rice. Immunocytological analyses reveal that MER3 only exists as foci in prophase I meiocytes. In addition, MER3 does not colocalize with PAIR2 at the beginning of prophase I, but locates on one end of PAIR2 fragments at later stages, whereas MER3 foci merely locate on one end of REC8 fragments when signals start to be seen in early prophase I. The normal loading of PAIR2 and REC8 in mer3 implies that their loading is independent of MER3. On the contrary, the absence of MER3 signal in pair2 mutants indicates that PAIR2 is essential for the loading and further function of MER3.

Mechanics of meiosis.

The present paper reviews our observations on the premeiotic DNA replication in Lilium and on the morphology and behaviour of chromosomes during the meiotic prophase in Schizophyllum, Lilium, Drosophila, Bombyx and Homo as revealed by three-dimensional reconstructions of synaptonemal complexes (SC). The main results and conclusions are the following: (1) Premeiotic DNA replication in Lilium is 6 times longer than the mitotic S-phase. (2) Presynaptic alignment of homologous chromosomes was not observed. (3) Initiation of synapsis was in Bombyx and Homo restricted to telomere regions while in Lilium complex formation was generally initiated interstitially. (4) Interlocking of chromosomes was frequent at zygotene. (5) Direct evidence for resolution of interlockings by breakage and reunion of chromosomes was obtained in Homo and Bombyx. (6) Two phases of chromosome pairing and SC formation were revealed in Bombyx. (7) A karyotype based on SC length, centromere index and chromatin markers was constructed for human spermatocytes. (8) The number and distribution of recombination nodules were in agreement with the hypothesis that these structures are involved in crossing over and/or gene conversion. (9) Fragments of the SC were found in prometaphase I bivalents of human spermatocytes corresponding in number to the number of chiasmata. (10) In the absence of crossing over, compensatory mechanisms have evolved in female Bombyx substituting for chiasmata in the retention of homologues.

Pairing and synapsis in wild type Arabidopsis thaliana

A spreading technique was used to perform a structural analysis of prophase I nuclei in pollen mother cells (PMCs) of wild-type Arabidopsis thaliana. In leptotene, all chromosomes developed fully axial elements before a presynaptic alignment was observed. Pairing and synapsis start in regions close to the telomeres at early zygotene. Interstitial synaptonemal complex (SC) stretches were found to occur at several sites per bivalent at mid zygotene. Within individual bivalents, extensive regions of SC formation often existed at the same time as other extensive regions that were unsynapsed. Also in the same nucleus, one bivalent might have several SC segments, while other bivalents have only a few. The classical bouquet was not so evident as in other plant species. Length measurements of the five pachytene bivalents have allowed the elaboration of a pachytene karyotype. Pachytene chromatin compaction in Arabidopsis was significantly less than that observed in the other species analysed and this is paralleled with a higher recombination rate (centimorgans per megabase).

From early homologue recognition to synaptonemal complex formation

This review focuses on various aspects of chromosome homology searching and their relationship to meiotic and vegetative pairing and to the silencing of unpaired copies of genes. Chromosome recognition and pairing is a prominent characteristic of meiosis; however, for some organisms, this association (complete or partial) is also a normal part of nuclear organization. The multiple mechanisms suggested to contribute to homologous pairing are analyzed. Recognition of DNA/DNA homology also plays an important role in detecting DNA segments that are present in inappropriate number of copies before and during meiosis. In this context, the mechanisms of methylation induced premeiotically, repeat-induced point mutation, meiotic silencing by unpaired DNA, and meiotic sex chromosome inactivation will be discussed. Homologue juxtaposition during meiotic prophase can be divided into three mechanistically distinct steps, namely, recognition, presynaptic alignment, and synapsis by the synaptonemal complex (SC). In most organisms, these three steps are distinguished by their dependence on DNA double-strand breaks (DSBs). The coupling of SC initiation to (and downstream effects of) DSB formation and the exceptions to this dependency are discussed. Finally, this review addresses the specific factors that appear to promote chromosome movement at various stages of meiotic prophase, most particularly at the bouquet stage, and on their significance for homologue pairing and/or achieving a final pachytene configuration.

Localization and roles of Ski8p protein in Sordaria meiosis and delineation of three mechanistically distinct steps of meiotic homolog juxtaposition.

Ski8p is implicated in degradation of non-poly(A) and double-stranded RNA, and in meiotic DNA recombination. We have identified the Sordaria macrospora SKI8 gene. Ski8p is cytoplasmically localized in all vegetative and sexual cycle cells, and is nuclear localized, specifically in early-mid-meiotic prophase, in temporal correlation with Spo11p, the meiotic double-strand break (DSB) transesterase. Localizations of Ski8p and Spo11p are mutually interdependent. ski8 mutants exhibit defects in vegetative growth, entry into the sexual program, and sporulation. Diverse meiotic defects, also seen in spo11 mutants, are diagnostic of DSB absence, and they are restored by exogenous DSBs. These results suggest that Ski8p promotes meiotic DSB formation by acting directly within meiotic prophase chromosomes. Mutant phenotypes also divide meiotic homolog juxtaposition into three successive, mechanistically distinct steps; recognition, presynaptic alignment, and synapsis, which are distinguished by their differential dependence on DSBs.

Genetic and cytological characterization of the recombination protein RAD-51 in Caenorhabditis elegans.

We investigated the role of Caenorhabditis elegans rad-51 during meiotic prophase. We showed that rad-51 mutant worms are viable, have no defects in meiotic homology recognition and synapsis but exhibit abnormal chromosomal morphology and univalent formation at diakinesis. During meiosis RAD-51 becomes localized to distinct foci in nuclei of the transition zone of the gonad and is most abundant in nuclei at late zygotene/early pachytene. Foci then gradually disappear from chromosomes and no foci are observed in late pachytene. RAD-51 localization requires the recombination genes spo-11 and mre-11 as well as chk-2, which is necessary for homology recognition and presynaptic alignment. Mutational analysis with synapsis- and recombination-defective strains, as well as the analysis of strains bearing heterozygous translocation chromosomes, suggests that presynaptic alignment may be required for RAD-51 focus formation, whereas homologous synaptonemal complex formation is required to remove RAD-51 foci.

A model for chromosome structure during the mitotic and meiotic cell cycles.

The chromosome scaffold model in which loops of chromatin are attached to a central, coiled chromosome core (scaffold) is the current paradigm for chromosome structure. Here we present a modified version of the chromosome scaffold model to describe chromosome structure and behavior through the mitotic and meiotic cell cycles. We suggest that a salient feature of chromosome structure is established during DNA replication when sister loops of DNA extend in opposite directions from replication sites on nuclear matrix strands. This orientation is maintained into prophase when the nuclear matrix strand is converted into two closely associated sister chromatid cores with sister DNA loops extending in opposite directions. We propose that chromatid cores are contractile and show, using a physical model, that contraction of cores during late prophase can result in coiled chromatids. Coiling accounts for the majority of chromosome shortening that is needed to separate sister chromatids within the confines of a cell. In early prophase I of meiosis, the orientation of sister DNA loops in opposite directions from axial elements assures that DNA loops interact preferentially with homologous DNA loops rather than with sister DNA loops. In this context, we propose a bar code model for homologous presynaptic chromosome alignment that involves weak paranemic interactions of homologous DNA loops. Opposite orientation of sister loops also suppresses crossing over between sister chromatids in favor of crossing over between homologous non-sister chromatids. After crossing over is completed in pachytene and the synaptonemal complex breaks down in early diplotene (= diffuse stage), new contractile cores are laid down along each chromatid. These chromatid cores are comparable to the chromatid cores in mitotic prophase chromosomes. As an aside, we propose that leptotene through early diplotene represent the 'missing' G2 period of the premeiotic interphase. The new chromosome cores, along with sister chromatid cohesion, stabilize chiasmata. Contraction of cores in late diplotene causes chromosomes to coil in a configuration that encourages subsequent syntelic orientation of sister kinetochores and amphitelic orientation of homologous kinetochore pairs on the spindle at metaphase I.

Synapsis in a natural autotetraploid

To test assumptions of the autotetraploid chromosome pairing model regarding events during synapsis, whole-mount spreads of synaptonemal complexes (SCs) of Machaeranthera pinnatifida (=Haplopappus spinulosus) (Asteraceae) (2n = 4x = 16) were analyzed by electron microscopy. On the assumption of one synaptic initiation per chromosome arm, each pachytene quadrivalent is expected to have one partner switch (PS), and the frequency of pachytene quadrivalents for each chromosome is predicted to be 2/3 (or 0.67). However, to the contrary, we observed a range of one to four PSs per pachytene quadrivalent with an overall mean of 1.56. This suggests that the number of synaptic initiations is greater than one per chromosome arm (or >two per chromosome), and the predicted frequency of pachytene quadrivalents should be >8/9 (based on a minimum of three initiations per chromosome). However, in close agreement with the model, the observed pachytene quadrivalent frequency from SCs in this study was 0.69. To explain the apparent discrepancy between the observed frequency of PSs and the observed frequency of quadrivalents, the possibility of nonindependent synaptic initiations and presynaptic alignment are discussed in the context of their potential influence on quadrivalent frequency. Recombination nodules (RNs), which were scored in about half the SC spreads, occurred at a frequency (9.6 per nucleus) comparable with the chiasma frequency at diakinesis (9.3 per nucleus). The frequency of RNs as well as their distribution is consistent with the hypothesis that RNs occur at sites of crossing over and chiasma formation.Key words: autopolyploid, Machaeranthera pinnatifida, meiosis, recombination nodules, synaptonemal complex.

Synapsis in a natural autotetraploid

To test assumptions of the autotetraploid chromosome pairing model regarding events during synapsis, whole-mount spreads of synaptonemal complexes (SCs) of Machaeranthera pinnatifida (=Haplopappus spinulosus) (Asteraceae) (2n = 4x = 16) were analyzed by electron microscopy. On the assumption of one synaptic initiation per chromosome arm, each pachytene quadrivalent is expected to have one partner switch (PS), and the frequency of pachytene quadrivalents for each chromosome is predicted to be 2/3 (or 0.67). However, to the contrary, we observed a range of one to four PSs per pachytene quadrivalent with an overall mean of 1.56. This suggests that the number of synaptic initiations is greater than one per chromosome arm (or >two per chromosome), and the predicted frequency of pachytene quadrivalents should be >8/9 (based on a minimum of three initiations per chromosome). However, in close agreement with the model, the observed pachytene quadrivalent frequency from SCs in this study was 0.69. To explain the apparent discrepancy between the observed frequency of PSs and the observed frequency of quadrivalents, the possibility of nonindependent synaptic initiations and presynaptic alignment are discussed in the context of their potential influence on quadrivalent frequency. Recombination nodules (RNs), which were scored in about half the SC spreads, occurred at a frequency (9.6 per nucleus) comparable with the chiasma frequency at diakinesis (9.3 per nucleus). The frequency of RNs as well as their distribution is consistent with the hypothesis that RNs occur at sites of crossing over and chiasma formation.Key words: autopolyploid, Machaeranthera pinnatifida, meiosis, recombination nodules, synaptonemal complex.

Effect of the pairing gene Ph1 and premeiotic colchicine treatment on intra- and interchromosome pairing of isochromosomes in common wheat.

The analysis of the pattern of isochromosome pairing allows one to distinguish factors affecting presynaptic alignment of homologous chromosomes from those affecting synapsis and crossing-over. Because the two homologous arms in an isochromosome are invariably associated by a common centromere, the suppression of pairing between these arms (intrachromosome pairing) would indicate that synaptic or postsynaptic events were impaired. In contrast, the suppression of pairing between an isochromosome and its homologous chromosome (interchromosome pairing), without affecting intrachromosome pairing, would suggest that homologous presynaptic alignment was impaired. We used such an isochromosome system to determine which of the processes associated with chromosome pairing was affected by the Ph1 gene of common wheat-the main gene that restricts pairing to homologues. Ph1 reduced the frequency of interchromosome pairing without affecting intrachromosome pairing. In contrast, intrachromosome pairing was strongly reduced in the absence of the synaptic gene Syn-B1. Premeiotic colchicine treatment, which drastically decreased pairing of conventional chromosomes, reduced interchromosome but not intrachromosome pairing. The results support the hypothesis that premeiotic alignment is a necessary stage for the regularity of meiotic pairing and that Ph1 relaxes this alignment. We suggest that Ph1 acts on premeiotic alignment of homologues and homeologues as a means of ensuring diploid-like meiotic behavior in polyploid wheat.

Compound synapses within the GABAergic innervation of the auditory inner hair cells in the adolescent mouse.

Ultrastructural investigation of the gamma-aminobutyric acid (GABA) component of the inner spiral bundle in adolescent mice revealed a pathway of glutamic acid decarboxylase (GAD)-positive and -negative fibers and vesiculated endings that contact inner hair cells and their afferents through a complex of axosomatic and axodendritic synapses. Ultrastructural details were investigated by using conventional electron microscopy. Several synaptic arrangements were observed: Main axosomatic synapses form between vesiculated endings and individual or adjoining inner hair cells (interreceptor synapses). Spinous synapses form on long, spinelike processes that protrude from inner hair cells to reach distant efferent endings. The efferent endings associate with inner hair cells and their synaptic afferents through compound synapses-serial, "converging," and triadic-otherwise characteristic of sensory relay nuclei. Serial synapses form by the sequential presynaptic alignment of the efferent-->receptor-->afferent components. Converging synapses result from the simultaneous apposition of a receptor ribbon synapse and a presynaptic efferent terminal on a recipient afferent dendrite. Triadic synapses comprise a vesiculated efferent ending in contact with an inner hair cell and with its synaptic afferent. Additionally, efferent endings may form simple axodendritic and axoaxonal synapses with GAD-negative vesiculated endings. The combination of different synaptic arrangements leads to short chains of compound synapses. It is assumed that these synaptic patterns seen in the adolescent mouse represent adult synaptology. The patterns of synaptic connectivity suggest an integrative role for the GABA/GAD lateral efferent system, and imply its involvement in the pre- and postsynaptic modulation of auditory signals.

Localized chiasmata and meiotic nodules in the tetraploid onionAllium porrum

Allium porrum L. (cultivated leek) (2n = 4x = 32) is a fertile tetraploid that forms bivalents with pericentric chiasmata at metaphase I. To investigate the basis of this unusual behavior for a tetraploid, we describe the karyotype, axial cores, synaptonemal complexes (SCs), and meiotic nodules of A. porrum. The karyotype appears to be autotetraploid. This conclusion is also supported by presynaptic alignment of axial cores in groups of four and partner trades between pairs of SCs. Numerous early nodules are distributed all along axial cores and SCs during zygonema, but they are lost by late zygonema - early pachynema. Late (recombination) nodules (RNs) are present on SCs near kinetochores throughout the remainder of pachynema. This pattern of RNs corresponds to the pattern of pericentric chiasmata. Pachytene quadrivalents usually are resolved into bivalents because partner trades between SC lateral elements rarely occur between RNs on the same segment of SC. Thus, the patterns of crossing-over and partner trades promote balanced disjunction and high fertility in autotetraploid A. porrum. Rare quadrivalents observed at metaphase I must be due to infrequent partner trades between RNs. Polycomplexes, unusual in their number and size, were observed during zygonema. Key words : synaptonemal complex, recombination nodules, localized chiasmata, polycomplex, Allium porrum.

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.

Synaptic patterns of rye B chromosomes. I: The standard type.

Chromosome pairing of standard B chromosomes (Bs) in rye has been studied by synaptonemal complex surface spreading of pollen mother cells containing from one to eight Bs. The main characteristics exhibited by the Bs are: (i) They are often located peripherally in surface spread nuclei and do not show presynaptic alignment. (ii) The period of pairing is delayed relative to that of the A set. (iii) Bs undergo extensive non-homol-ogous pairing at pachytene. (iv) The frequency of trivalents and quadrivalents in plants with 3Bs and 4Bs is about 66% following the expectations on the basis of random pairing among the short and the long arms with a single distal pairing initiation site per arm. Some quadrivalents containing two partner switches were observed, but on the same basis of randomness there was a deficiency of pachytene multivalents in plants with 5-8 Bs. The possible causes of the discrepancy between these observations are discussed. (v) There is a reduction in the frequency of multivalents between pachytene and metaphase I.

The initiation of meiotic chromosome pairing: the cytological view

Opposing views are held with respect to the time when and the mechanisms whereby homologous chromosomes find each other for meiotic synapsis. On the one hand, some evidence has been presented for somatic homologous associations or some other kind of relationship between chromosomes in somatic cells as a preliminary to meiotic pairing. On the other hand, it is argued by many that homologous contacts are first established at meiotic prophase prior to, or in the course of, synaptonemal complex formation. The present paper reviews the controversial cytological evidence, hypotheses, and ideas on how the first contact between homologous chromosomes comes about.

EM analysis of meiotic chromosome pairing in a pentaploid Achillea hybrid

Presynaptic alignment occurs at zygotene and early pachytene in a pentaploid hybrid between Achillea collina (4x) and A. millefolium (6x). Mostly, the five corresponding chromosomes of each set are assorted into a group of three (two synapsed plus one aligned) and a group of two (synapsed); groups of five axes by alignment and/or pairing are extremely rare. This reflects different degrees of homology between the five genomes and allows inferences about the possible phylogeny and genome formulas of the tetraploid (AASS) and hexaploid (AASSTT) involved. The alignment as subgroups of three and two chromosomes in their 5x hybrid also indicates that synaptic alignment responds to slightly different degrees of homology and is decisive regarding which chromosomes will finally pair effectively and cross over. At early zygotene, alignment is weakly expressed, arguing against premeiotic associations as a precondition for homologous alignment and subsequent synaptonemal complex formation. At late pachytene, synapsis is maximized by autosynapsis of chromosomes which do not take part in the preceding homologous two-by-two synapsis.

Colchicine action at meiotic prophase revealed by SC-spreading

In the present paper earlier findings on the interference of colchicine with presynaptic alignment and synaptonemal complex formation in Allium ursinum are corroborated and several other direct or indirect consequences of colchicine treatment, like deformations and decomposition of lateral elements and a high incidence of non-homologous associations are reported. Mechanisms of meiotic alignment and pairing are discussed with respect to their susceptibility to the known cytological and biochemical activities of colchicine.

Synaptonemal complex spreading in Allium ursinum: pericentric asynapsis and axial thickenings.

In Allium ursinum meiotic pairing of homologues is always incomplete; a proximal region on each bivalent remains regularly unsynapsed even in late pachytene. The spatial correlation of the unsynapsed region with the kinetochore suggests that the kinetochore itself exerts an inhibitory effect on synapsis in its vicinity. This can be interpreted as the cytological basis of the 'centromere effect' on recombination in this species. Moreover, the high incidence of a pericentric inversion loop in a heterozygous chromosome pair shows that proximal pairing initiation is possible and that its failure cannot be responsible for pericentric asynapsis. The formation of the inversion loop is complicated by the need for two independent pairing initiation sites because synapsis cannot proceed across the pericentric region. It is proposed that the meiotic bouquet polarization helps in establishing the presynaptic alignment of the homologous sites within the inverted regions and hence to achieve a high rate of inversion loop formation. Thickenings of the axial/lateral elements are not distributed equally along the synaptonemal complex. They are underrepresented in unpaired axes but strikingly abundant at the borders with synapsed regions, suggesting their origin in the pairing forks during the process of synapsis. They are virtually always present at nucleolus-organizing regions and often they appear at corresponding sites on opposite lateral elements. Besides the thickenings several other kinds of axial deformities are present in unpaired axes.