Three stages of meiotic homologous chromosome pairing in wheat: cognition, alignment and synapsis

 Chromosome painting enabled the study of homologous chromosome behaviour prior to and during meiosis. Total genomic DNA from rye, used as a probe for in situ hybridization, identified the rye chromosome arm in a wheat-rye translocation line (T5AS·5RL) at meiotic prophase and the preceding interphase. Accurate staging of the development of the meiocytes was attained by parallel studies of chromatin morphology, nucleolar behaviour and synaptonemal complex formation in electron microscopy thin sections and silver-stained surface spreads. Three stages of pairing were identified for the large cereal genomes that are organized in a Rabl configuration: first, cognition occurs during the long interphase before leptotene, bringing the homologous chromosome domains into close proximity and possibly starting at the centromere; second, homologous chromosome segments align at late leptotene; and third, zygotene synapsis initiates near the telomere, although it was also observed to occur near the centromere. A pairing model is proposed for wheat, with a genome size of 17000 Mbp, that shows prallels to and notable differences from yeast and mammalian models of meiosis. Received: 25 January 1997 / Revision accepted: 14 July 1997     

Gene conversion, recombination nodules, and the initiation of meiotic synapsis

The nature of the relationship between the two types of meiotic recombination outcomes, exchange (crossing-over) and simple gene conversion, has been debated for years. I here propose that these two types of events are not necessarily causally related and hypothesize that the primary role of events detected as simple gene conversion is in the recognition of homology during synapsis.     

Heterologous synapsis in C. elegans is regulated by meiotic double-strand breaks and crossovers

Chromosoma - Alignment of the parental chromosomes during meiotic prophase is key to the formation of genetic exchanges, or crossovers, and consequently to the successful production of gametes. In...     

Yeast axial-element protein,Red1, binds SUMO chains to promote meiotic interhomologue recombination and chromosome synapsis

The synaptonemal complex (SC) is a tripartite protein structure consisting of two parallel axial elements (AEs) and a central region. During meiosis, the SC connects paired homologous chromosomes, promoting interhomologue (IH) recombination. Here, we report that, like the CE component Zip1, Saccharomyces cerevisiae axial-element structural protein, Red1, can bind small ubiquitin-like modifier (SUMO) polymeric chains. The Red1–SUMO chain interaction is dispensable for the initiation of meiotic DNA recombination, but it is essential for Tel1- and Mec1-dependent Hop1 phosphorylation, which ensures IH recombination by preventing the inter-sister chromatid DNA repair pathway. Our results also indicate that Red1 and Zip1 may directly sandwich the SUMO chains to mediate SC assembly. We suggest that Red1 and SUMO chains function together to couple homologous recombination and Mec1–Tel1 kinase activation with chromosome synapsis during yeast meiosis.     

C. elegans HIM-17 links chromatin modification and competence for initiation of meiotic recombination

Initiation of meiotic recombination by double-strand breaks (DSBs) must occur in a controlled fashion to avoid jeopardizing genome integrity. Here, we identify chromatin-associated protein HIM-17 as a link between chromatin state and DSB formation during C. elegans meiosis. Dependencies of several meiotic prophase events on HIM-17 parallel those seen for DSB-generating enzyme SPO-11: HIM-17 is essential for DSB formation but dispensable for homolog synapsis. Crossovers and chiasmata are eliminated in him-17 null mutants but are restored by artificially induced DSBs, indicating that all components required to convert DSBs into chiasmata are present. Unlike SPO-11, HIM-17 is also required for proper accumulation of histone H3 methylation at lysine 9 on meiotic prophase chromosomes. HIM-17 shares structural features with three proteins that interact genetically with LIN-35/Rb, a known component of chromatin-modifying complexes. Furthermore, DSB levels and incidence of chiasmata can be modulated by loss of LIN-35/Rb. These and other data suggest that chromatin state governs the timing of DSB competence.     

Hanging on to your homolog: the roles of pairing, synapsis and recombination in the maintenance of homolog adhesion

Homologous chromosomes initially undergo weak alignments that bring homologous sequences into register during meiosis. These alignments can be facilitated by two types of mechanisms: interstitial homology searches and telomere-telomere alignments. As prophase (and chromatin compaction) proceeds, these initial pairings or alignments need to be stabilized. In at least some organisms, such as Saccharomyces cerevisiae and S. pombe, these pairings can apparently be maintained by the creation of recombination intermediates. In contrast, synapsis during zygotene may be able to facilitate and/or maintain chromosome pairing even in the absence of exchange in several higher organisms. It thus seems possible that the synaptonemal complex plays a role both in maintaining homolog adhesion during meiotic prophase and, more speculatively, in facilitating meiotic exchange. Received: 15 November 1999; in revised form: 17 January 2000 / Accepted: 18 January 2000     

A mouse homolog of the Saccharomyces cerevisiae meiotic recombination DNA transesterase Spo11p.

The Saccharomyces cerevisiae Spo11 protein is thought to catalyze formation of the DNA double-strand breaks that initiate meiotic recombination. We have cloned cDNA and genomic DNA for a mouse gene encoding a protein with significant sequence similarity to conserved domains found in proteins of the Spo11p family. This putative mouse Spo11 gene maps to the distal region of chromosome 2 (homologous to human chromosome 20q13.2-q13.3) and comprises at least 12 exons, spanning approximately 15-18 kb. Strong expression of the Spo11 message is seen in juvenile and adult testis by RNA in situ hybridization, RT-PCR, and Northern blot, with much weaker expression in thymus and brain. In situ hybridization detects expression in oocytes of embryonic ovary, but not of adult ovary. RT-PCR and in situ hybridization analyses of a time course of juvenile testis development indicate that Spo11 expression begins in early meiotic Prophase I, prior to the pachytene stage, with increasing accumulation of mRNA through the pachytene stage. Taken together, these results strongly suggest that this gene encodes the functional homolog of yeast Spo11p, which in turn suggests that the mechanism of meiotic recombination initiation is conserved between yeast and mammals.     

Differential timing of S phases, X chromosome replication, and meiotic prophase in the C. elegans germ line

The replication of chromosomes in meiosis is an important first step for subsequent chromosomal interactions that promote accurate disjunction in the first of two segregation events to generate haploid gametes. We have developed an assay to monitor DNA replication in vivo in mitotic and meiotic germline nuclei of the nematode Caenorhabditis elegans. Using mutants that affect the mitosis/meiosis switch, we show that meiotic S phase is at least twice as long as mitotic S phase in C. elegans germ cell nuclei. Furthermore, our assay reveals that different regions of the genome replicate at different times, with the heterochromatic-like X chromosomes replicating at a distinct time from the autosomes. Finally, we have exploited S-phase labeling to monitor the timing of progression through meiotic prophase. Meiotic prophase for oocyte production in hermaphrodites lasts 54-60 h. Further, we find that the duration of the pachytene sub-stage is modulated by the presence of sperm. On the other hand, meiotic prophase for sperm production in males is completed by 20-24 h. Possible sources for the sex-specific differences in meiotic prophase kinetics are discussed.     

ZHP-3 acts at crossovers to couple meiotic recombination with synaptonemal complex disassembly and bivalent formation in C. elegans

Crossover recombination and the formation of chiasmata normally ensure the proper segregation of homologous chromosomes during the first meiotic division. zhp-3, the Caenorhabditis elegans ortholog of the budding yeast ZIP3 gene, is required for crossover recombination. We show that ZHP-3 protein localization is highly dynamic. At a key transition point in meiotic prophase, the protein shifts from along the length of the synaptonemal complex (SC) to an asymmetric localization on the SC and eventually becomes restricted to foci that mark crossover recombination events. A zhp-3::gfp transgene partially complements a null mutation and reveals a separation of function; although the fusion protein can promote nearly wild-type levels of recombination, aneuploidy among the progeny is high, indicating defects in meiotic chromosome segregation. The structure of bivalents is perturbed in this mutant, suggesting that the chromosome segregation defect results from an inability to properly remodel chromosomes in response to crossovers. smo-1 mutants exhibit phenotypes similar to zhp-3::gfp mutants at higher temperatures, and smo-1; zhp-3::gfp double mutants exhibit more severe meiotic defects than either single mutant, consistent with a role for SUMO in the process of SC disassembly and bivalent differentiation. We propose that coordination of crossover recombination with SC disassembly and bivalent formation reflects a conserved role of Zip3/ZHP-3 in coupling recombination with SC morphogenesis.     

A C. elegans homolog of the Cockayne syndrome complementation group A gene

Cockayne syndrome (CS) is a debilitating and complex disorder that results from inherited mutations in the CS complementation genes A and B, CSA and CSB. The links between the molecular functions of the CS genes and the complex pathophysiology of CS are as of yet poorly understood and are the subject of intense debate. While mouse models reflect the complexity of CS, studies on simpler genetic models might shed new light on the consequences of CS mutations. Here we describe a functional homolog of the human CSA gene in Caenorhabditis elegans. Similar to its human counterpart, mutations in the nematode csa-1 gene lead to developmental growth defects as a consequence of DNA lesions.     

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.     

Mad1’s ability to interact with Mad2 is essential to regulate and monitor meiotic synapsis in C. elegans

Meiotic homolog synapsis is essential to ensure accurate segregation of chromosomes during meiosis. In C. elegans, proper regulation of synapsis and a checkpoint that monitors synapsis relies on the spindle checkpoint components, Mad1 and Mad2, and Pairing Centers (PCs), cis-acting loci that interact with the nuclear envelope to mobilize chromosomes within the nucleus. Here, we test what specific functions of Mad1 and Mad2 are required to regulate and monitor synapsis. We find that a mutation that prevents Mad1’s localization to the nuclear periphery abolishes the synapsis checkpoint but has no effect on Mad2’s localization to the nuclear periphery or synapsis. By contrast, a mutation that prevents Mad1’s interaction with Mad2 abolishes the synapsis checkpoint, delays synapsis and fails to localize Mad2 to the nuclear periphery. These data indicate that Mad1’s primary role in regulating synapsis is through control of Mad2 and that Mad2 can bind other factors at the nuclear periphery. We also tested whether Mad2’s ability to adopt a specific conformation associated with its activity during spindle checkpoint function is required for its role in meiosis. A mutation that prevents Mad2 from adopting its active conformer fails to localize to the nuclear periphery, abolishes the synapsis checkpoint and exhibits substantial defects in meiotic synapsis. Thus, Mad2, and its regulation by Mad1, is an important regulator of meiotic synapsis in C. elegans.     

A family of zinc-finger proteins is required for chromosome-specific pairing and synapsis during meiosis in C. elegans

Homologous chromosome pairing and synapsis are prerequisite for accurate chromosome segregation during meiosis. Here, we show that a family of four related C2H2 zinc-finger proteins plays a central role in these events in C. elegans. These proteins are encoded within a tandem gene cluster. In addition to the X-specific HIM-8 protein, three additional paralogs collectively mediate the behavior of the five autosomes. Each chromosome relies on a specific member of the family to pair and synapse with its homolog. These "ZIM" proteins concentrate at special regions called meiotic pairing centers on the corresponding chromosomes. These sites are dispersed along the nuclear envelope during early meiotic prophase, suggesting a role analogous to the telomere-mediated meiotic bouquet in other organisms. To gain insight into the evolution of these components, we characterized homologs in C. briggsae and C. remanei, which revealed changes in copy number of this gene family within the nematode lineage.     

A cytoplasmic dynein heavy chain is required for oscillatory nuclear movement of meiotic prophase and efficient meiotic recombination in fission yeast.

Meiotic recombination requires pairing of homologous chromosomes, the mechanisms of which remain largely unknown. When pairing occurs during meiotic prophase in fission yeast, the nucleus oscillates between the cell poles driven by astral microtubules. During these oscillations, the telomeres are clustered at the spindle pole body (SPB), located at the leading edge of the moving nucleus and the rest of each chromosome dangles behind. Here, we show that the oscillatory nuclear movement of meiotic prophase is dependent on cytoplasmic dynein. We have cloned the gene encoding a cytoplasmic dynein heavy chain of fission yeast. Most of the cells disrupted for the gene show no gross defect during mitosis and complete meiosis to form four viable spores, but they lack the nuclear movements of meiotic prophase. Thus, the dynein heavy chain is required for these oscillatory movements. Consistent with its essential role in such nuclear movement, dynein heavy chain tagged with green fluorescent protein (GFP) is localized at astral microtubules and the SPB during the movements. In dynein-disrupted cells, meiotic recombination is significantly reduced, indicating that the dynein function is also required for efficient meiotic recombination. In accordance with the reduced recombination, which leads to reduced crossing over, chromosome missegregation is increased in the mutant. Moreover, both the formation of a single cluster of centromeres and the colocalization of homologous regions on a pair of homologous chromosomes are significantly inhibited in the mutant. These results strongly suggest that the dynein-driven nuclear movements of meiotic prophase are necessary for efficient pairing of homologous chromosomes in fission yeast, which in turn promotes efficient meiotic recombination.     

Chromosomal synapsis and the meiotic process in male mesquite lizards, Sceloporus grammicus complex.

Meiosis in males of the F5 cytotype of Sceloporus grammicus was examined through the analysis of synaptonemal complexes (SCs), diakinetic (metaphase I) nuclei, and secondary spermatocytes (metaphase II configurations). These data allowed the establishment of criteria for substaging of zygonema and pachynema, morphological characterization of the SC complement, and comparison of the orientation and segregation of the autosomes and sex chromosomes. The analysis of nuclei from all stages of meiotic prophase I (leptonema through diakinesis) provided a useful means of partitioning the temporal sequence of early meiotic events. Three substages of zygonema (Z1-Z3) were established, based on the extent of synapsis of the microchromosomal and macrochromosomal elements. Synaptic initiation of the autosomes and sex chromosomes was synchronous. Two patterns of macrochromosomal synapsis were observed. Whereas synapsis of the biarmed elements was biterminal (i.e., progressing from both ends of the homologs), synapsis of the acrocentric elements was uniterminal involving only the distal (noncentromeric) ends of the homologs. Unique sex-chromosomal characteristics were not observed in S. grammicus and, therefore, the substaging of pachynema was based on subjective criteria. Examination of diakinesis--metaphase I and metaphase II configurations indicated low levels of diakinetic irregularities with balanced segregation of the autosomal bivalents and the sex-chromosomal trivalent.     

CRA-1 uncovers a double-strand break-dependent pathway promoting the assembly of central region proteins on chromosome axes during C. elegans meiosis

The synaptonemal complex (SC), a tripartite proteinaceous structure that forms between homologous chromosomes during meiosis, is crucial for faithful chromosome segregation. Here we identify CRA-1, a novel and conserved protein that is required for the assembly of the central region of the SC during C. elegans meiosis. In the absence of CRA-1, central region components fail to extensively localize onto chromosomes at early prophase and instead mostly surround the chromatin at this stage. Later in prophase, central region proteins polymerize along chromosome axes, but for the most part fail to connect the axes of paired homologous chromosomes. This defect results in an inability to stabilize homologous pairing interactions, altered double-strand break (DSB) repair progression, and a lack of chiasmata. Surprisingly, DSB formation and repair are required to promote the polymerization of the central region components along meiotic chromosome axes in cra-1 mutants. In the absence of both CRA-1 and any one of the C. elegans homologs of SPO11, MRE11, RAD51, or MSH5, the polymerization observed along chromosome axes is perturbed, resulting in the formation of aggregates of the SC central region proteins. While radiation-induced DSBs rescue this polymerization in cra-1; spo-11 mutants, they fail to do so in cra-1; mre-11, cra-1; rad-51, and cra-1; msh-5 mutants. Taken together, our studies place CRA-1 as a key component in promoting the assembly of a tripartite SC structure. Moreover, they reveal a scenario in which DSB formation and repair can drive the polymerization of SC components along chromosome axes in C. elegans.