The molecular features of chromosome pairing at meiosis: the polyploid challenge using wheat as a reference

During meiosis, chromosome numbers are halved, leading to haploid gametes, a process that is crucial for the maintenance of a stable genome through successive generations. The process for the accurate segregation of the homologues starts in pre-meiosis as each homologue is replicated and the respective products are held together as two sister chromatids via specific cohesion proteins. At the start of meiosis, each chromosome must recognise its homologue from amongst all the chromosomes present in the nucleus and then associate or pair with that homologue. This process of homologue recognition in meiosis is more complicated in polyploids because of the greater number of related chromosomes. Despite the presence of these related chromosomes, for polyploids such as wheat to produce viable gametes, they must behave as diploids during meiosis with only true homologues pairing. In this review, the relationship between the Ph1 cyclin-dependent kinase (CDK)-like genes in wheat and the CDK2 genes in mammals and their involvement in controlling this process at meiosis is examined.     

Nuclear architecture and chromosome dynamics in the search of the pairing partner in meiosis in plants

The formation of haploid gametes in organisms with sexual reproduction requires regular bivalent chromosome pairing in meiosis. In many species, homologous chromosomes occupy separate territories at the onset of meiosis. To be paired at metaphase I, they need to be brought into a close proximity for interactions that include homology recognition and the establishment of some form of bonds. How homologues find each other is one of the least understood meiotic events. Plant species with large or medium sized genomes, such as wheat or maize, are excellent materials for the cytological analysis of chromosome dynamics at early meiosis, but genes that control meiosis have been identified mainly in small genome species such as Arabidopsis thaliana. This review is focused on the contribution studies on plants are providing to the knowledge of the initial steps of the meiotic process.     

Chromosomes associate premeiotically and in xylem vessel cells via their telomeres and centromeres in diploid rice (<Emphasis Type="Italic">Oryza sativa</Emphasis>)

Studies of the meiosis of diploid plants such as Arabidopsis, maize and diploid progenitors of wheat have revealed no premeiotic association of chromosomes. Premeiotic and somatic association of chromosomes has only been previously observed in the anther tissues and xylem vessel cells of developing roots in polyploid plants such as hexaploid and tetraploid wheat, polyploid relatives of wheat and artificial polyploids made from the progenitor diploids of wheat. This suggested that this association was confined specifically to polyploids or was induced by polyploidy. However, we developed procedures for in situ hybridization on structurally well-preserved tissue sections of rice, and analysed two diploid rice species (Oryza sativa and O. punctata). Contrary to expectation, this has revealed that centromeres and telomeres also associate both in the xylem vessel cells of developing root and in undifferentiated anther cells in these diploids. However, in contrast to wheat and related polyploids, where the initial association in undifferentiated anthers is between either non-homologous or related chromosomes, and not homologous chromosomes, the initial association of rice chromosomes seems to be between homologues. Thus, in contrast to the diploid dicot model Arabidopsis, meiotic studies on the diploid model cereal, rice, will now need to take into account the effects of premeiotic chromosome association.Pilar Prieto and Ana Paula Santos are joint first authors.     

Suppression of Homoeologous Pairing by <Emphasis Type="Italic">B</Emphasis> Chromosomes in a <Emphasis Type="Italic">Lolium</Emphasis> Species Hybrid

INTERSPECIFIC hybridization together with polyploidy has been an important force in the evolution of many of our graminaceous crop plants. Both wheat (Triticum aestivum) and oats (Avena sativa), for example, are natural allohexaploids derived in each case from the hybridization of three separate but related diploid species. The efforts of plant breeders to synthesize stable and fertile polyploids of this kind have, on the whole, been unsuccessful. The main reason for this is that whereas meiosis in natural allopolyploids such as wheat is extremely regular this is not the case with “synthetic” polyploids. In wheat precise control over pairing at meiosis is achieved by a gene or a cluster of genes on chromosome SB. The gene acts by restricting the pairing to homologous chromosomes with the result that only bivalents are formed, disjunction is regular and inheritance is completely disomic^1,2. In the artificial polyploids at pachytene there is pairing between both homologous chromosomes (from the same species) and “corresponding” homoeologous chromosomes (from different species). The result is an extremely irregular metaphase 1 comprising multivalents and univalents as well as bivalents. Segregation is irregular and a certain degree of infertility is inevitable.     

Dynamics of DNA Replication during Premeiosis and Early Meiosis in Wheat

Meiosis is a specialised cell division that involves chromosome replication, two rounds of chromosome segregation and results in the formation of the gametes. Meiotic DNA replication generally precedes chromosome pairing, recombination and synapsis in sexually developing eukaryotes. In this work, replication has been studied during premeiosis and early meiosis in wheat using flow cytometry, which has allowed the quantification of the amount of DNA in wheat anther in each phase of the cell cycle during premeiosis and each stage of early meiosis. Flow cytometry has been revealed as a suitable and user-friendly tool to detect and quantify DNA replication during early meiosis in wheat. Chromosome replication was detected in wheat during premeiosis and early meiosis until the stage of pachytene, when chromosomes are associated in pairs to further recombine and correctly segregate in the gametes. In addition, the effect of the Ph1 locus, which controls chromosome pairing and affects replication in wheat, was also studied by flow cytometry. Here we showed that the Ph1 locus plays an important role on the length of meiotic DNA replication in wheat, particularly affecting the rate of replication during early meiosis in wheat.     

Meiotic restitution in wheat polyhaploids (amphihaploids): a potent evolutionary force

Polyploidy is well recognized as a major force in plant speciation. Among the polyploids in nature, allopolyploids are preponderant and include important crop plants like bread wheat, Triticum aestivum L. (2n = 6x = 42; AABBDD genomes). Allopolyploidy must result through concomitant or sequential events that entail interspecific or intergeneric hybridization and chromosome doubling in the resultant hybrids. To gain insight into the mechanism of evolution of wheat, we extracted polyhaploids of 2 cultivars, Chinese Spring (CS) and Fukuhokomugi (Fuko), of bread wheat by crossing them with maize, Zea mays L. ssp. mays. The derived Ph1-polyhaploids (2n = 3x = 21; ABD) showed during meiosis mostly univalents, which produced first-division restitution (FDR) nuclei that in turn gave rise to unreduced (2n) male gametes with 21 chromosomes. The haploids on maturity set some viable seed. The mean number of seeds per spike was 1.45 +/- 0.161 in CS and 2.3 +/- 0.170 in Fuko. Mitotic chromosome preparations from root tips of the derived plantlets revealed 2n = 42 chromosomes, that is, twice that of the parental polyhaploid, which indicated that they arose by fusion of unreduced male and female gametes formed by the polyhaploid. The Ph1-induced univalency must have produced 2n gametes and hence bilateral sexual polyploidization and reconstitution of disomic bread wheat. These findings highlight the quantum jump by which bread wheat evolved from durum wheat in nature. Thus, bread wheat offers an excellent example of rapid evolution by allopolyploidy. In the induced polyhaploids (ABD) that are equivalent of amphihaploids, meiotic phenomena such as FDR led to regeneration of parental bread wheat, perhaps a simulation of the evolutionary steps that occurred in nature at the time of the origin of hexaploid wheat.     

Terminal regions of wheat chromosomes select their pairing partners in meiosis

Many plant species, including important crops like wheat, are polyploids that carry more than two sets of genetically related chromosomes capable of meiotic pairing. To safeguard a diploid-like behavior at meiosis, many polyploids evolved genetic loci that suppress incorrect pairing and recombination of homeologues. The Ph1 locus in wheat was proposed to ensure homologous pairing by controlling the specificity of centromere associations that precede chromosome pairing. Using wheat chromosomes that carry rye centromeres, we show that the centromere associations in early meiosis are not based on homology and that the Ph1 locus has no effect on such associations. Although centromeres indeed undergo a switch from nonhomologous to homologous associations in meiosis, this process is driven by the terminally initiated synapsis. The centromere has no effect on metaphase I chiasmate chromosome associations: homologs with identical or different centromeres, in the presence and absence of Ph1, pair the same. A FISH analysis of the behavior of centromeres and distal chromomeres in telocentric and bi-armed chromosomes demonstrates that it is not the centromeric, but rather the subtelomeric, regions that are involved in the correct partner recognition and selection.     

A bivalent polyploid model for linkage analysis in outcrossing tetraploids

Polyploids can be classified as either allopolyploids or autopolyploids based on their presumed origins. From a perspective of linkage analysis, however, the nature of polyploids can be better described as bivalent polyploids, in which two chromosomes pair at meiosis, multivalent polyploids, in which more than two chromosomes pair, and general polyploids, in which bivalent and multivalent formations occur simultaneously. In this paper, we develop a statistical method for linkage analysis of polymorphic markers in bivalent polyploids. This method takes into account a unique cytological pairing mechanism for the formation of diploid gametes in tetraploids-preferential bivalent pairings at meiosis during which two homologous chromosomes pair with a higher probability than two homoeologous chromosomes. The higher frequency of homologous over homoeologous pairing, defined as the preferential pairing factor, affects the segregation patterns and linkage analysis of different genes on the same chromosome. A maximum likelihood method implemented with the EM algorithm is proposed to simultaneously estimate linkage and parental linkage phases over a pair of markers from any possible marker cross type between two outbred bivalent tetraploid parents demonstrating preferential bivalent pairings. Simulation studies display that the method can be well used to estimate the recombination fraction between different marker types and the preferential pairing factor typical of bivalent tetraploids. The implications of this method for current genome projects in polyploid species are discussed.     

SYCE2 is required for synaptonemal complex assembly, double strand break repair, and homologous recombination

Synapsis is the process by which paired chromosome homologues closely associate in meiosis before crossover. In the synaptonemal complex (SC), axial elements of each homologue connect through molecules of SYCP1 to the central element, which contains the proteins SYCE1 and -2. We have derived mice lacking SYCE2 protein, producing males and females in which meiotic chromosomes align and axes form but do not synapse. Sex chromosomes are unaligned, not forming a sex body. Additionally, markers of DNA breakage and repair are retained on the axes, and crossover is impaired, culminating in both males and females failing to produce gametes. We show that SC formation can initiate at sites of SYCE1/SYCP1 localization but that these points of initiation cannot be extended in the absence of SYCE2. SC assembly is thus dependent on SYCP1, SYCE1, and SYCE2. We provide a model to explain this based on protein-protein interactions.     

Chromosome banding in amphibia

The distribution of constitutive heterochromatin on the chromosomes of Triturus a. alpestris, T. v. vulgaris and T. h. helveticus (Amphibia, Urodela) was investigated. Sex-specific chromosomes were determined in the karyotypes of T. a. alpestris (chromosomes 4) and T. v. vulgaris (chromosomes 5). The male animals have one heteromorphic chromosome pair, of which only one homologue displays heterochromatic telomeres in the long arms; the telomeres of the other homologue are euchromatic. This chromosome pair is always homomorphic and without telomeric heterochromatin in the female animals. There is a highly reduced crossing-over frequency between the heteromorphic chromosome arms in the male meiosis of T. a. alpestris; in T. v. vulgaris no crossing-over at all occurs between the heteromorphic chromosome arms. No heteromorphisms between the homologues exist on the corresponding lampbrush chromosomes of the female meiosis. In T. h. helveticus no sex-specific heteromorphism of the constitutive heterochromatin could be determined. The male animals of this species, however, already possess a chromosome pair with a greatly reduced frequency of chiasma-formation in the long arms. The C-band patterns and the pairing configurations of the sex-specific chromosomes in the male meiosis indicate an XX/XY-type of sex-determination for the three species. A revision of the literature about experimental interspecies hybridizations, gonadic structure of haploid and polyploid animals, and sex-linked genes yielded further evidence in favor of male heterogamety. The results moreover suggest that the heterochromatinization of the Y-chromosome was the primary step in the evolution of the sex chromosomes.     

Inducing chromosome pairing through premature condensation: analysis of wheat interspecific hybrids

At the onset of meiosis, chromosomes first decondense and then condense as the process of recognition and intimate pairing occurs between homologous chromosomes. We show here that okadaic acid, a drug known to induce chromosome condensation, can be introduced into wheat interspecific hybrids prior to meiosis to induce chromosome pairing. This pairing occurs in the presence of the Ph1 locus, which usually suppresses pairing of related chromosomes and which we show here delays condensation. Thus the timing of chromosome condensation during the onset of meiosis is an important factor in controlling chromosome pairing.     

Condensin restructures chromosomes in preparation for meiotic divisions

The production of haploid gametes from diploid germ cells requires two rounds of meiotic chromosome segregation after one round of replication. Accurate meiotic chromosome segregation involves the remodeling of each pair of homologous chromosomes around the site of crossover into a highly condensed and ordered structure. We showed that condensin, the protein complex needed for mitotic chromosome compaction, restructures chromosomes during meiosis in Caenorhabditis elegans. In particular, condensin promotes both meiotic chromosome condensation after crossover recombination and the remodeling of sister chromatids. Condensin helps resolve cohesin-independent linkages between sister chromatids and alleviates recombination-independent linkages between homologues. The safeguarding of chromosome resolution by condensin permits chromosome segregation and is crucial for the formation of discrete, individualized bivalent chromosomes.     

High-throughput knockout screen in Schizosaccharomyces pombe identifies a novel gene required for efficient homolog disjunction during meiosis I

Meiosis is the process which produces haploid gametes from diploid precursor cells. This reduction of chromosome number is achieved by two successive divisions. Whereas homologs segregate during meiosis I, sister chromatids segregate during meiosis II. To identify novel proteins required for proper segregation of chromosomes during meiosis, we applied a high-throughput knockout technique to delete 87 S. pombe genes whose expression is upregulated during meiosis and analyzed the mutant phenotypes. Using this approach, we identified a new protein, Dil1, which is required to prevent meiosis I homolog non-disjunction. We show that Dil1 acts in the dynein pathway to promote oscillatory nuclear movement during meiosis.     

Meiotically asynapsis-induced aneuploidy in autopolyploid Arabidopsis thaliana

The patterns of homologue segregation are the basis for euploidy or aneuploidy formation in diploids and allo-/auto-polyploids. Homologue segregation in diploids resembles that in allopolyploids during meiosis; however, meiotic chromosome behavior in autopolyploids is complicated by multiplication of homologous chromosome components. Obviously, loss of single chromosomes (or segmented chromosomes) frequently leads to abortion of reproductive gametes in diploids and allopolyploids. In contrast, the consequence of chromosome loss in autopolyploids is effortlessly compensated for by the presence of multiplied chromosome complements. Here, we use the meiotically asynaptic gene asy1, in combination with polyploidization, to elucidate aneuploidy formation in autotetraploid Arabidopsis. The results indicate that, due to homologous asynapsis in meiotic prophase I, retarded chromosome losses could induce aneuploidy during gametogenesis in autotetraploid asy1. The severe loss of individual chromosomes probably reaches the haploid genome among selfed or backcrossed progeny, leading to stochastic chromosome loss in Arabidopsis. Reciprocal crosses of autotetraploid asy1 with wild-type prove a pathway of duoparental transmission of aneuploidy (hypoploidy and hyperploidy). Viable hypoploids over-transmit via male gametes; conversely, viable hyperploids transmit mainly in female gametogenesis. This result suggests a more stringent maternal restriction of ploidy transmission in autopolyploid Arabidopsis.     

Homoeologous Chromosome Sorting and Progression of Meiotic Recombination in Brassica napus: Ploidy Does Matter!

Meiotic recombination is the fundamental process that produces balanced gametes and generates diversity within species. For successful meiosis, crossovers must form between homologous chromosomes. This condition is more difficult to fulfill in allopolyploid species, which have more than two sets of related chromosomes (homoeologs). Here, we investigated the formation, progression, and completion of several key hallmarks of meiosis in Brassica napus (AACC), a young polyphyletic allotetraploid crop species with closely related homoeologous chromosomes. Altogether, our results demonstrate a precocious and efficient sorting of homologous versus homoeologous chromosomes during early prophase I in two representative B. napus accessions that otherwise show a genotypic difference in the progression of homologous recombination. More strikingly, our detailed comparison of meiosis in near isogenic allohaploid and euploid plants showed that the mechanism(s) promoting efficient chromosome sorting in euploids is adjusted to promote crossover formation between homoeologs in allohaploids. This suggests that, in contrast to other polyploid species, chromosome sorting is context dependent in B. napus.     

Meiosis in allopolyploids -- the importance of 'Teflon' chromosomes

Polyploids possess two or more sets of related chromosomes as a result of either the doubling of chromosomes following sexual hybridization within the same species (autopolyploidy), or between closely related species containing related but not completely homologous (homoeologous) genomes (allopolyploidy). For allopolyploids to produce viable gametes and be fertile, they must behave as diploids during meiosis, so that only identical chromosomes (homologues) pair. A solution to this problem is an enhanced ability to resolve incorrect pairing, which in turn promotes correct pairing. This gives nonhomologous chromosomes an almost 'Teflon'-like status, so that only the correct pairs 'stick'.     

Rec8 cleavage by separase is required for meiotic nuclear divisions in fission yeast

Sister chromatid cohesion in meiosis is established by cohesin complexes, including the Rec8 subunit. During meiosis I, sister chromatid cohesion is destroyed along the chromosome arms to release connections of recombined homologous chromosomes (homologues), whereas centromeric cohesion persists until it is finally destroyed at anaphase II. In fission yeast, as in mammals, distinct cohesin complexes are used depending on the chromosomal region; Rec8 forms a complex with Rec11 (equivalent to SA3) mainly along chromosome arms, while Psc3 (equivalent to SA1 and SA2) forms a complex mainly in the vicinity of the centromeres. Here we show that separase activation and resultant Rec8 cleavage are required for meiotic chromosome segregation in fission yeast. A non-cleavable form of Rec8 blocks disjunction of homologues at meiosis I. However, displacing non-cleavable Rec8 restrictively from the chromosome arm by genetically depleting Rec11 alleviated the blockage of homologue segregation, but not of sister segregation. We propose that the segregation of homologues at meiosis I and of sisters at meiosis II requires the cleavage of Rec8 along chromosome arms and at the centromeres, respectively.     

Leagues of their own: sexually dimorphic features of meiotic prophase I

Meiosis is a conserved cell division process that is used by sexually reproducing organisms to generate haploid gametes. Males and females produce different end products of meiosis: eggs (females) and sperm (males). In addition, these unique end products demonstrate sex-specific differences that occur throughout meiosis to produce the final genetic material that is packaged into distinct gametes with unique extracellular morphologies and nuclear sizes. These sexually dimorphic features of meiosis include the meiotic chromosome architecture, in which both the lengths of the chromosomes and the requirement for specific meiotic axis proteins being different between the sexes. Moreover, these changes likely cause sex-specific changes in the recombination landscape with the sex that has the longer chromosomes usually obtaining more crossovers. Additionally, epigenetic regulation of meiosis may contribute to sexually dimorphic recombination landscapes. Here we explore the sexually dimorphic features of both the chromosome axis and crossing over for each stage of meiotic prophase I in Mus musculus, Caenorhabditis elegans, and Arabidopsis thaliana. Furthermore, we consider how sex-specific changes in the meiotic chromosome axes and the epigenetic landscape may function together to regulate crossing over in each sex, indicating that the mechanisms controlling crossing over may be different in oogenesis and spermatogenesis.

     

Homologous chromosome interactions in meiosis: diversity amidst conservation

Proper chromosome segregation is crucial for preventing fertility problems, birth defects and cancer. During mitotic cell divisions, sister chromatids separate from each other to opposite poles, resulting in two daughter cells that each have a complete copy of the genome. Meiosis poses a special problem in which homologous chromosomes must first pair and then separate at the first meiotic division before sister chromatids separate at the second meiotic division. So, chromosome interactions between homologues are a unique feature of meiosis and are essential for proper chromosome segregation. Pairing and locking together of homologous chromosomes involves recombination interactions in some cases, but not in others. Although all organisms must match and lock homologous chromosomes to maintain genome integrity throughout meiosis, recent results indicate that the underlying mechanisms vary in different organisms.     

Polyploidy induces centromere association

Many species exhibit polyploidy. The presence of more than one diploid set of similar chromosomes in polyploids can affect the assortment of homologous chromosomes, resulting in unbalanced gametes. Therefore, a mechanism is required to ensure the correct assortment and segregation of chromosomes for gamete formation. Ploidy has been shown to affect gene expression. We present in this study an example of a major effect on a phenotype induced by ploidy within the Triticeae. We demonstrate that centromeres associate early during anther development in polyploid species. In contrast, centromeres in diploid species only associate at the onset of meiotic prophase. We propose that this mechanism provides a potential route by which chromosomes can start to be sorted before meiosis in polyploids. This explains previous reports indicating that meiotic prophase is shorter in polyploids than in their diploid progenitors. Even artificial polyploids exhibit this phenotype, suggesting that the mechanism must be present in diploids, but only expressed in the presence of more than one diploid set of chromosomes.