Per-Nucleus Crossover Covariation and Implications for Evolution.

Crossing over is a nearly universal feature of sexual reproduction. Here, analysis of crossover numbers on a per-chromosome and per-nucleus basis reveals a fundamental, evolutionarily conserved feature of meiosis: within individual nuclei, crossover frequencies covary across different chromosomes. This effect results from per-nucleus covariation of chromosome axis lengths. Crossovers can promote evolutionary adaptation. However, the benefit of creating favorable new allelic combinations must outweigh the cost of disrupting existing favorable combinations. Covariation concomitantly increases the frequencies of gametes with especially high, or especially low, numbers of crossovers, and thus might concomitantly enhance the benefits of crossing over while reducing its costs. A four-locus population genetic model suggests that such an effect can pertain in situations where the environment fluctuates: hyper-crossover gametes are advantageous when the environment changes while hypo-crossover gametes are advantageous in periods of environmental stasis. These findings reveal a new feature of the basic meiotic program and suggest a possible adaptive advantage.

Inefficient Crossover Maturation Underlies Elevated Aneuploidy in Human Female Meiosis.

Meiosis is the cellular program that underlies gamete formation. For this program, crossovers between homologous chromosomes play an essential mechanical role to ensure regular segregation. We present a detailed study of crossover formation in human male and female meiosis, enabled by modeling analysis. Results suggest that recombination in the two sexes proceeds analogously and efficiently through most stages. However, specifically in female (but not male), ∼25% of the intermediates that should mature into crossover products actually fail to do so. Further, this "female-specific crossover maturation inefficiency" is inferred to make major contributions to the high level of chromosome mis-segregation and resultant aneuploidy that uniquely afflicts human female oocytes (e.g., giving Down syndrome). Additionally, crossover levels on different chromosomes in the same nucleus tend to co-vary, an effect attributable to global per-nucleus modulation of chromatin loop size. Maturation inefficiency could potentially reflect an evolutionary advantage of increased aneuploidy for human females.

Canonical and noncanonical roles of Hop1 are crucial for meiotic prophase in the fungus Sordaria macrospora.

We show here that in the fungus Sordaria macrospora, the meiosis-specific HORMA-domain protein Hop1 is not essential for the basic early events of chromosome axis development, recombination initiation, or recombination-mediated homolog coalignment/pairing. In striking contrast, Hop1 plays a critical role at the leptotene/zygotene transition which is defined by transition from pairing to synaptonemal complex (SC) formation. During this transition, Hop1 is required for maintenance of normal axis structure, formation of SC from telomere to telomere, and development of recombination foci. These hop1Δ mutant defects are DSB dependent and require Sme4/Zip1-mediated progression of the interhomolog interaction program, potentially via a pre-SC role. The same phenotype occurs not only in hop1Δ but also in absence of the cohesin Rec8 and in spo76-1, a non-null mutant of cohesin-associated Spo76/Pds5. Thus, Hop1 and cohesins collaborate at this crucial step of meiotic prophase. In addition, analysis of 4 non-null mutants that lack this transition defect reveals that Hop1 also plays important roles in modulation of axis length, homolog-axis juxtaposition, interlock resolution, and spreading of the crossover interference signal. Finally, unexpected variations in crossover density point to the existence of effects that both enhance and limit crossover formation. Links to previously described roles of the protein in other organisms are discussed.

Sister cohesion and structural axis components mediate homolog bias of meiotic recombination.

Meiotic double-strand break (DSB)-initiated recombination must occur between homologous maternal and paternal chromosomes ("homolog bias"), even though sister chromatids are present. Through physical recombination analyses, we show that sister cohesion, normally mediated by meiotic cohesin Rec8, promotes "sister bias"; that meiosis-specific axis components Red1/Mek1kinase counteract this effect, thereby satisfying an essential precondition for homolog bias; and that other components, probably recombinosome-related, directly ensure homolog partner selection. Later, Rec8 acts positively to ensure maintenance of bias. These complexities mirror opposing dictates for global sister cohesion versus local separation and differentiation of sisters at recombination sites. Our findings support DSB formation within axis-tethered recombinosomes containing both sisters and ensuing programmed sequential release of "first" and "second" DSB ends. First-end release would create a homology-searching "tentacle." Rec8 and Red1/Mek1 also independently license recombinational progression and abundantly localize to different domains. These domains could comprise complementary environments that integrate inputs from DSB repair and mitotic chromosome morphogenesis into the complete meiotic program.

Dna2 removes toxic ssDNA-RPA filaments generated from meiotic recombination-associated DNA synthesis.

During the repair of DNA double-strand breaks (DSBs), de novo synthesized DNA strands can displace the parental strand to generate single-strand DNAs (ssDNAs). Many programmed DSBs and thus many ssDNAs occur during meiosis. However, it is unclear how these ssDNAs are removed for the complete repair of meiotic DSBs. Here, we show that meiosis-specific depletion of Dna2 (dna2-md) results in an abundant accumulation of RPA and an expansion of RPA from DSBs to broader regions in Saccharomyces cerevisiae. As a result, DSB repair is defective and spores are inviable, although the levels of crossovers/non-crossovers seem to be unaffected. Furthermore, Dna2 induction at pachytene is highly effective in removing accumulated RPA and restoring spore viability. Moreover, the depletion of Pif1, an activator of polymerase δ required for meiotic recombination-associated DNA synthesis, and Pif1 inhibitor Mlh2 decreases and increases RPA accumulation in dna2-md, respectively. In addition, blocking DNA synthesis during meiotic recombination dramatically decreases RPA accumulation in dna2-md. Together, our findings show that meiotic DSB repair requires Dna2 to remove ssDNA-RPA filaments generated from meiotic recombination-associated DNA synthesis. Additionally, we showed that Dna2 also regulates DSB-independent RPA distribution.

The ubiquitin-proteasome system regulates meiotic chromosome organization.

Meiotic crossover (CO) recombination is tightly regulated by chromosome architecture to ensure faithful chromosome segregation and to reshuffle alleles between parental chromosomes for genetic diversity of progeny. However, regulation of the meiotic chromosome loop/axis organization is poorly understood. Here, we identify a molecular pathway for axis length regulation. We show that the cohesin regulator Pds5 can interact with proteasomes. Meiosis-specific depletion of proteasomes and/or Pds5 results in a similarly shortened chromosome axis, suggesting proteasomes and Pds5 regulate axis length in the same pathway. Protein ubiquitination is accumulated in pds5 and proteasome mutants. Moreover, decreased chromosome axis length in these mutants can be largely rescued by decreasing ubiquitin availability and thus decreasing protein ubiquitination. Further investigation reveals that two ubiquitin E3 ligases, SCF (Skp­Cullin­F-box) and Ufd4, are involved in this Pds5­ubiquitin/proteasome pathway to cooperatively control chromosome axis length. These results support the hypothesis that ubiquitination of chromosome proteins results in a shortened chromosome axis, and cohesin­Pds5 recruits proteasomes onto chromosomes to regulate ubiquitination level and thus axis length. These findings reveal an unexpected role of the ubiquitin­proteasome system in meiosis and contribute to our knowledge of how Pds5 regulates meiotic chromosome organization. A conserved regulatory mechanism probably exists in higher eukaryotes.

Negative supercoils regulate meiotic crossover patterns in budding yeast.

Interference exists ubiquitously in many biological processes. Crossover interference patterns meiotic crossovers, which are required for faithful chromosome segregation and evolutionary adaption. However, what the interference signal is and how it is generated and regulated is unknown. We show that yeast top2 alleles which cannot bind or cleave DNA accumulate a higher level of negative supercoils and show weaker interference. However, top2 alleles which cannot religate the cleaved DNA or release the religated DNA accumulate less negative supercoils and show stronger interference. Moreover, the level of negative supercoils is negatively correlated with crossover interference strength. Furthermore, negative supercoils preferentially enrich at crossover-associated Zip3 regions before the formation of meiotic DNA double-strand breaks, and regions with more negative supercoils tend to have more Zip3. Additionally, the strength of crossover interference and homeostasis change coordinately in mutants. These findings suggest that the accumulation and relief of negative supercoils pattern meiotic crossovers.

MEIOK21 regulates oocyte quantity and quality via modulating meiotic recombination.

The reproductive life span of females is largely determined by the number and quality of oocytes. Previously, we identified MEIOK21 as a meiotic recombination regulator required for male fertility. Here, we characterize the important roles of MEIOK21 in regulating female meiosis and oocyte number and quality. MEIOK21 localizes at recombination sites as a component of recombination bridges in oogenesis like in spermatogenesis. Meiok21-/- female mice show subfertility. Consistently, the size of the primordial follicle pool in Meiok21-/- females is only ~40% of wild-type females because a great number of oocytes with defects in meiotic recombination and/or synapsis are eliminated. Furthermore, the numbers of primordial and growing follicles show a more marked decrease in an age-dependent manner compared with wild-type females. Further analysis shows Meiok21-/- oocytes also have reduced rates of germinal vesicle breakdown and the first polar body extrusion when cultured in vitro, indicating poor oocyte quality. Additionally, Meiok21-/- oocytes have more chromosomes bearing a single distally localized crossover (chiasmata), suggesting a possible defect in crossover maturation. Taken together, our findings indicate critical roles for MEIOK21 in ensuring the number and quality of oocytes in the follicles.

ESA1 regulates meiotic chromosome axis and crossover frequency via acetylating histone H4.

Meiotic recombination is integrated into and regulated by meiotic chromosomes, which is organized as loop/axis architecture. However, the regulation of chromosome organization is poorly understood. Here, we show Esa1, the NuA4 complex catalytic subunit, is constitutively expressed and localizes on chromatin loops during meiosis. Esa1 plays multiple roles including homolog synapsis, sporulation efficiency, spore viability, and chromosome segregation in meiosis. Detailed analyses show the meiosis-specific depletion of Esa1 results in decreased chromosome axis length independent of another axis length regulator Pds5, which further leads to a decreased number of Mer2 foci, and consequently a decreased number of DNA double-strand breaks, recombination intermediates, and crossover frequency. However, Esa1 depletion does not impair the occurrence of the obligatory crossover required for faithful chromosome segregation, or the strength of crossover interference. Further investigations demonstrate Esa1 regulates chromosome axis length via acetylating the N-terminal tail of histone H4 but not altering transcription program. Therefore, we firstly show a non-chromosome axis component, Esa1, acetylates histone H4 on chromatin loops to regulate chromosome axis length and consequently recombination frequency but does not affect the basic meiotic recombination process. Additionally, Esa1 depletion downregulates middle induced meiotic genes, which probably causing defects in sporulation and chromosome segregation.

Meiotic chromosome organization and crossover patterns†.

Meiosis is the foundation of sexual reproduction, and crossover recombination is one hallmark of meiosis. Crossovers establish the physical connections between homolog chromosomes (homologs) for their proper segregation and exchange DNA between homologs to promote genetic diversity in gametes and thus progenies. Aberrant crossover patterns, e.g., absence of the obligatory crossover, are the leading cause of infertility, miscarriage, and congenital disease. Therefore, crossover patterns have to be tightly controlled. During meiosis, loop/axis organized chromosomes provide the structural basis and regulatory machinery for crossover patterning. Accumulating evidence shows that chromosome axis length regulates the numbers and the positions of crossovers. In addition, recent studies suggest that alterations in axis length and the resultant alterations in crossover frequency may contribute to evolutionary adaptation. Here, current advances regarding these issues are reviewed, the possible mechanisms for axis length regulating crossover frequency are discussed, and important issues that need further investigations are suggested.

MEIOK21: a new component of meiotic recombination bridges required for spermatogenesis.

Repair of DNA double-strand breaks (DSBs) with homologous chromosomes is a hallmark of meiosis that is mediated by recombination 'bridges' between homolog axes. This process requires cooperation of DMC1 and RAD51 to promote homology search and strand exchange. The mechanism(s) regulating DMC1/RAD51-ssDNA nucleoprotein filament and the components of 'bridges' remain to be investigated. Here we show that MEIOK21 is a newly identified component of meiotic recombination bridges and is required for efficient formation of DMC1/RAD51 foci. MEIOK21 dynamically localizes on chromosomes from on-axis foci to 'hanging foci', then to 'bridges', and finally to 'fused foci' between homolog axes. Its chromosome localization depends on DSBs. Knockout of Meiok21 decreases the numbers of HSF2BP and DMC1/RAD51 foci, disrupting DSB repair, synapsis and crossover recombination and finally causing male infertility. Therefore, MEIOK21 is a novel recombination factor and probably mediates DMC1/RAD51 recruitment to ssDNA or their stability on chromosomes through physical interaction with HSF2BP.

1-Nitropyrene exposure induces mitochondria dysfunction and impairs oocyte maturation in mice.

Oocyte quality is essential for a successful pregnancy. 1-Nitropyrene (1-NP) is a widely distributed pollutant in the environment and is well-known for its mutagenicity and carcinogenicity. However, whether 1-NP has toxic effects on mammalian oocyte quality remains unknown. In the present study, we focused on the effect of 1-NP on oocyte maturation using mouse oocytes as an in vitro model. Our study showed that 1-NP exposure disrupted the meiotic spindle assembly and caused chromosome misalignment, further impaired first polar body extrusion, and significantly decreased the fertilization capability in mouse oocytes. Further investigation showed that the mitochondrial membrane potential (MMP) and ATP levels were decreased, and the expression of genes encoding components of the mitochondrial respiratory chain was inhibited in 1-NP exposed oocytes. Meanwhile, 1-NP exposure increased the levels of reactive oxygen species (ROS), inhibited the expression of genes encoding antioxidant enzymes, and increased the frequency of early apoptotic oocytes. Overall, our data suggest that 1-NP exposure disrupts mitochondrial function and intracellular redox balance, ultimately impairing oocyte maturation. These findings reveal the adverse effect of 1-NP exposure on oocyte quality.

E3 ligase Hei10: a multifaceted structure-based signaling molecule with roles within and beyond meiosis.

Human enhancer of invasion-10 (Hei10) mediates meiotic recombination and also plays roles in cell proliferation. Here we explore Hei10's roles throughout the sexual cycle of the fungus Sordaria with respect to localization and effects of null, RING-binding, and putative cyclin-binding (RXL) domain mutations. Hei10 makes three successive types of foci. Early foci form along synaptonemal complex (SC) central regions. At some of these positions, depending on its RING and RXL domains, Hei10 mediates development and turnover of two sequential types of recombination complexes, each demarked by characteristic amplified Hei10 foci. Integration with ultrastructural data for recombination nodules further reveals that recombination complexes differentiate into three types, one of which corresponds to crossover recombination events during or prior to SC formation. Finally, Hei10 positively and negatively modulates SUMO localization along SCs by its RING and RXL domains, respectively. The presented findings suggest that Hei10 integrates signals from the SC, associated recombination complexes, and the cell cycle to mediate both the development and programmed turnover/evolution of recombination complexes via SUMOylation/ubiquitination. Analogous cell cycle-linked assembly/disassembly switching could underlie localization and roles for Hei10 in centrosome/spindle pole body dynamics and associated nuclear trafficking. We suggest that Hei10 is a unique type of structure-based signal transduction protein.

Crossover Interference, Crossover Maturation, and Human Aneuploidy.

A striking feature of human female sexual reproduction is the high level of gametes that exhibit an aberrant number of chromosomes (aneuploidy). A high baseline observed in women of prime reproductive age is followed by a dramatic increase in older women. Proper chromosome segregation requires one or more DNA crossovers (COs) between homologous maternal and paternal chromosomes, in combination with cohesion between sister chromatid arms. In human females, CO designations occur normally, according to the dictates of CO interference, giving early CO-fated intermediates. However, ≈25% of these intermediates fail to mature to final CO products. This effect explains the high baseline of aneuploidy and is predicted to synergize with age-dependent cohesion loss to explain the maternal age effect. Here, modern advances in the understanding of crossing over and CO interference are reviewed, the implications of human female CO maturation inefficiency are further discussed, and areas of interest for future studies are suggested.

SCRE serves as a unique synaptonemal complex fastener and is essential for progression of meiosis prophase I in mice.

Meiosis is a specialized cell division for producing haploid gametes from diploid germ cells. During meiosis, synaptonemal complex (SC) mediates the alignment of homologs and plays essential roles in homologous recombination and therefore in promoting accurate chromosome segregation. In this study, we have identified a novel protein SCRE (synaptonemal complex reinforcing element) as a key molecule in maintaining the integrity of SC during meiosis prophase I in mice. Deletion of Scre (synaptonemal complex reinforcing element) caused germ cell death in both male and female mice, resulting in infertility. Our mechanistic studies showed that the synapses and SCs in Scre knockout mice were unstable due to the lack of the SC reinforcing function of SCRE, which is sparsely localized as discrete foci along the central elements in normal synaptic homologous chromosomes. The lack of Scre leads to meiosis collapse at the late zygotene stage. We further showed that SCRE interacts with synaptonemal complex protein 1 (SYCP1) and synaptonemal complex central element 3 (SYCE3). We conclude that the function of SCRE is to reinforce the integrity of the central elements, thereby stabilizing the SC and ensuring meiotic cell cycle progression. Our study identified SCRE as a novel SC fastener protein that is distinct from other known SC proteins.

Topoisomerase II mediates meiotic crossover interference.

Spatial patterning is a ubiquitous feature of biological systems. Meiotic crossovers provide an interesting example, defined by the classic phenomenon of crossover interference. Here we identify a molecular pathway for interference by analysing crossover patterns in budding yeast. Topoisomerase II plays a central role, thus identifying a new function for this critical molecule. SUMOylation (of topoisomerase II and axis component Red1) and ubiquitin-mediated removal of SUMOylated proteins are also required. The findings support the hypothesis that crossover interference involves accumulation, relief and redistribution of mechanical stress along the protein/DNA meshwork of meiotic chromosome axes, with topoisomerase II required to adjust spatial relationships among DNA segments.

Meiotic prophase roles of Rec8 in crossover recombination and chromosome structure.

Rec8 is a prominent component of the meiotic prophase chromosome axis that mediates sister chromatid cohesion, homologous recombination and chromosome synapsis. Here, we explore the prophase roles of Rec8. (i) During the meiotic divisions, Rec8 phosphorylation mediates its separase-mediated cleavage. We show here that such cleavage plays no detectable role for chromosomal events of prophase. (ii) We have analyzed in detail three rec8 phospho-mutants, with 6, 24 or 29 alanine substitutions. A distinct 'separation of function' phenotype is revealed. In the mutants, axis formation and recombination initiation are normal, as is non-crossover recombination; in contrast, crossover (CO)-related events are defective. Moreover, the severities of these defects increase coordinately with the number of substitution mutations, consistent with the possibility that global phosphorylation of Rec8 is important for these effects. (iii) We have analyzed the roles of three kinases that phosphorylate Rec8 during prophase. Timed inhibition of Dbf4-dependent Cdc7 kinase confers defects concordant with rec8 phospho-mutant phenotypes. Inhibition of Hrr25 or Cdc5/polo-like kinase does not. Our results suggest that Rec8's prophase function, independently of cohesin cleavage, contributes to CO-specific events in conjunction with the maintenance of homolog bias at the leptotene/zygotene transition of meiotic prophase.

Crossover patterning by the beam-film model: analysis and implications.

Crossing-over is a central feature of meiosis. Meiotic crossover (CO) sites are spatially patterned along chromosomes. CO-designation at one position disfavors subsequent CO-designation(s) nearby, as described by the classical phenomenon of CO interference. If multiple designations occur, COs tend to be evenly spaced. We have previously proposed a mechanical model by which CO patterning could occur. The central feature of a mechanical mechanism is that communication along the chromosomes, as required for CO interference, can occur by redistribution of mechanical stress. Here we further explore the nature of the beam-film model, its ability to quantitatively explain CO patterns in detail in several organisms, and its implications for three important patterning-related phenomena: CO homeostasis, the fact that the level of zero-CO bivalents can be low (the "obligatory CO"), and the occurrence of non-interfering COs. Relationships to other models are discussed.

Interference-mediated synaptonemal complex formation with embedded crossover designation.

Biological systems exhibit complex patterns at length scales ranging from the molecular to the organismic. Along chromosomes, events often occur stochastically at different positions in different nuclei but nonetheless tend to be relatively evenly spaced. Examples include replication origin firings, formation of chromatin loops along chromosome axes and, during meiosis, localization of crossover recombination sites ("crossover interference"). We present evidence in the fungus Sordaria macrospora that crossover interference is part of a broader pattern that includes synaptonemal complex (SC) nucleation. This pattern comprises relatively evenly spaced SC nucleation sites, among which a subset are crossover sites that show a classical interference distribution. This pattern ensures that SC forms regularly along the entire length of the chromosome as required for the maintenance of homolog pairing while concomitantly having crossover interactions locally embedded within the SC structure as required for both DNA recombination and structural events of chiasma formation. This pattern can be explained by a threshold-based designation and spreading interference process. This model can be generalized to give diverse types of related and/or partially overlapping patterns, in two or more dimensions, for any type of object.