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A central feature of meiosis is the pairing of homologous maternal and paternal chromosomes ('homologues') along their lengths 1-3 . Recognition between homologues and their juxtaposition in space is mediated by axis-associated recombination complexes. Also, pairing must occur without entanglements among unrelated chromosomes. Here we examine homologue juxtaposition in real time by four-dimensional fluorescence imaging of tagged chromosomal loci at high spatio-temporal resolution in budding yeast. We discover that corresponding loci come together from a substantial distance (1.8 µm) and complete pairing in a very short time, about 6 min (thus, rapid homologue juxtaposition or RHJ). Homologue loci first move rapidly together (in 30 s, at speeds of roughly 60 nm s -1 ) into an intermediate stage corresponding to canonical 400 nm axis coalignment. After a short pause, crossover/non-crossover differentiation (crossover interference) mediates a second short, rapid transition that ultimately gives close pairing of axes at 100 nm by means of synaptonemal complex formation. Furthermore, RHJ (1) occurs after chromosomes acquire prophase chromosome organization, (2) is nearly synchronous over thirds of chromosome lengths, but (3) is asynchronous throughout the genome. Finally, cytoskeleton-mediated movement is important for the timing and distance of RHJ onset and for ensuring its normal progression. General implications for local and global aspects of pairing are discussed., (© 2024. The Author(s), under exclusive licence to Springer Nature Limited.)

Meiotic recombination is initiated by programmed double-strand breaks (DSBs). Studies in Saccharomyces cerevisiae have shown that, following rapid resection to generate 3' single-stranded DNA (ssDNA) tails, one DSB end engages a homolog partner chromatid and is extended by DNA synthesis, whereas the other end remains associated with its sister. Then, after regulated differentiation into crossover- and noncrossover-fated types, the second DSB end participates in the reaction by strand annealing with the extended first end, along both pathways. This second-end capture is dependent on Rad52, presumably via its known capacity to anneal two ssDNAs. Here, using physical analysis of DNA recombination, we demonstrate that this process is dependent on direct interaction of Rad52 with the ssDNA binding protein, replication protein A (RPA). Furthermore, the absence of this Rad52-RPA joint activity results in a cytologically-prominent RPA spike, which emerges from the homolog axes at sites of crossovers during the pachytene stage of the meiotic prophase. Our findings suggest that this spike represents the DSB end of a broken chromatid caused by either the displaced leading DSB end or the second DSB end, which has been unable to engage with the partner homolog-associated ssDNA. These and other results imply a close correspondence between Rad52-RPA roles in meiotic recombination and mitotic DSB repair., (© The Author(s) 2024. Published by Oxford University Press on behalf of Nucleic Acids Research.)

Meiosis is the specialized cellular program that underlies gamete formation for sexual reproduction. It is therefore not only interesting but also a fundamentally important subject for investigation. An especially attractive feature of this program is that many of the processes of special interest involve organized chromosomes, thus providing the possibility to see chromosomes "in action". Analysis of meiosis has also proven to be useful in discovering and understanding processes that are universal to all chromosomal programs. Here we provide an overview of the different historical moments when the gap between observation and understanding of mechanisms and/or roles for the new discovered molecules was bridged. This review reflects also the synergy of thinking and discussion among our three laboratories during the past several decades., (© 2024. The Author(s).)

The raison d'être of meiosis is shuffling of genetic information via Mendelian segregation and, within individual chromosomes, by DNA crossing-over. These outcomes are enabled by a complex cellular program in which interactions between homologous chromosomes play a central role. We first provide a background regarding the basic principles of this program. We then summarize the current understanding of the DNA events of recombination and of three processes that involve whole chromosomes: homolog pairing, crossover interference, and chiasma maturation. All of these processes are implemented by direct physical interaction of recombination complexes with underlying chromosome structures. Finally, we present convergent lines of evidence that the meiotic program may have evolved by coupling of this interaction to late-stage mitotic chromosome morphogenesis.

We have explored the meiotic roles of cohesin modulators Pds5 and Rad61/Wapl, in relation to one another, and to meiotic kleisin Rec8, for homolog pairing, all physically definable steps of recombination, prophase axis length and S-phase progression, in budding yeast. We show that Pds5 promotes early steps of recombination and thus homolog pairing, and also modulates axis length, with both effects independent of a sister chromatid. [Pds5+Rec8] promotes double-strand break formation, maintains homolog bias for crossover formation and promotes S-phase progression. Oppositely, the unique role of Rad61/Wapl is to promote non-crossover recombination by releasing [Pds5+Rec8]. For this effect, Rad61/Wapl probably acts to maintain homolog bias by preventing channeling into sister interactions. Mysteriously, each analyzed molecule has one role that involves neither of the other two. Overall, the presented findings suggest that Pds5's role in maintenance of sister chromatid cohesion during the mitotic prophase-analogous stage of G2/M is repurposed during meiosis prophase to promote interactions between homologs., (© The Author(s) 2019. Published by Oxford University Press on behalf of Nucleic Acids Research.)

Meiosis is the cellular program by which a diploid cell gives rise to haploid gametes for sexual reproduction. Meiotic progression depends on tight physical and functional coupling of recombination steps at the DNA level with specific organizational features of meiotic-prophase chromosomes. The present study reveals that every step of this coupling is mediated by a single molecule: Asy2/Mer2. We show that Mer2, identified so far only in budding and fission yeasts, is in fact evolutionarily conserved from fungi (Mer2/Rec15/Asy2/Bad42) to plants (PRD3/PAIR1) and mammals (IHO1). In yeasts, Mer2 mediates assembly of recombination-initiation complexes and double-strand breaks (DSBs). This role is conserved in the fungus Sordaria However, functional analysis of 13 mer2 mutants and successive localization of Mer2 to axis, synaptonemal complex (SC), and chromatin revealed, in addition, three further important functions. First, after DSB formation, Mer2 is required for pairing by mediating homolog spatial juxtaposition, with implications for crossover (CO) patterning/interference. Second, Mer2 participates in the transfer/maintenance and release of recombination complexes to/from the SC central region. Third, after completion of recombination, potentially dependent on SUMOylation, Mer2 mediates global chromosome compaction and post-recombination chiasma development. Thus, beyond its role as a recombinosome-axis/SC linker molecule, Mer2 has important functions in relation to basic chromosome structure., (© 2017 Tessé et al.; Published by Cold Spring Harbor Laboratory Press.)

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., (Copyright © 2017 Elsevier Inc. All rights reserved.)

Many morphological features, in both physical and biological systems, exhibit spatial patterns that are specifically characterized by a tendency to occur with even spacing (in one, two, or three dimensions). The positions of crossover (CO) recombination events along meiotic chromosomes provide an interesting biological example of such an effect. In general, mechanisms that explain such patterns may (a) be mechanically based, (b) occur by a reaction-diffusion mechanism in which macroscopic mechanical effects are irrelevant, or (c) involve a combination of both types of effects. We have proposed that meiotic CO patterns arise by a mechanical mechanism, have developed mathematical expressions for such a process based on a particular physical system with analogous properties (the so-called beam-film model), and have shown that the beam-film model can very accurately explain experimental CO patterns as a function of the values of specific defined parameters. Importantly, the mathematical expressions of the beam-film model can apply quite generally to any mechanism, whether it involves mechanical components or not, as long as its logic and component features correspond to those of the beam-film system. Furthermore, via its various parameters, the beam-film model discretizes the patterning process into specific components. Thus, the model can be used to explore the theoretically predicted effects of various types of changes in the patterning process. Such predictions can expand detailed understanding of the bases for various biological effects. We present here a new MATLAB program that implements the mathematical expressions of the beam-film model with increased robustness and accessibility as compared to programs presented previously. As in previous versions, the presented program permits both (1) simulation of predicted CO positions along chromosomes of a test population and (2) easy analysis of CO positions, both for experimental data sets and for data sets resulting from simulations. The goal of the current presentation is to make these approaches more readily accessible to a wider audience of researchers. Also, the program is easily modified, and we encourage interested users to make changes to suit their specific needs. A link to the program is available on the Kleckner laboratory website: http://projects.iq.harvard.edu/kleckner_lab .

Recombination is a prominent feature of meiosis in which it plays an important role in increasing genetic diversity during inheritance. Additionally, in most organisms, recombination also plays mechanical roles in chromosomal processes, most notably to mediate pairing of homologous chromosomes during prophase and, ultimately, to ensure regular segregation of homologous chromosomes when they separate at the first meiotic division. Recombinational interactions are also subject to important spatial patterning at both early and late stages. Recombination-mediated processes occur in physical and functional linkage with meiotic axial chromosome structure, with interplay in both directions, before, during, and after formation and dissolution of the synaptonemal complex (SC), a highly conserved meiosis-specific structure that links homolog axes along their lengths. These diverse processes also are integrated with recombination-independent interactions between homologous chromosomes, nonhomology-based chromosome couplings/clusterings, and diverse types of chromosome movement. This review provides an overview of these diverse processes and their interrelationships., (Copyright © 2015 Cold Spring Harbor Laboratory Press; all rights reserved.)

During meiosis, crossover recombination is tightly regulated. A spatial patterning phenomenon known as interference ensures that crossovers are well-spaced along the chromosomes. Additionally, every pair of homologs acquires at least one crossover. A third feature, crossover homeostasis, buffers the system such that the number of crossovers remains steady despite decreases or increases in the number of earlier recombinational interactions. Here we summarize recent work from our laboratory supporting the idea that all 3 of these aspects are intrinsic consequences of a single basic process and suggesting that the underlying logic of this process corresponds to that embodied in a particular (beam-film) model.

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.

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., (© 2014 De Muyt et al.; Published by Cold Spring Harbor Laboratory Press.)

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.

Recombinational repair of spontaneous double-strand breaks (DSBs) exhibits sister bias. DSB-initiated meiotic recombination exhibits homolog bias. Physical analysis in yeast reveals that, in both cases, the recombination reaction intrinsically gives homolog bias. From this baseline default, cohesin intervenes to confer sister bias, likely independent of cohesion. In meiosis, cohesin's sister-biasing effect is counteracted by RecA homolog Rad51 and its mediators, plus meiotic RecA homolog Dmc1, which thereby restore intrinsic homolog bias. Meiotic axis complex Red1/Mek1/Hop1 participates by cleanly switching recombination from mitotic to meiotic mode, concomitantly activating Dmc1. We propose that a Rad51/DNA filament at one DSB end captures the intact sister, creating an anchor pad. This filament extends across the DSB site on the intact partner, precluding intersister strand exchange, thus forcing use of the homolog. Cohesin and Dmc1 interactively modulate this extension, with program-appropriate effects. In accord with this model, Rad51-mediated recombination in vivo requires the presence of a sister., (Copyright © 2013 Elsevier Inc. All rights reserved.)

Meiotic recombination initiates via programmed double-strand breaks (DSBs). We investigate whether, at a given initiation site, DSBs occur independently among the four available chromatids. For a single DSB "hot spot", the proportions of nuclei exhibiting zero, one, or two (or more) observable events were defined by tetrad analysis and compared with those predicted by different DSB distribution scenarios. Wild-type patterns are incompatible with independent distribution of DSBs among the four chromatids. In most or all nuclei, DSBs occur one-per-pair of chromatids, presumptively sisters. In many nuclei, only one DSB occurs per four chromatids, confirming the existence of trans inhibition where a DSB on one chromosome interactively inhibits DSB formation on the partner chromosome. Several mutants exhibit only a one-per-pair constraint, a phenotype we propose to imply loss of trans inhibition. Signal transduction kinases Mec1 (ATR) and Tel1 (ATM) exhibit this phenotype and thus could be mediators of this effect. Spreading trans inhibition can explain even spacing of total recombinational interactions and implies that establishment of interhomolog interactions and DSB formation are homeostatic processes. The two types of constraints on DSB formation provide two different safeguards against recombination failure during meiosis.

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., (Copyright © 2010 Elsevier Inc. All rights reserved.)

Meiotic chromosome pairing involves not only recognition of homology but also juxtaposition of entire chromosomes in a topologically regular way. Analysis of filamentous fungus Sordaria macrospora reveals that recombination proteins Mer3, Msh4, and Mlh1 play direct roles in all of these aspects, in advance of their known roles in recombination. Absence of Mer3 helicase results in interwoven chromosomes, thereby revealing the existence of features that specifically ensure "entanglement avoidance." Entanglements that remain at zygotene, i.e., "interlockings," require Mlh1 for resolution, likely to eliminate constraining recombinational connections. Patterns of Mer3 and Msh4 foci along aligned chromosomes show that the double-strand breaks mediating homologous alignment have spatially separated ends, one localized to each partner axis, and that pairing involves interference among developing interhomolog interactions. We propose that Mer3, Msh4, and Mlh1 execute all of these roles during pairing by modulating the state of nascent double-strand break/partner DNA contacts within axis-associated recombination complexes., (Copyright 2010 Elsevier Inc. All rights reserved.)

Dramatic chromosome motion is a characteristic of mid-prophase of meiosis that is observed across broadly divergent eukaryotic phyla. Although the specific mechanisms underlying chromosome motions vary among organisms studied to date, the outcome is similar in all cases: vigorous back-and-forth movement (as fast as approximately 1mum/sec for budding yeast), led by chromosome ends (or near-end regions), and directed by cytoskeletal components via direct association through the nuclear envelope. The exact role(s) of these movements remains unknown, although an idea gaining currency is that movement serves as a stringency factor, eliminating unwanted inter-chromosomal associations or entanglements that have arisen as part of the homolog pairing process and, potentially, unwanted associations of chromatin with the nuclear envelope. Turbulent chromosome movements observed during bipolar orientation of chromosomes for segregation could also serve similar roles during mitosis. Recent advances shed light on the contribution of protein complexes involved in the meiotic movements in chromosome dynamics during the mitotic program.

Chromosome movements are a general feature of mid-prophase of meiosis. In budding yeast, meiotic chromosomes exhibit dynamic movements, led by nuclear envelope (NE)-associated telomeres, throughout the zygotene and pachytene stages. Zygotene motion underlies the global tendency for colocalization of NE-associated chromosome ends in a "bouquet." In this study, we identify Csm4 as a new molecular participant in these processes and show that, unlike the two previously identified components, Ndj1 and Mps3, Csm4 is not required for meiosis-specific telomere/NE association. Instead, it acts to couple telomere/NE ensembles to a force generation mechanism. Mutants lacking Csm4 and/or Ndj1 display the following closely related phenotypes: (i) elevated crossover (CO) frequencies and decreased CO interference without abrogation of normal pathways; (ii) delayed progression of recombination, and recombination-coupled chromosome morphogenesis, with resulting delays in the MI division; and (iii) nondisjunction of homologs at the MI division for some reason other than absence of (the obligatory) CO(s). The recombination effects are discussed in the context of a model where the underlying defect is chromosome movement, the absence of which results in persistence of inappropriate chromosome relationships that, in turn, results in the observed mutant phenotypes., Competing Interests: The authors have declared that no competing interests exist.

During meiosis, DNA events of recombination occur in direct physical association with underlying chromosome axes. Meiotic cohesin Rec8 and cohesin-associated Spo76/Pds5 are prominent axis components. Two observations indicate that recombination complexes can direct the local destabilization of underlying chromosome axes. First, in the absence of Rec8, Spo76/Pds5 is lost locally at sites of late-persisting Msh4 foci, with a concomitant tendency for loosening of intersister and interhomolog connectedness at the affected sites. This loss is dependent on initiation of recombination. Second, in wild-type prophase, local separation of sister axes is seen at sites of synaptonemal complex-associated recombination nodules. Additional findings reveal that Rec8 localizes to both axis and bulk chromatin and is required for chromatin compactness. Further, Rec8 is essential for maintenance of sister cohesion, along arms and centromeres, during the pachytene-to-diplotene transition, revealing an intrinsic tendency for destabilization of sister cohesion during this period. This finding shows how the loss of sister connectedness, in arm and/or centric regions, could lead to the segregation defects that are seen in the human "maternal age effect" and how Rec8 could be a target of that effect. Finally, Rec8 plays related, but synergistic roles with Spo76/Pds5, indicating auxiliary roles for meiotic and mitotic cohesion-associated components.

We show that, during budding yeast meiosis, axis ensemble Hop1/Red1 and synaptonemal complex (SC) component Zip1 tend to occur in alternating strongly staining domains. The widely conserved AAA+-ATPase Pch2 mediates this pattern, likely by means of direct intervention along axes. Pch2 also coordinately promotes timely progression of cross-over (CO) and noncross-over (NCO) recombination. Oppositely, in a checkpoint-triggering aberrant situation (zip1Delta), Pch2 mediates robust arrest of stalled recombination complexes, likely via nucleolar localization. We suggest that, during WT meiosis, Pch2 promotes progression of SC-associated CO and NCO recombination complexes at a regulated early-midpachytene transition that is rate-limiting for later events; in contrast, during defective meiosis, Pch2 ensures that aberrant recombination complexes fail to progress so that intermediates can be harmlessly repaired during eventual return to growth. Positive vs. negative roles of Pch2 in the two situations are analogous to positive vs. negative roles of Mec1/ATR, suggesting that Pch2 might mediate Mec1/ATR activity. We further propose that regulatory surveillance of normal and abnormal interchromosomal interactions in mitotic and meiotic cells may involve "structure-dependent interchromosomal interaction" (SDIX) checkpoints.

Meiotic recombination-related DNA synthesis (MRDS) was analyzed in Saccharomyces cerevisiae by specifically timed incorporation of thymidine analogs into chromosomes. Lengths and positions of incorporation tracts were determined relative to a known recombination hot spot along DNA, as was the timing and localization of incorporation relative to forming and formed synaptonemal complex in spread chromosomes. Distinct patterns could be specifically associated with the majority cross-over and non-cross-over recombination processes. The results obtained provide direct evidence for key aspects of current consensus recombination models, provide information regarding temporal and spatial relationships between non-cross-over formation and the synaptonemal complex, and raise the possibility that removal of RecA homolog Rad51 plays a key role in regulating onset of MRDS. Finally, classical observations on MRDS in Drosophila, mouse, and lily are readily mapped onto the findings presented here, providing further evidence for a broadly conserved meiotic recombination process.

Zip2 and Zip3 are meiosis-specific proteins that, in collaboration with several partners, act at the sites of crossover-designated, axis-associated recombinational interactions to mediate crossover/chiasma formation. Here, Spo22 (also called Zip4) is identified as a probable functional collaborator of Zip2/3. The molecular roles of Zip2, Zip3, and Spo22/Zip4 are unknown. All three proteins are part of a small evolutionary cohort comprising similar homologs in four related yeasts. Zip3 is shown to contain a RING finger whose structural features most closely match those of known ubiquitin E3s. Further, Zip3 exhibits major domainal homologies to Rad18, a known DNA-binding ubiquitin E3. Also described is an approach to the identification and mapping of repeated protein sequence motifs, Alignment Based Repeat Annotation (ABRA), that we have developed. When ABRA is applied to Zip2 and Spo22/Zip4, they emerge as a 14-blade WD40-like repeat protein and a 22-unit tetratricopeptide repeat protein, respectively. WD40 repeats of Cdc20, Cdh1, and Cdc16 and tetratricopeptide repeats of Cdc16, Cdc23, and Cdc27, all components of the anaphase-promoting complex, are also analyzed. These and other findings suggest that Zip2, Zip3, and Zip4 act together to mediate a process that involves Zip3-mediated ubiquitin labeling, potentially as a unique type of ubiquitin-conjugating complex.

Yeast mutants lacking meiotic proteins Zip1, Zip2, Zip3, Mer3, and/or Msh5 (ZMMs) were analyzed for recombination, synaptonemal complex (SC), and meiotic progression. At 33 degrees C, recombination-initiating double-strand breaks (DSBs) and noncrossover products (NCRs) form normally while formation of single-end invasion strand exchange intermediates (SEIs), double Holliday junctions, crossover products (CRs), and SC are coordinately defective. Thus, during wild-type meiosis, recombinational interactions are differentiated into CR and NCR types very early, prior to onset of stable strand exchange and independent of SC. By implication, crossover interference does not require SC formation. We suggest that SC formation may require interference. Subsequently, CR-designated DSBs undergo a tightly coupled, ZMM-promoted transition that yields SEI-containing recombination complexes embedded in patches of SC. zmm mutant phenotypes differ strikingly at 33 degrees C and 23 degrees C, implicating higher temperature as a positive effector of recombination and identifying a checkpoint that monitors local CR-specific events, not SC formation, at late leptotene.

Chromosomal processes related to formation and function of meiotic chiasmata have been analyzed in Sordaria macrospora. Double-strand breaks (DSBs), programmed or gamma-rays-induced, are found to promote four major events beyond recombination and accompanying synaptonemal complex formation: (1) juxtaposition of homologs from long-distance interactions to close presynaptic coalignment at midleptotene; (2) structural destabilization of chromosomes at leptotene/zygotene, including sister axis separation and fracturing, as revealed in a mutant altered in the conserved, axis-associated cohesin-related protein Spo76/Pds5p; (3) exit from the bouquet stage, with accompanying global chromosome movements, at zygotene/pachytene (bouquet stage exit is further found to be a cell-wide regulatory transition and DSB transesterase Spo11p is suggested to have a new noncatalytic role in this transition); (4) normal occurrence of both meiotic divisions, including normal sister separation. Functional interactions between DSBs and the spo76-1 mutation suggest that Spo76/Pds5p opposes local destabilization of axes at developing chiasma sites and raise the possibility of a regulatory mechanism that directly monitors the presence of chiasmata at metaphase I. Local chromosome remodeling at DSB sites appears to trigger an entire cascade of chromosome movements, morphogenetic changes, and regulatory effects that are superimposed upon a foundation of DSB-independent processes.

Analysis of meiotic recombination by functional genomic approaches reveals prominent spatial and functional interactions among diverse organizational determinants. Recombination occurs between chromatin loop sequences; however, these sequences are spatially tethered to underlying chromosome axes via their recombinosomes. Meiotic chromosomal protein, Red1, localizes to chromosome axes; however, Red1 loading is modulated by R/G-bands isochores and thus by bulk chromatin state. Recombination is also modulated by isochore determinants: R-bands differentially favor double-strand break (DSB) formation but disfavor subsequent loading of meiotic RecA homolog, Dmc1. Red1 promotes DSB formation in both R- and G-bands and then promotes Dmc1 loading, specifically counteracting disfavoring R-band effects. These complexities are discussed in the context of chiasma formation as a series of coordinated local changes at the DNA and chromosome-axis levels.

We describe an approach to detect the frequency of interaction between any two genomic loci. Generation of a matrix of interaction frequencies between sites on the same or different chromosomes reveals their relative spatial disposition and provides information about the physical properties of the chromatin fiber. This methodology can be applied to the spatial organization of entire genomes in organisms from bacteria to human. Using the yeast Saccharomyces cerevisiae, we could confirm known qualitative features of chromosome organization within the nucleus and dynamic changes in that organization during meiosis. We also analyzed yeast chromosome III at the G1 stage of the cell cycle. We found that chromatin is highly flexible throughout. Furthermore, functionally distinct AT- and GC-rich domains were found to exhibit different conformations, and a population-average 3D model of chromosome III could be determined. Chromosome III emerges as a contorted ring.

A central feature of meiosis is pairing of homologous chromosomes, which occurs in two stages: coalignment of axes followed by installation of the synaptonemal complex (SC). Concomitantly, recombination complexes reposition from on-axis association to the SC central region. We show here that, in the fungus Sordaria macrospora, this critical transition is mediated by robust interaxis bridges that contain an axis component (Spo76/Pds5), DNA, plus colocalizing Mer3/Msh4 recombination proteins and the Zip2-Zip4 mediator complex. Mer3-Msh4-Zip2-Zip4 colocalizing foci are first released from their tight axis association, dependent on the SC transverse-filament protein Sme4/Zip1, before moving to bridges and thus to a between-axis position. Ensuing shortening of bridges and accompanying juxtaposition of axes to 100 nm enables installation of SC central elements at sites of between-axisMer3-Msh4-Zip2-Zip4 complexes. We show also that the Zip2-Zip4 complex has an intrinsic affinity for chromosome axes at early leptotene, where it localizes independently of recombination, but is dependent on Mer3. Then, later, Zip2-Zip4 has an intrinsic affinity for the SC central element, where it ultimately localizes to sites of crossover complexes at the end of pachytene. These and other findings suggest that the fundamental role of Zip2-Zip4 is to mediate the recombination/structure interface at all post-double-strand break stages. We propose that Zip2-Zip4 directly mediates a molecular handoff of Mer3-Msh4 complexes, from association with axis components to associationwith SC central components, at the bridge stage, and then directly mediates central region installation during SC nucleation. [ABSTRACT FROM AUTHOR]

The article reports that geneticist Nancy Kleckner has received the Thomas Hunt Morgan Medal by the Genetics Society of America for her work in the field of genetics. It informs that Kleckner has made achievements in several different research areas, including bacterial transposition, chromosome organization, and meiosis. It also mentions that she has combined traditional genetic approaches with molecular biology, microscopy and modeling.

Meiosis presents many important mysteries that await elucidation. Here we discuss two such aspects. First, we consider how the current meiotic program might have evolved. We emphasize the central feature of this program: how homologous chromosomes find one another (“pair”) so as to create the connections required for their regular segregation at Meiosis I. Points of emphasis include the facts that: (i) the classical “bouquet stage” is not required for initial homolog contacts in the current evolved meiotic program; and (ii) diverse observations point to commonality between molecules that mediate meiotic inter-homolog interactions and molecules that are integral to centromeres and/or to microtubule organizing centers (a.k.a. spindle pole bodies or centrosomes). Second, we provide an overview of the classical phenomenon of crossover (CO) interference in an effort to bridge the gap between description on the one hand versus logic and mechanism on the other. [ABSTRACT FROM AUTHOR]

Repair of DNA double-strand breaks (DSBs) by homologous recombination is crucial for cell proliferation and tumour suppression. However, despite its importance, the molecular intermediates of mitotic DSB repair remain undefined. The double Holliday junction (DHJ), presupposed to be the central intermediate for more than 25 years, has only been identified during meiotic recombination. Moreover, evidence has accumulated for alternative, DHJ-independent mechanisms, raising the possibility that DHJs are not formed during DSB repair in mitotically cycling cells. Here we identify intermediates of DSB repair by using a budding-yeast assay system designed to mimic physiological DSB repair. This system uses diploid cells and provides the possibility for allelic recombination either between sister chromatids or between homologues, as well as direct comparison with meiotic recombination at the same locus. In mitotically cycling cells, we detect inter-homologue joint molecule (JM) intermediates whose strand composition and size are identical to those of the canonical DHJ structures observed in meiosis. However, in contrast to meiosis, JMs between sister chromatids form in preference to those between homologues. Moreover, JMs seem to represent a minor pathway of DSB repair in mitotic cells, being detected at about tenfold lower levels (per DSB) than during meiotic recombination. Thus, although DHJs are identified as intermediates of DSB-promoted recombination in both mitotic and meiotic cells, their formation is distinctly regulated according to the specific dictates of the two cellular programs. [ABSTRACT FROM AUTHOR]

We show that, during budding yeast meiosis, axis ensemble Hopi/Redi and synaptonemal complex (SC) component Zipi tend to occur in alternating strongly staining domains. The widely conserved AAA+-ATPase Pch2 mediates this pattern, likely by means of direct intervention along axes. Pch2 also coordinately promotes timely progression of cross-over (CO) and noncross-over (NCO) recombination. Oppositely, in a checkpoint-triggering aberrant situation (zip1Ɗ), Pch2 mediates robust arrest of stalled recombination complexes, likely via nucleolar localization. We suggest that, during WT meiosis, Pch2 promotes progression of SC-associated CO and NCO recombination complexes at a regulated early-midpachytene transition that is rate-limiting for later events; in contrast, during defective meiosis, Pch2 ensures that aberrant recombination complexes fail to progress so that intermediates can be harmlessly repaired during eventual return to growth. Positive vs. negative roles of Pch2 in the two situations are analogous to positive vs. negative roles of Mecl/ATR, suggesting that Pch2 might mediate Mecl/ ATR activity. We further propose that regulatory surveillance of normal and abnormal interchromosomal interactions in mitotic and meiotic cells may involve "structure-dependent interchromosomal interaction" (SDIX) checkpoints. [ABSTRACT FROM AUTHOR]

Meiotic recombination proceeds in biochemical complexes that are physically associated with underlying chromosome structural axes. In this study, we discuss the organizational basis for these axes, the timing and nature of recombinosome/axis organization with respect to the prophase program of DNA and to structural changes, and the possible significance of axis organization. Furthermore, we discuss implications and extensions of our recently proposed mechanical model for chiasma formation. Finally, we give a broader consideration to past and present models for the role of the synaptonemal complex. [ABSTRACT FROM AUTHOR]

Examines the occurrence of single-end invasion (SEI) during the transition from double-strand breaks (DSB) to double-Holliday junction. Modulation of meiotic DNA replication by interchromosomal interaction proteins; Structural asymmetry of SEI; Establishment of interhomolog bias prior to DSB formation.

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, designation of crossover recombination sites ("crossover interference"). We present evidence, in the fungus Sordaria macrospora, that crossover interference is part of a broader patterning program that includes synaptonemal complex (SC) nucleation. This program yields relatively evenly-spaced SC nucleation sites; among these, a subset is also crossover sites that show a classical interference distribution. This pattern ensures that SC forms regularly along the entire lengths of the chromosomes as required for homolog pairing maintenance and interlock sensing while concomitantly embedding crossover interactions 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 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., Other Research Unit