Mouse pachytene checkpoint 2 (trip13) is required for completing meiotic recombination but not synapsis

In mammalian meiosis, homologous chromosome synapsis is coupled with recombination. As in most eukaryotes, mammalian meiocytes have checkpoints that monitor the fidelity of these processes. We report that the mouse ortholog (Trip13) of pachytene checkpoint 2 (PCH2), an essential component of the synapsis checkpoint in Saccharomyces cerevisiae and Caenorhabditis elegans, is required for completion of meiosis in both sexes. TRIP13-deficient mice exhibit spermatocyte death in pachynema and loss of oocytes around birth. The chromosomes of mutant spermatocytes synapse fully, yet retain several markers of recombination intermediates, including RAD51, BLM, and RPA. These chromosomes also exhibited the chiasmata markers MLH1 and MLH3, and okadaic acid treatment of mutant spermatocytes caused progression to metaphase I with bivalent chromosomes. Double mutant analysis demonstrated that the recombination and synapsis genes Spo11, Mei1, Rec8, and Dmc1 are all epistatic to Trip13, suggesting that TRIP13 does not have meiotic checkpoint function in mice. Our data indicate that TRIP13 is required after strand invasion for completing a subset of recombination events, but possibly not those destined to be crossovers. To our knowledge, this is the first model to separate recombination defects from asynapsis in mammalian meiosis, and provides the first evidence that unrepaired DNA damage alone can trigger the pachytene checkpoint response in mice.     

Presence of an extra chromosome alters meiotic double-stranded break repair dynamics and MLH1 foci distribution in human oocytes

Studies performed on human trisomic 21 oocytes have revealed that during meiosis, the three homologues 21 synapse and, in some cases, achieve what looks like a trivalent. This implies that meiotic recombination takes place among the three homologous chromosomes 21, and to some extent, crossovers form between them. To see how meiotic recombination is in the presence of an extra chromosome 21, we analyzed the distribution of three recombination markers (γH2AX, RPA, and MLH1) on trisomic 21 oocytes at pachynema and, in particular, on chromosomes 21. Results clearly show how the presence of an extra chromosome 21 alters meiotic recombination progression, leading to the presence of a higher number of early recombination markers at pachynema. Moreover, the distribution on these chromosomes 21 of some of these markers is different in aneuploid oocytes. Finally, there is a substantial increase in the number of MLH1 foci, a marker of most crossovers in mammals, which is related to the number of synapsed chromosomes in pachynema. Thus, bivalents 21 had fewer MLH1 foci than partial or total trivalents, suggesting a close relationship between synapsis and crossover designation. All of the data presented suggest that the presence of an extra chromosome alters meiotic recombination globally in aneuploid human oocytes.     

Patterns of meiotic recombination in human fetal oocytes

Abnormal patterns of meiotic recombination (i.e., crossing-over) are believed to increase the risk of chromosome nondisjunction in human oocytes. To date, information on recombination has been obtained using indirect, genetic methods. Here we use an immunocytological approach, based on detection of foci of a DNA mismatch-repair protein, MLH1, on synaptonemal complexes at prophase I of meiosis, to provide the first direct estimate of the frequency of meiotic recombination in human oocytes. At pachytene, the stage of maximum homologous chromosome pairing, we found a mean of 70.3 foci (i.e., crossovers) per oocyte, with considerable intercell variability (range 48-102 foci). This mean equates to a genetic-map length of 3,515 cM. The numbers and positions of foci were determined for chromosomes 21, 18, 13, and X. These chromosomes yielded means of 1.23 foci (61.5 cM), 2.36 foci (118 cM), 2.5 foci (125 cM), and 3.22 foci (161 cM), respectively. The foci were almost invariably located interstitially and were only occasionally located close to chromosome ends. These data confirm the large difference, in recombination frequency, between human oocytes and spermatocytes and demonstrate a clear intersex variation in distribution of crossovers. In a few cells, chromosomes 21 and 18 did not have any foci (i.e., were presumptively noncrossover); however, configurations that lacked foci were not observed for chromosomes 13 and X. For the latter two chromosome pairs, the only instances of absence of foci were observed in abnormal cells that showed chromosome-pairing errors affecting these chromosomes. We speculate that these abnormal fetal oocytes may be the source of the nonrecombinant chromosomes 13 and X suggested, by genetic studies, to be associated with maternally derived chromosome nondisjunction.     

Male mouse recombination maps for each autosome identified by chromosome painting

Linkage maps constructed from genetic analysis of gene order and crossover frequency provide few clues to the basis of genomewide distribution of meiotic recombination, such as chromosome structure, that influences meiotic recombination. To bridge this gap, we have generated the first cytological recombination map that identifies individual autosomes in the male mouse. We prepared meiotic chromosome (synaptonemal complex [SC]) spreads from 110 mouse spermatocytes, identified each autosome by multicolor fluorescence in situ hybridization of chromosome-specific DNA libraries, and mapped >2,000 sites of recombination along individual autosomes, using immunolocalization of MLH1, a mismatch repair protein that marks crossover sites. We show that SC length is strongly correlated with crossover frequency and distribution. Although the length of most SCs corresponds to that predicted from their mitotic chromosome length rank, several SCs are longer or shorter than expected, with corresponding increases and decreases in MLH1 frequency. Although all bivalents share certain general recombination features, such as few crossovers near the centromeres and a high rate of distal recombination, individual bivalents have unique patterns of crossover distribution along their length. In addition to SC length, other, as-yet-unidentified, factors influence crossover distribution leading to hot regions on individual chromosomes, with recombination frequencies as much as six times higher than average, as well as cold spots with no recombination. By reprobing the SC spreads with genetically mapped BACs, we demonstrate a robust strategy for integrating genetic linkage and physical contig maps with mitotic and meiotic chromosome structure.     

Chromosome-wide control of meiotic crossing over in C. elegans

A central event in sexual reproduction is the reduction in chromosome number that occurs at the meiosis I division. Most eukaryotes rely on crossing over between homologs, and the resulting chiasmata, to direct meiosis I chromosome segregation, yet make very few crossovers per chromosome pair. This indicates that meiotic recombination must be tightly regulated to ensure that each chromosome pair enjoys the crossover necessary to ensure correct segregation. Here, we investigate control of meiotic crossing over in Caenorhabditis elegans, which averages only one crossover per chromosome pair per meiosis, by constructing genetic maps of end-to-end fusions of whole chromosomes. Fusion of chromosomes removes the requirement for a crossover in each component chromosome segment and thereby reveals a propensity to restrict the number of crossovers such that pairs of fusion chromosomes composed of two or even three whole chromosomes enjoy but a single crossover in the majority of meioses. This regulation can operate over physical distances encompassing half the genome. The meiotic behavior of heterozygous fusion chromosomes further suggests that continuous meiotic chromosome axes, or structures that depend on properly assembled axes, may be important for crossover regulation.     

Meiotic recombination protein Rec12: functional conservation, crossover homeostasis and early crossover/non-crossover decision

In fission yeast and other eukaryotes, Rec12 (Spo11) is thought to catalyze the formation of dsDNA breaks (DSBs) that initiate homologous recombination in meiosis. Rec12 is orthologous to the catalytic subunit of topoisomerase VI (Top6A). Guided by the crystal structure of Top6A, we engineered the rec12 locus to encode Rec12 proteins each with a single amino acid substitution in a conserved residue. Of 21 substitutions, 10 significantly reduced or abolished meiotic DSBs, gene conversion, crossover recombination and the faithful segregation of chromosomes. Critical residues map within the metal ion-binding pocket toprim (E179A, D229A, D231A), catalytic region 5Y-CAP (R94A, D95A, Y98F) and the DNA-binding interface (K201A, G202E, R209A, K242A). A subset of substitutions reduced DSBs but maintained crossovers, demonstrating crossover homeostasis. Furthermore, a strong separation of function mutation (R304A) suggests that the crossover/non-crossover decision is established early by a protein-protein interaction surface of Rec12. Fission yeast has multiple crossovers per bivalent, and chromosome segregation was robust above a threshold of about one crossover per bivalent, below which non-disjunction occurred. These results support structural and functional conservation among Rec12/Spo11/Top6A family members for the catalysis of DSBs, and they reveal how Rec12 regulates other features of meiotic chromosome dynamics.     

Recombination map of the common shrew, Sorex araneus (Eulipotyphla, Mammalia)

The Eurasian common shrew (Sorex araneus L.) is characterized by spectacular chromosomal variation, both autosomal variation of the Robertsonian type and an XX/XY(1)Y(2) system of sex determination. It is an important mammalian model of chromosomal and genome evolution as it is one of the few species with a complete genome sequence. Here we generate a high-precision cytological recombination map for the species, the third such map produced in mammals, following those for humans and house mice. We prepared synaptonemal complex (SC) spreads of meiotic chromosomes from 638 spermatocytes of 22 males of nine different Robertsonian karyotypes, identifying each autosome arm by differential DAPI staining. Altogether we mapped 13,983 recombination sites along 7095 individual autosomes, using immunolocalization of MLH1, a mismatch repair protein marking recombination sites. We estimated the total recombination length of the shrew genome as 1145 cM. The majority of bivalents showed a high recombination frequency near the telomeres and a low frequency near the centromeres. The distances between MLH1 foci were consistent with crossover interference both within chromosome arms and across the centromere in metacentric bivalents. The pattern of recombination along a chromosome arm was a function of its length, interference, and centromere and telomere effects. The specific DNA sequence must also be important because chromosome arms of the same length differed substantially in their recombination pattern. These features of recombination show great similarity with humans and mice and suggest generality among mammals. However, contrary to a widespread perception, the metacentric bivalent tu usually lacked an MLH1 focus on one of its chromosome arms, arguing against a minimum requirement of one chiasma per chromosome arm for correct segregation. With regard to autosomal chromosomal variation, the chromosomes showing Robertsonian polymorphism display MLH1 foci that become increasingly distal when comparing acrocentric homozygotes, heterozygotes, and metacentric homozygotes. Within the sex trivalent XY(1)Y(2), the autosomal part of the complex behaves similarly to other autosomes.     

Sex-dependent frequency and type of autosomal univalency at the first meiotic metaphase in mouse germ cells

Univalents at the first meiotic metaphase in mouse spermatocytes occur mainly in the XY pair, making it difficult to compare the amounts of univalency in males and females. In this study, the amounts of autosomal univalency in male and female meiosis were compared using the model strain CBA-T6, in which univalency of the small marker autosome pair T6 has been shown to occur very frequently in spermatocytes. Mice from inbred CBA and DBA strains were also analysed. The total frequencies of univalency (sex chromosomes plus autosomes) in metaphase I spermatocytes were 45.6% in CBA, 36.9% in CBA-T6, and 37.3% in DBA males. The aneuploidy in metaphase II spermatocytes ranged from 1.4 to 3% in these strains, which was in agreement with previous findings that most primary spermatocytes with abnormal chromosome configurations are arrested in their development before metaphase II. In the CBA-T6 strain, autosomal univalency at metaphase I mostly involved chromosome pair T6; however, its frequency differed significantly between the sexes, amounting to 18.9% in spermatocytes and 4.3% in oocytes. In the CBA strain, autosomal univalents at metaphase I were seen in 7.7% of the spermatocytes and 1.4% of the oocytes and, in DBA mice, in 4.9% of the spermatocytes and 3.8% of the oocytes. However, in DBA oocytes, when univalency occurred it usually concerned a greater number of bivalents in one cell (range: 2-19 disjoined bivalents), a phenomenon very rare in males of this strain. This study shows that univalent formation differs between the male and female types of meiosis.     

Homeostatic control of recombination is implemented progressively in mouse meiosis

Humans suffer from high rates of fetal aneuploidy, often arising from the absence of meiotic crossover recombination between homologous chromosomes. Meiotic recombination is initiated by double-strand breaks (DSBs) generated by the SPO11 transesterase. In yeast and worms, at least one buffering mechanism, crossover homeostasis, maintains crossover numbers despite variation in DSB numbers. We show here that mammals exhibit progressive homeostatic control of recombination. In wild-type mouse spermatocytes, focus numbers for early recombination proteins (RAD51, DMC1) were highly variable from cell to cell, whereas foci of the crossover marker MLH1 showed little variability. Furthermore, mice with greater or fewer copies of the Spo11 gene--with correspondingly greater or fewer numbers of early recombination foci--exhibited relatively invariant crossover numbers. Homeostatic control is enforced during at least two stages, after the formation of early recombination intermediates and later while these intermediates mature towards crossovers. Thus, variability within the mammalian meiotic program is robustly managed by homeostatic mechanisms to control crossover formation, probably to suppress aneuploidy. Meiotic recombination exemplifies how order can be progressively implemented in a self-organizing system despite natural cell-to-cell disparities in the underlying biochemical processes.     

A link between meiotic prophase progression and crossover control

During meiosis, most organisms ensure that homologous chromosomes undergo at least one exchange of DNA, or crossover, to link chromosomes together and accomplish proper segregation. How each chromosome receives a minimum of one crossover is unknown. During early meiosis in Caenorhabditis elegans and many other species, chromosomes adopt a polarized organization within the nucleus, which normally disappears upon completion of homolog synapsis. Mutations that impair synapsis even between a single pair of chromosomes in C. elegans delay this nuclear reorganization. We quantified this delay by developing a classification scheme for discrete stages of meiosis. Immunofluorescence localization of RAD-51 protein revealed that delayed meiotic cells also contained persistent recombination intermediates. Through genetic analysis, we found that this cytological delay in meiotic progression requires double-strand breaks and the function of the crossover-promoting heteroduplex HIM-14 (Msh4) and MSH-5. Failure of X chromosome synapsis also resulted in impaired crossover control on autosomes, which may result from greater numbers and persistence of recombination intermediates in the delayed nuclei. We conclude that maturation of recombination events on chromosomes promotes meiotic progression, and is coupled to the regulation of crossover number and placement. Our results have broad implications for the interpretation of meiotic mutants, as we have shown that asynapsis of a single chromosome pair can exert global effects on meiotic progression and recombination frequency.     

Meiotic chromosome structure. Kinetochores and chromatid cores in standard and B chromosomes of Arcyptera fusca (Orthoptera) revealed by silver staining.

The behaviour of two chromosome structures in silver-stained chromosomes was analyzed through the first meiotic division in spermatocytes of the acridoid species Arcyptera fusca. Results showed that at diakinesis kinetochores and chromatid cores are individualized while they associate in bivalents of metaphase I; only kinetochores and distal core spots associate in the sex chromosome. Metaphase I is characterized by morphological and localization changes of both kinetochores and cores which define the onset of anaphase I. These changes analyzed in both autosomes and in the sex chromosome allow us to distinguish among three different substages in metaphase I spermatocytes. B chromosomes may be present as univalents, bivalents, or trivalents. Metaphase I B univalents are characterized by separated cores except at their distal ends and individualized and flat sister kinetochores. At anaphase I sister kinetochores of lagging B chromatids remain connected through a silver-stained strand. The behaviour of cores and kinetochores of B bivalents is identical with that found in the autosomal bivalents. The differences in the morphology of kinetochores of every chromosome shown by B trivalents at metaphase I may be related to the balanced forces acting on the multivalent. The results show dramatic changes in chromosome organization of bivalents during metaphase I. These changes suggest that chromatid cores are not involved in the maintenance of bivalents. Moreover, the changes in morphology of kinetochores are independent of the stage of meiosis but correlate with the kind of division (amphitelic-syntelic) that chromosomes undergo.     

Inter-sex variation in synaptonemal complex lengths largely determine the different recombination rates in male and female germ cells

Meiotic chromosomes in human oocytes are packaged differently than in spermatocytes at the pachytene stage of meiosis I, when crossing-over takes place. Thus the meiosis-specific pairing structure, the synaptonemal complex (SC), is considerably longer in oocytes in comparison to spermatocytes. The aim of the present study was to examine the influence of this length factor on meiotic recombination in male and female human germ cells. The positions of crossovers were identified by the DNA mismatch repair protein MLH1. Spermatocytes have approximately 50 crossovers per cell in comparison to more than 70 in oocytes. Analyses of inter-crossover distances (and presumptively crossover interference) along SCs suggested that while there might be inter-individual variation, there was no consistent difference between sexes. Thus the higher rate of recombination in human oocytes is not a consequence of more closely spaced crossovers along the SCs. The rate of recombination per unit length of SC is higher in spermatocytes than oocytes. However, when the so-called obligate chiasma is excluded from the analysis, then the rates of recombination per unit length of SC are essentially identical in the two sexes. Our analyses indicate that the inter-sex difference in recombination is largely a consequence of the difference in meiotic chromosome architecture in the two sexes. We propose that SC length per se, and therefore the size of the physical platform for crossing-over (and not the DNA content) is the principal factor determining the difference in rate of recombination in male and female germ cells. A preliminary investigation of SC loop size by fluorescence in situ hybridization (FISH) indicated loops may be shorter in oocytes than in spermatocytes.     

Competing crossover pathways act during meiosis in Saccharomyces cerevisiae

In Saccharomyces cerevisiae the MSH4-MSH5, MLH1-MLH3, and MUS81-MMS4 complexes act to promote crossing over during meiosis. MSH4-MSH5, but not MUS81-MMS4, promotes crossovers that display interference. A role for MLH1-MLH3 in crossover control is less clear partly because mlh1Delta mutants retain crossover interference yet display a decrease in crossing over that is only slightly less severe than that seen in msh4Delta and msh5Delta mutants. We analyzed the effects of msh5Delta, mlh1Delta, and mms4Delta single, double, and triple mutants on meiotic crossing over at four consecutive genetic intervals on chromosome XV using newly developed computer software. mlh1Delta mms4Delta double mutants displayed the largest decrease in crossing over (13- to 15-fold) of all mutant combinations, yet these strains displayed relatively high spore viability (42%). In contrast, msh5Delta mms4Delta and msh5Delta mms4Delta mlh1Delta mutants displayed smaller decreases in crossing over (4- to 6-fold); however, spore viability (18-19%) was lower in these strains than in mlh1Delta mms4Delta strains. These data suggest that meiotic crossing over can occur in yeast through three distinct crossover pathways. In one pathway, MUS81-MMS4 promotes interference-independent crossing over; in a second pathway, both MSH4-MSH5 and MLH1-MLH3 promote interference-dependent crossovers. A third pathway, which appears to be repressed by MSH4-MSH5, yields deleterious crossovers.     

Differential immunolocalization of a putative Rec8p in meiotic autosomes and sex chromosomes of triatomine bugs

Hemipteran chromosomes are holocentric and show regular, special behavior at meiosis. While the autosomes pair at pachytene, have synaptonemal complexes (SCs) and recombination nodules (RNs) and segregate at anaphase I, the sex chromosomes do not form an SC or RNs, divide equationally at anaphase I, and their chromatids segregate at anaphase II. Here we show that this behavior is shared by the X and Y chromosomes of Triatoma infestans and the X(1)X(2)Y chromosomes of Triatoma pallidipennis. As Rec8p is a widely occurring component of meiotic cohesin, involved in meiotic homolog segregation, we used an antibody against Rec8p of Caenorhabditis elegans for immunolocalization in these triatomines. We show that while Rec8p is colocalized with SCs in the autosomes, no Rec8p can be found by immunolabeling in the sex chromosomes at any stage of meiosis. Furthermore, Rec8p labeling is lost from autosomal bivalents prior to metaphase I. In both triatomine species the sex chromosomes conjoin with each other during prophase I, and lack any SC, but they form "fuzzy cores", which are observed with silver staining and with light and electron microscopy during pachytene. Thin, serial sectioning and electron microscopy of spermatocytes at metaphases I and II reveals differential behavior of the sex chromosomes. At metaphase I the sex chromosomes form separate entities, each surrounded by a membranous sheath. On the other hand, at metaphase II the sex chromatids are closely tied and surrounded by a shared membranous sheath. The peculiar features of meiosis in these hemipterans suggest that they depart from the standard meiotic mechanisms proposed for other organisms.     

An improved method for cytogenetic analysis of meiotic aneuploidy in rodent and frog spermatocytes

Methods are described for the attachment of isolated spermatocytes to glass slides and the subsequent hypotonic swelling and gradual fixation of the metaphase I and metaphase II cells. The methods minimize cell loss and cell disruption and meiotic metaphase chromosomes become spread within residual cytoplasm thus reducing artefactual chromosome loss. Metaphase II complements from mouse, rat and frog spermatocytes prepared by these procedures had relatively low frequencies of hypoploidy (0.5-1.6%). Bivalent loss was not detected in 916 metaphase I complements. Injection of 0.1 mg/kg demecolcine into mice increased the incidence of metaphase II hypoploidy 8-fold. The hypoploid and hyperploid frequencies here increased equally. The results suggest that the methods described may be useful for the analysis of mechanisms of meiotic aneuploidy including aneuploidy resulting from chromosome loss during meiosis I.     

Chromosome-wide regulation of meiotic crossover formation in Caenorhabditis elegans requires properly assembled chromosome axes

Most sexually reproducing organisms depend on the regulated formation of crossovers, and the consequent chiasmata, to accomplish successful segregation of homologous chromosomes at the meiosis I division. A robust, chromosome-wide crossover control system limits chromosome pairs to one crossover in most meioses in the nematode Caenorhabditis elegans; this system has been proposed to rely on structural integrity of meiotic chromosome axes. Here, we test this hypothesis using a mutant, him-3(me80), that assembles reduced levels of meiosis-specific axis component HIM-3 along cohesin-containing chromosome axes. Whereas pairing, synapsis, and crossing over are eliminated when HIM-3 is absent, the him-3(me80) mutant supports assembly of synaptonemal complex protein SYP-1 along some paired chromosomes, resulting in partial competence for chiasma formation. We present both genetic and cytological evidence indicating that the him-3(me80) mutation leads to an increased incidence of meiotic products with two crossovers. These results indicate that limiting the amount of a major axis component results in a reduced capacity to communicate the presence of a (nascent) crossover and/or to discourage others in response.     

Meiotic checkpoints and the interchromosomal effect on crossing over in Drosophila females

During prophase of meiosis I, genetic recombination is initiated with a Spo11-dependent DNA double-strand break (DSB). Repair of these DSBs can generate crossovers, which become chiasmata and are important for the process of chromosome segregation. To ensure at least one chiasma per homologous pair of chromosomes, the number and distribution of crossovers is regulated. One system contributing to the distribution of crossovers is the pachytene checkpoint, which requires the conserved gene pch2 that encodes an AAA+ATPase family member. Pch2-dependent pachytene checkpoint function causes delays in pachytene progression when there are defects in processes required for crossover formation, such as mutations in DSB-repair genes and when there are defects in the structure of the meiotic chromosome axis. Thus, the pachytene checkpoint appears to monitor events leading up to the generation of crossovers. Interestingly, heterozygous chromosome rearrangements cause Pch2-dependent pachytene delays and as little as two breaks in the continuity of the paired chromosome axes are sufficient to evoke checkpoint activity. These chromosome rearrangements also cause an interchromosomal effect on recombination whereby crossing over is suppressed between the affected chromosomes but is increased between the normal chromosome pairs. We have shown that this phenomenon is also due to pachytene checkpoint activity.     

Spo13 maintains centromeric cohesion and kinetochore coorientation during meiosis I

BACKGROUND: The meiotic cell cycle, the cell division cycle that leads to the generation of gametes, is unique in that a single DNA replication phase is followed by two chromosome segregation phases. During meiosis I, homologous chromosomes are segregated, and during meiosis II, as in mitosis, sister chromatids are partitioned. For homolog segregation to occur during meiosis I, physical linkages called chiasmata need to form between homologs, sister chromatid cohesion has to be lost in a stepwise manner, and sister kinetochores must attach to microtubules emanating from the same spindle pole (coorientation). RESULTS: Here we show that the meiosis-specific factor Spo13 functions in two key aspects of meiotic chromosome segregation. In cells lacking SPO13, cohesin, which is the protein complex that holds sister chromatids together, is not protected from removal around kinetochores during meiosis I but is instead lost along the entire length of the chromosomes. We furthermore find that Spo13 promotes sister kinetochore coorientation by maintaining the monopolin complex at kinetochores. In the absence of SPO13, Mam1 and Lrs4 disassociate from kinetochores prematurely during pro-metaphase I and metaphase I, resulting in a partial defect in sister kinetochore coorientation in spo13 Delta cells. CONCLUSIONS: Our results indicate that Spo13 has the ability to regulate both the stepwise loss of sister chromatid cohesion and kinetochore coorientation, two essential features of meiotic chromosome segregation.     

Deficiency of X and Y chromosomal pairing at meiotic prophase in spermatocytes of sterile interspecific hybrids between laboratory mice (Mus domesticus) and Mus spretus

The normal association between the X and Y chromosomes at metaphase I of meiosis, as seen in air-dried light microscope preparations of mouse spermatocytes, is frequently lacking in the spermatocytes of the sterile interspecific hybrid between the laboratory mouse strains C57BL/6 and Mus spretus. The purpose of this work is to determine whether the separate X and Y chromosomes in the hybrid are asynaptic, caused by failure to pair, or desynaptic, caused by precocious dissociation. Unpaired X-Y chromosomes were observed in air-dried preparations at diakinesis, just prior to metaphase I. Furthermore, immunocytology and electron microscopy studies of surface-spread pachytene spermatocytes indicate that the X and Y chromosomes frequently fail to initiate synapsis as judged by the failure to form a synaptonemal complex between the pairing regions of the X and Y Chromosomes. Several additional chromosomal abnormalities were observed in the hybrid. These include fold-backs of the unpaired X or Y cores, associations between the autosome and sex chromosome cores, and autosomal univalents. The occurrence of abnormal autosomal and XY-autosomal associations was also correlated with cell degeneration during meiotic prophase. The primary breakdown in hybrid spermatogenesis occurs at metaphase I (MI), with the appearance of degenerated cells at late MI. In those cells, the X and Y are decondensed rather than condensed as they are in normal mouse MI spermatocytes. These results, in combination with the previous genetic analysis of spermatogenesis in hybrids and backcrosses with fertile female hybrids, suggest that the spermatogenic breakdown in the interspecific hybrid is primarily correlated with the failure of XY pairing at meiotic prophase, asynapsis, followed by the degeneration of spermatocytes at metaphase I. Secondarily, the failure of XY pairing can be accompanied by failure of autosomal pairing, which appears to involve an abnormal sex vesicle and degeneration at pachytene or diplotene.by C. Heyting     

Crossover distribution and high interference for both the X chromosome and an autosome during oogenesis and spermatogenesis in Caenorhabditis elegans

Regulation of both the number and the location of crossovers during meiosis is important for normal chromosome segregation. We used sequence-tagged site polymorphisms to examine the distribution of all crossovers on the X chromosome during oogenesis and on one autosome during both oogenesis and spermatogenesis in Caenorhabditis elegans. The X chromosome has essentially one crossover during oogenesis, with only three possible double crossover exceptions among 220 recombinant X chromosomes. All three had one of the two crossovers in the same chromosomal interval, suggesting that crossovers in that interval do not cause interference. No other interval was associated with double crossovers. Very high interference was also found on an autosome during oogenesis, implying that each chromosome has only one crossover during oogenesis. During spermatogenesis, recombination on this autosome was reduced by approximately 30% compared to oogenesis, but the relative distribution of the residual crossovers was only slightly different. In contrast to previous results with other autosomes, no double crossover chromosomes were observed. Despite an increased frequency of nonrecombinant chromosomes, segregation of a nonrecombinant autosome during spermatogenesis appears to occur normally. This indicates that an achiasmate segregation system helps to ensure faithful disjunction of autosomes during spermatogenesis.