Sperm abnormalities in the PL/J mouse strain: A description and proposed mechanism for malformation

A high frequency (42%) of sperm from the inbred homozygous mouse strain PL/J are abnormal. Head shape abnormalities occur in 15% of the total sperm; and 27% of the sperm are headless, with the mitochondria condensed into a mass at the caudal end of the midpiece region. The sperm without heads exhibit relatively normal motility. Electron microscopy of the testes indicates that some of the abnormal sperm in PL/J males result from a failure of the paired centrioles to attain a normal position on the nucleus opposite the acrosome prior to implantation, or to attach at all. The centrioles that are not attached to the nuclear envelope can differentiate to form the principal piece and midpiece region. The frequency of headless variants in heterozygous F₁ indicates that the trait is mainly recessive. The offspring from the backcross of the F₁ to homozygous PL/J parents did not give a clear-cut segregation pattern. The frequency of abnormal sperm in the F₁ and the backcross is higher when the female parent is a PL/J.     

Microtubule organization and distribution of gamma-tubulin in male meiosis of lepidoptera

Meiotic spindles in males of higher Lepidotera are unusual in that the bulk of the spindle microtubules (MTs) ends about halfway between the equatorial plate and the centrosomes in metaphase. It appears worthwhile to determine how the MTs are nucleated, while their pole proximal ends are distant from the centrosomes. To this end, spermatocytes of Phragmatobia fuliginosa (Arctiidae), collected in the field, were double-labeled with antibodies to beta- and gamma-tubulin. The former antibody reveals the entire microtubular cytoskeleton, and the latter is directed against a newly-discovered tublin isoform that is prevalent in microtubule-organizing centers (MTOCs). The immunocytochemical work was supplemented by a fine structural analysis of MTOCs and spindles. Gamma-tubulin was clearly detected at the spindle poles, and prominent microtubular asters originated from these sites. Additionally, MT arrays at both sides of the equatorial plate in metaphase spermatocytes contained gamma-tubulin. The staining persisted in late anaphase, when kinetochore MTs are depolymerized. This indicates that at least nonkinetochore MTs contain gamma-tubulin. The analysis of ultrathin sections through spindles revealed large amounts of pericentriolar material at the spindles poles, in prometaphase through anaphase. The spindle MTs appeared as regular, straight elements in longitudinal sections. We assume that gamma-tubulin is located at the pole proximal ends of the MTs and/or is associated with the spindle MTs throughout their lengths. In order to distinguish between these possibilities, testes of Ephestia kuehniella (Pyralidae), a laboratory species, were cold-treated prior to double-labeling with antibodies to beta- and gamma-tubulin. The treatment was expected to depolymerize MTs. Astral MTs, which were nucleated end-on by gamma-tubulin-containing material, indeed depolymerized. In contrast, the gamma-tubulin-containing spindle MTs persisted. It is, therefore, conceivable that gamma-tubulin is associated with MTs throughout their lengths in male meiosis of Lepidoptera species. It is plausible that this association stabilizes the MTs against cold-induced disassembly. © 1996 Wiley-Liss, Inc.     

Genetic Interactions between the Aurora Kinases Reveal New Requirements for AURKB and AURKC during Oocyte Meiosis

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Meiosis I in Xenopus oocytes is not error-prone despite lacking spindle assembly checkpoint

The spindle assembly checkpoint, SAC, is a surveillance mechanism to control the onset of anaphase during cell division. SAC prevents anaphase initiation until all chromosome pairs have achieved bipolar attachment and aligned at the metaphase plate of the spindle. In doing so, SAC is thought to be the key mechanism to prevent chromosome nondisjunction in mitosis and meiosis. We have recently demonstrated that Xenopus oocyte meiosis lacks SAC control. This prompted the question of whether Xenopus oocyte meiosis is particularly error-prone. In this study, we have karyotyped a total of 313 Xenopus eggs following in vitro oocyte maturation. We found no hyperploid egg, out of 204 metaphase II eggs with countable chromosome spreads. Therefore, chromosome nondisjunction is very rare during Xenopus oocyte meiosis I, despite the lack of SAC.     

Meeting the meiotic challenge: Specializations in mammalian oocyte spindle formation

Oocytes uniquely accumulate cytoplasmic constituents to support early embryogenesis. This unique specialization is accompanied by acquisition of a large size and by execution of asymmetric meiotic divisions that preserve precious ooplasm through the expulsion of minimal size polar bodies. While often taken for granted, these basic features of oogenesis necessitate unique specializations of the meiotic apparatus. These include a chromatin‐sourced RanGTP gradient that restricts spindle size by defining a spatial domain where meiotic spindles form, acentriolar centrosomes that rely on microtubule organizing centers to form spindle poles, and an actin‐based mechanism for asymmetric spindle positioning. Additionally, localized protein synthesis to support spindle formation is achieved in the spindle forming region, whilst protein synthesis is reduced elsewhere in the ooplasm. This is achieved through enrichment of spindle‐related mRNAs in the spindle forming region combined with local PLK1‐mediated phosphorylation and inactivation of the translational repressor EIF4EBP1. This allows PLK1 to function as an important regulatory nexus through which endogenous and exogenous signals can impact spindle formation and function, and highlights the important role that PLK1 may have in maintaining oocyte quality and fertility.     

Age‐dependent aneuploidy in mammalian oocytes instigated at the second meiotic division

Ageing severely affects the chromosome segregation process in human oocytes resulting in aneuploidy, infertility and developmental disorders. A considerable amount of segregation errors in humans are introduced at the second meiotic division. We have here compared the chromosome segregation process in young adult and aged female mice during the second meiotic division. More than half of the oocytes in aged mice displayed chromosome segregation irregularities at anaphase II, resulting in dramatically increased level of aneuploidy in haploid gametes, from 4% in young adult mice to 30% in aged mice. We find that the post‐metaphase II process that efficiently corrects aberrant kinetochore‐microtubule attachments in oocytes in young adult mice is approximately 10‐fold less efficient in aged mice, in particular affecting chromosomes that show small inter‐centromere distances at the metaphase II stage in aged mice. Our results reveal that post‐metaphase II processes have critical impact on age‐dependent aneuploidy in mammalian eggs.     

RBFOX2缺失抑制雄性小鼠的减数分裂起始

减数分裂起始是配子发生的关键步骤。目前人们已经发现了一些减数分裂起始所必需的基因,但是对于该过程的调控基因及其作用机制还知之甚少。本实验室先前建立了精原干细胞（spermatogonial stem cells,SSCs）体外培养以及体外诱导减数分裂起始的技术体系,为更好地探究减数分裂起始调控基因及作用机理提供了良好的条件。实验室前期研究发现RNA结合蛋白RBFOX2可能调控减数分裂起始,然而RBFOX2在生殖细胞的减数分裂起始过程中的功能及作用机制还需深入研究。本研究利用慢病毒介导的转基因技术产生了RBFOX2敲降的精原干细胞系;发现RBFOX2敲降后,精原干细胞的自我更新、增殖以及分化未发生显著变化,但是减数分裂无法启动,分化型精原细胞发生明显凋亡。这一结果进一步显示了RNA结合蛋白在雄性动物减数分裂起始中的重要功能。     

MEIOSIN Directs the Switch from Mitosis to Meiosis in Mammalian Germ Cells

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The evolutionary ecology of testicular function: size isn't everything

Larger testes are considered the quintessential adaptation to sperm competition. However, the strong focus on testis size in evolutionary research risks ignoring other potentially adaptive features of testicular function, many of which will also be shaped by post‐mating sexual selection. Here we advocate a more integrated research programme that simultaneously takes into account the developmental machinery of spermatogenesis and the various selection pressures that act on this machinery and its products. The testis is a complex organ, and so we begin by outlining how we can think about the evolution of testicular function both in terms of the composition and spatial organisation of the testis (‘testicular histology’), as well as in terms of the logical organisation of cell division during spermatogenesis (‘testicular architecture’). We then apply these concepts to ask which aspects of testicular function we can expect to be shaped by post‐mating sexual selection. We first assess the impact of selection on those traits most strongly associated with sperm competition, namely the number and kind of sperm produced. A broad range of studies now support our contention that post‐mating sexual selection affects many aspects of testicular function besides gross testis size, for example, to maximise spermatogenic efficiency or to enable the production of particular sperm morphologies. We then broaden our focus to ask how testicular function is affected by fluctuation in sperm demand. Such fluctuation can occur over an individual's lifetime (for example due to seasonality in reproduction) and may select for particular types of testicular histology and architecture depending on the particular reproductive ecology of the species in question. Fluctuation in sperm demand also occurs over evolutionary time, due to shifts in the mating system, and this may have various consequences for testicular function, for example on rates of proliferation‐induced mutation and for dealing with intragenomic conflict. We end by suggesting additional approaches that could be applied to study testicular function, and conclude that simultaneously considering the machinery, products and scheduling of spermatogenesis will be crucial as we seek to understand more fully the evolution of this most fundamental of male reproductive traits.     

Anatomy and ultrastructure of the male reproductive system in Pleioplana atomata (Platyhelminthes: Polycladida)

Abstract. The ultrastructure of the male reproductive system in the polyclad flatworm Pleioplana atomata is described. Numerous testes are scattered throughout the entire body but are heavily concentrated on the ventral side. All stages of differentiating sperm cells are present in all testes follicles. Intercellular bridges connect spermatocytes and spermatids derived from a single spermatogonium. In the distal part of spermatids, a zone of differentiation develops with a row of microtubules beneath the plasmalemma. Adjacent to these microtubules, an intercentriolar body is flanked by two basal bodies that give rise to two axonemes (each with a 9+“1” microtubular pattern) that face in opposite directions. The Golgi complex appears in the central portion of the spermatid and produces numerous small and large electron-dense bodies. The small bodies surround the nucleus, whereas the large bodies cluster along with the mitochondria in the central part of the spermatid. Development of the spermatid leads to cell elongation and formation of a filiform, biflagellate mature spermatozoon with cortical microtubules all along the sperm shaft. The male canal system consists of paired vasa deferentia that separately enter a single seminal vesicle. A single prostatic canal connects the seminal vesicle to the prostatic vesicle. Ultrastructurally, the seminal vesicle and prostatic canal are very similar, and along with the prostatic vesicle and stylet pocket, are lined by a ciliated epithelium. The ultrastructure of the prostatic vesicle indicates that it probably produces a large volume of seminal fluid that, along with spermatozoa, is transferred to the mating partner through a stylet. Some of the findings, particularly on sperm ultrastructure, may provide characters useful for phylogenetic analysis.     

Reduced ability to recover from spindle disruption and loss of kinetochore spindle assembly checkpoint proteins in oocytes from aged mice

Currently, maternal aging in women, based on mouse models, is thought to raise oocyte aneuploidy rates, because chromosome cohesion deteriorates during prophase arrest, and Sgo2, a protector of centromeric cohesion, is lost. Here we show that the most common mouse strain, C57Bl6/J, is resistant to maternal aging, showing little increase in aneuploidy or Sgo2 loss. Instead it demonstrates significant kinetochore-associated loss in the spindle assembly checkpoint protein Mad2 and phosphorylated Aurora C, which is involved in microtubule–kinetochore error correction. Their loss affects the fidelity of bivalent segregation but only when spindle organization is impaired during oocyte maturation. These findings have an impact clinically regarding the handling of human oocytes ex vivo during assisted reproductive techniques and suggest there is a genetic basis to aneuploidy susceptibility.     

Deficiency of ovarian ornithine decarboxylase contributes to aging‐related egg aneuploidy in mice

It has been known for more than four decades that during mammalian estrous cycles, luteinizing hormone stimulates a transitory rise in the ovaries of ornithine decarboxylase (ODC) activity and its enzymatic product putrescine, concurrent with oocyte maturation in vivo. Inhibition of this transitory ODC/putrescine rise, however, does not appear to affect oocyte maturation or ovulation. Using several mouse models and combining in vitro and in vivo approaches, we demonstrated that deficiency of ODC during oocyte maturation is correlated with increased levels of egg aneuploidies. These results suggest that the transitory ovarian ODC rise in late proestrus is important for ensuring proper chromosome segregation during oocyte maturation. Older mice (8 months of age) exhibited about 1/3 that of young mice in LH‐stimulated ovarian ODC activity and a corresponding increase in egg aneuploidies. Moreover, a combination of putrescine supplementation in mouse drinking water leading up to oocyte retrieval and in oocyte maturation medium reduced egg aneuploidies of the older mice from 12.7% to 5.3%. Therefore, ovarian ODC deficiency might be an important etiology of maternal aging‐related aneuploidies, and peri‐ovulatory putrescine supplementation might reduce the risk of aneuploid conceptions in older women.     

同翅类昆虫的雄性生殖系统及精子发生(昆虫纲:半翅目)

本文比较了同翅类昆虫雄性生殖系统的结构、减数分裂期间染色体的行为和精子尾部的超微结构。研究表明蜡蝉总科和异翅类的精巢具有被膜,而蝉总科、叶蝉总科、沫蝉总科、角蝉总科、木虱总科、蚜总科、粉虱总科和蚧总科的精巢均不具有被膜。也可以根据精巢小叶的形状将精巢分为三类,蝉总科、叶蝉总科、沫蝉总科、角蝉总科、蚜总科和粉虱总科的精巢小叶为球形,蜡蝉总科、木虱总科和蚧总科的精巢小叶为管状,而异翅类的精巢小叶为片层状。减数分裂可以被分为5类:①蝉型(Cicadoidtype);②蜡蝉型(Fulgoroidtype);③木虱型(Psyloidtype);④蚜型(Aphidoidtype);⑤粉虱型(Aleyrodoidtype)和⑥蚧型(Coccoidtype),至少具有四个类群的减数分裂前期I具有弥散期,它们是:木虱总科、蜡蝉总科、蚧总科和异翅类。除粉虱总科和蚧总科的精子尾部退化以外,其余种类的精子鞭毛均具有典型的9 9 2轴丝结构。     

X-Y chromosome univalency in the testes of hyperthermic mice: I. Concomitant formation of multinucleated giant cells

Exposure of mice to an environment of approximately 35°C and 65% relative humidity for 2–5 days caused dissociation of the X-Y chromosome bivalent in diakinesis-metaphase I (primary) spermatocytes. Dissociation of autosomal bivalents, chromosome fragmentation, aneuploidy, and polyploidy were not significantly affected by heat stress. Heat stress also caused formation of multinucleated giant cells in the mouse testis. Giant cell formation followed a distribution similar to X-Y dissociation, reaching highest levels 5 days after heat stress and returning to control levels 50–70 days after heat stress. Spermatogonial activity was not permanently damaged by these conditions, and the germinal epithelium was able to repopulate. We believe that cell aberration or cell death resulting from either or both of these anomalies causes much of the decreased fertility found after subjecting scrotal mammals to high temperatures.     

Spermatozoa of chromosomally heterozygous mice and their fate in male and female genital tracts

The fate of morphologically normal but chromosomally abnormal spermatozoa derived from mice with variable degrees and complexity of Robertsonian heterozygosity was studied at different sites along the male and female genital tract by Feulgen-DNA measurements. In addition, the percentage frequencies of morphologically abnormal spermatozoa in transit along the male and female genital tracts were studied. It was found that during transit from the epididymis to the vas deferens the distribution of the Feulgen-DNA contents of morphologically normal spermatozoa changed: Spermatozoa with chromatin with the extremely low or high Feulgen staining intensity disappeared. The percentages of morphologically abnormal sperm cells did not change at these levels. In the female genital tracts, the distribution of Feulgen-DNA content of morphologically normal spermatozoa did not show significant changes. This indicates that spermatozoa are able to reach the fallopian tube in spite of gross genome unbalance. There is evidence that unbalanced spermatozoa take part in the fertilization process, producing abnormal zygotes subject to postzygotic loss. Conversely morphologically abnormal spermatozoa were preferentially lost before they reached the fallopian tube, suggesting they had been eliminated prezygotically.     

Meiosis and speciation: A study in a speciatingMus terricolor complex

The three chromosomal species of theMus terricolor complex possess 2n = 40 chromosomes. We show that their karyotypes differ in stable heterochromatin variations fixed in homozygous condition as prominent short arms in autosomes 1, 3 and 6. The three chromosomal species exhibit a high incidence of polymorphisms for Robertsonian fusions and pericentric inversions. Breeding experiments and histological analysis of testis show that heterozygosity for pericentric inversions and Robertsonian fusions had no effect on fertility. Meiotic analysis shows normal overall progression of meiosis in the heterozygotes, which is consistent with their normal gametogenesis. Nevertheless, both the inversion and fusion heterozygotes had undergone some alterations in the regular process of homologous synapsis, and it appeared that certain features of the meiotic system circumvented the potential negative effects of these polymorphic chromosomal rearrangements. The results indicate that the attributes of the meiotic system in a given organism could modulate the potential of a chromosomal rearrangement as reproductive barrier. The meiotic modulation hypothesis offers an explanation for the contradictory effects of the similar kinds of chromosomal mutations reported in different species.     

Loss of TDP-43 in male germ cells causes meiotic failure and impairs fertility in mice

Meiotic arrest is a common cause of human male infertility, but the causes of this arrest are poorly understood. Transactive response DNA-binding protein of 43 kDa (TDP-43) is highly expressed in spermatocytes in the preleptotene and pachytene stages of meiosis. TDP-43 is linked to several human neurodegenerative disorders wherein its nuclear clearance accompanied by cytoplasmic aggregates underlies neurodegeneration. Exploring the functional requirement for TDP-43 for spermatogenesis for the first time, we show here that conditional KO (cKO) of the Tardbp gene (encoding TDP-43) in male germ cells of mice leads to reduced testis size, depletion of germ cells, vacuole formation within the seminiferous epithelium, and reduced sperm production. Fertility trials also indicated severe subfertility. Spermatocytes of cKO mice showed failure to complete prophase I of meiosis with arrest at the midpachytene stage. Staining of synaptonemal complex protein 3 and γH2AX, markers of the meiotic synaptonemal complex and DNA damage, respectively, and super illumination microscopy revealed nonhomologous pairing and synapsis defects. Quantitative RT–PCR showed reduction in the expression of genes critical for prophase I of meiosis, including Spo11 (initiator of meiotic double-stranded breaks), Rec8 (meiotic recombination protein), and Rad21L (RAD21-like, cohesin complex component), as well as those involved in the retinoic acid pathway critical for entry into meiosis. RNA-Seq showed 1036 upregulated and 1638 downregulated genes (false discovery rate <0.05) in the Tardbp cKO testis, impacting meiosis pathways. Our work reveals a crucial role for TDP-43 in male meiosis and suggests that some forms of meiotic arrest seen in infertile men may result from the loss of function of TDP-43.