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Staged developmental mapping and X chromosome transcriptional dynamics during mouse spermatogenesis.Nature Communications Mar 2019Male gametes are generated through a specialised differentiation pathway involving a series of developmental transitions that are poorly characterised at the molecular...
Male gametes are generated through a specialised differentiation pathway involving a series of developmental transitions that are poorly characterised at the molecular level. Here, we use droplet-based single-cell RNA-Sequencing to profile spermatogenesis in adult animals and at multiple stages during juvenile development. By exploiting the first wave of spermatogenesis, we both precisely stage germ cell development and enrich for rare somatic cell-types and spermatogonia. To capture the full complexity of spermatogenesis including cells that have low transcriptional activity, we apply a statistical tool that identifies previously uncharacterised populations of leptotene and zygotene spermatocytes. Focusing on post-meiotic events, we characterise the temporal dynamics of X chromosome re-activation and profile the associated chromatin state using CUT&RUN. This identifies a set of genes strongly repressed by H3K9me3 in spermatocytes, which then undergo extensive chromatin remodelling post-meiosis, thus acquiring an active chromatin state and spermatid-specific expression.
Topics: Animals; Cell Separation; Chromatin; Chromosome Mapping; Chromosomes, Human, Pair 21; Epigenesis, Genetic; Female; Flow Cytometry; Gene Expression Profiling; Gene Expression Regulation, Developmental; Histones; Humans; Male; Meiotic Prophase I; Mice; Mice, Inbred C57BL; Mice, Transgenic; Sequence Analysis, RNA; Single-Cell Analysis; Spermatocytes; Spermatogenesis; Testis; Transcription, Genetic; X Chromosome
PubMed: 30890697
DOI: 10.1038/s41467-019-09182-1 -
Molecular Human Reproduction Sep 2010Mammalian oocytes are arrested at prophase I until puberty when luteinizing hormone (LH) induces resumption of meiosis of follicle-enclosed oocytes. Resumption of... (Review)
Review
Mammalian oocytes are arrested at prophase I until puberty when luteinizing hormone (LH) induces resumption of meiosis of follicle-enclosed oocytes. Resumption of meiosis is tightly coupled with regulating cyclin-dependent kinase 1 (CDK1) activity. Prophase I arrest depends on inhibitory phosphorylation of CDK1 and anaphase-promoting complex-(APC-CDH1)-mediated regulation of cyclin B levels. Prophase I arrest is maintained by endogenously produced cyclic adenosine monophosphate (cAMP), which activates protein kinase A (PKA) that in turn phosphorylates (and activates) the nuclear kinase WEE2. In addition, PKA-mediated phosphorylation of the phosphatase CDC25B results in its cytoplasmic retention. The combined effect maintains low levels of CDK1 activity that are not sufficient to initiate resumption of meiosis. LH triggers synthesis of epidermal growth factor-like factors in mural granulosa cells and leads to reduced cGMP transfer from cumulus cells to oocytes via gap junctions that couple the two cell types. cGMP inhibits oocyte phosphodiesterase 3A (PDE3A) and a decline in oocyte cGMP results in increased PDE3A activity. The ensuing decrease in oocyte cAMP triggers maturation by alleviating the aforementioned phosphorylations of WEE2 and CDC25B. As a direct consequence CDC25B translocates into the nucleus. The resulting activation of CDK1 also promotes extrusion of WEE2 from the nucleus thereby providing a positive amplification mechanism for CDK1 activation. Other kinases, e.g. protein kinase B, Aurora kinase A and polo-like kinase 1, also participate in resumption of meiosis. Mechanisms governing meiotic prophase I arrest and resumption of meiosis share common features with DNA damage-induced mitotic G2-checkpoint arrest and checkpoint recovery, respectively. These common features include CDC14B-dependent activation of APC-CDH1 in prophase I arrested oocytes or G2-arrested somatic cells, and CDC25B-dependent cell cycle resumption in both oocytes and somatic cells.
Topics: Age Factors; Anaphase-Promoting Complex-Cyclosome; Animals; Cdh1 Proteins; Cell Cycle Proteins; Cell Proliferation; Cyclic AMP; Cyclin B1; Cyclin-Dependent Kinase Inhibitor Proteins; Epidermal Growth Factor; Female; G2 Phase; Meiosis; Meiotic Prophase I; Metaphase; Mice; Oocytes; Oogenesis; Signal Transduction; Ubiquitin-Protein Ligase Complexes
PubMed: 20453035
DOI: 10.1093/molehr/gaq034 -
Epigenetic regulator Cfp1 safeguards male meiotic progression by regulating meiotic gene expression.Experimental & Molecular Medicine Aug 2022Meiosis occurs specifically in germ cells to produce sperm and oocytes that are competent for sexual reproduction. Multiple factors are required for successful meiotic...
Meiosis occurs specifically in germ cells to produce sperm and oocytes that are competent for sexual reproduction. Multiple factors are required for successful meiotic entry, progression, and termination. Among them, trimethylation of histone H3 on lysine 4 (H3K4me3), a mark of active transcription, has been implicated in spermatogenesis by forming double-strand breaks (DSBs). However, the role of H3K4me in transcriptional regulation during meiosis remains poorly understood. Here, we reveal that mouse CXXC finger protein 1 (Cfp1), a component of the H3K4 methyltransferase Setd1a/b, is dynamically expressed in differentiating male germ cells and safeguards meiosis by controlling gene expression. Genetic ablation of mouse CFP1 in male germ cells caused complete infertility with failure in prophase I of the 1st meiosis. Mechanistically, CFP1 binds to genes essential for spermatogenesis, and its loss leads to a reduction in H3K4me3 levels and gene expression. Importantly, CFP1 is highly enriched within the promoter/TSS of target genes to elevate H3K4me3 levels and gene expression at the pachytene stage of meiotic prophase I. The most enriched genes were associated with meiosis and homologous recombination during the differentiation of spermatocytes to round spermatids. Therefore, our study establishes a mechanistic link between CFP1-mediated transcriptional control and meiotic progression and might provide an unprecedented genetic basis for understanding human sterility.
Topics: Animals; Epigenesis, Genetic; Gene Expression; Histone-Lysine N-Methyltransferase; Humans; Male; Meiosis; Methylation; Mice; Semen; Trans-Activators
PubMed: 35918532
DOI: 10.1038/s12276-022-00813-0 -
Frontiers in Cell and Developmental... 2021Defects in crossover (CO) formation during meiosis are a leading cause of birth defects, embryonic lethality, and infertility. In a wide range of species, maternal aging...
Defects in crossover (CO) formation during meiosis are a leading cause of birth defects, embryonic lethality, and infertility. In a wide range of species, maternal aging increases aneuploidy and decreases oocyte quality. In which produce oocytes throughout the first half of adulthood, aging both decreases oocytes quality and increases meiotic errors. Phenotypes of mutations in genes encoding double-strand break (DSB)-associated proteins get more severe with maternal age suggesting that early meiosis reflects a particularly sensitive node during reproductive aging in the worm. We observed that aging has a direct effect on the integrity of meiotic CO formation, as observed by an increase of univalent chromosomes and fusions at diakinesis, with a considerable increase starting at 4 days. We also characterize the possible causes for the age-related changes in CO formation by analyzing both steady-state levels and kinetics of the ssDNA binding proteins RPA-1 and RAD-51. Profound reductions in numbers of both RPA-1 and RAD-51 foci suggests that both DSB formation and early meiotic repair are compromised in aging worms. Using laser microirradiation and γ-irradiation to induce exogenous damage, we show specifically that recruitment of these homologous recombination proteins is altered. Repair defects can be seen in two-and-one-half day-old adults making the loss of germline repair capacity among the earliest aging phenotypes in the worm.
PubMed: 34422819
DOI: 10.3389/fcell.2021.695333 -
International Review of Cell and... 2011Fertilization results from the fusion of male and female gametes in all sexually reproducing organisms. Much of nematode fertility work was focused on Caenorhabditis... (Review)
Review
Fertilization results from the fusion of male and female gametes in all sexually reproducing organisms. Much of nematode fertility work was focused on Caenorhabditis elegans and Ascaris suum. The C. elegans hermaphrodite produces a limited number of sperm initially and then commits to the exclusive production of oocytes. The postmeiotic differentiation called spermiogenesis converts sessile spermatids into motile spermatozoa. The motility of spermatozoa depends on dynamic assembly and disassembly of a major sperm protein-based cytoskeleton uniquely found in nematodes. Both self-derived and male-derived spermatozoa are stored in spermatheca, the site of fertilization in hermaphrodites. The oocyte is arrested in meiotic prophase I until a sperm-derived signal relieves the inhibition allowing the meiotic maturation to occur. Oocyte undergoes meiotic maturation, enters into spermatheca, gets fertilized, completes meiosis, and exits into uterus as a zygote. This review focuses on our current understanding of the events around fertilization in nematodes.
Topics: Animals; Fertilization; Nematoda; Oocytes; Sperm Motility; Sperm-Ovum Interactions; Spermatogenesis
PubMed: 21749902
DOI: 10.1016/B978-0-12-386039-2.00006-7 -
American Journal of Physiology. Cell... Oct 2022In female mammals, the size of the initially established primordial follicle pool within the ovaries determines the reproductive life span. Interestingly, the...
In female mammals, the size of the initially established primordial follicle pool within the ovaries determines the reproductive life span. Interestingly, the establishment of the primordial follicle pool is accompanied by a remarkable programmed oocyte loss for unclear reasons. Here, we identify a new role of ASH1-like histone lysine methyltransferase (ASH1L) in controlling the apoptosis of oocytes during meiotic prophase I in mice. Our results showed that overexpression of led to a dramatic loss of fetal oocytes via apoptosis, which subsequently resulted in a reduced capacity of the primordial follicle pool. Overexpression of also led to a deficiency in DNA double-strand break repair associated with premature upregulation of p63 and phosphorylated checkpoint kinase 2 (p-CHK2), the major genome guardian of the female germline, following overexpression in fetal ovaries. In summary, ASH1L is one of the indispensable epigenetic molecules that acts as a guardian of the genome. It protects oocyte genome integrity and removes oocytes with serious DNA damage by regulating the expression of p63 and p-CHK2 during meiotic prophase I in mice. Our study provides a perspective on the physiological regulatory role of DNA damage checkpoint signaling in fetal oocyte guardianship and female fertility.
Topics: Animals; Apoptosis; Checkpoint Kinase 2; DNA Damage; DNA-Binding Proteins; Female; Histone-Lysine N-Methyltransferase; Mammals; Meiosis; Mice; Oocytes
PubMed: 36094439
DOI: 10.1152/ajpcell.00196.2022 -
Cells Dec 2022Cilia are hair-like projections of the plasma membrane with an inner microtubule skeleton known as axoneme. Motile cilia and flagella beat to displace extracellular...
Cilia are hair-like projections of the plasma membrane with an inner microtubule skeleton known as axoneme. Motile cilia and flagella beat to displace extracellular fluids, playing important roles in the airways and reproductive system. On the contrary, primary cilia function as cell-type-dependent sensory organelles, detecting chemical, mechanical, or optical signals from the extracellular environment. Cilia dysfunction is associated with genetic diseases called ciliopathies and with some types of cancer. Cilia have been recently identified in zebrafish gametogenesis as an important regulator of conformation and recombination. However, there is little information about the structure and functions of cilia in mammalian meiosis. Here we describe the presence of cilia in male mouse meiotic cells. These solitary cilia formed transiently in 20% of zygotene spermatocytes and reached considerable lengths (up to 15-23 µm). CEP164 and CETN3 localization studies indicated that these cilia emanate from the mother centriole prior to centrosome duplication. In addition, the study of telomeric TFR2 suggested that cilia are not directly related to the conformation during early male mouse meiosis. Instead, based on TEX14 labeling of intercellular bridges in spermatocyte cysts, we suggest that mouse meiotic cilia may have sensory roles affecting cyst function during prophase I.
Topics: Mice; Male; Animals; Female; Humans; Centrioles; Cilia; Centrosome; Meiotic Prophase I; Zebrafish; Mothers; Meiosis; Mammals; Transcription Factors
PubMed: 36611937
DOI: 10.3390/cells12010142 -
Development (Cambridge, England) Jun 2021Spermatogenesis is precisely controlled by complex gene-expression programs. During mammalian male germ-cell development, a crucial feature is the repression of...
Spermatogenesis is precisely controlled by complex gene-expression programs. During mammalian male germ-cell development, a crucial feature is the repression of transcription before spermatid elongation. Previously, we discovered that the RNA-binding protein EWSR1 plays an important role in meiotic recombination in mouse, and showed that EWSR1 is highly expressed in late meiotic cells and post-meiotic cells. Here, we used an Ewsr1 pachytene stage-specific knockout mouse model to study the roles of Ewsr1 in late meiotic prophase I and in spermatozoa maturation. We show that loss of EWSR1 in late meiotic prophase I does not affect proper meiosis completion, but does result in defective spermatid elongation and chromocenter formation in the developing germ cells. As a result, male mice lacking EWSR1 after pachynema are sterile. We found that, in Ewsr1 CKO round spermatids, transition from a meiotic gene-expression program to a post-meiotic and spermatid gene expression program related to DNA condensation is impaired, suggesting that EWSR1 plays an important role in regulation of spermiogenesis-related mRNA synthesis necessary for spermatid differentiation into mature sperm.
Topics: Animals; Gene Expression Regulation, Developmental; Male; Meiosis; Meiotic Prophase I; Mice; Mice, Knockout; RNA-Binding Protein EWS; Spermatids; Spermatogenesis; Spermatozoa
PubMed: 34100066
DOI: 10.1242/dev.199414 -
G3 (Bethesda, Md.) Feb 2020A trispecific hybrid, MTP (hereafter called tripsazea), was developed from intergeneric crosses involving tetraploid (2 = 4 = 40, genome: MMMM), tetraploid (2 = 4 =...
A trispecific hybrid, MTP (hereafter called tripsazea), was developed from intergeneric crosses involving tetraploid (2 = 4 = 40, genome: MMMM), tetraploid (2 = 4 = 72, TTTT), and tetraploid (2 = 4 = 40, PPPP). On crossing maize- (2 = 4 = 56, MMTT) with , 37 progenies with varying chromosome numbers (36-74) were obtained, and a special one (, tripsazea) possessing 2 = 74 chromosomes was generated. Tripsazea is perennial and expresses phenotypic characteristics affected by its progenitor parent. Flow cytometry analysis of tripsazea and its parents showed that tripsazea underwent DNA sequence elimination during allohexaploidization. Of all the chromosomes in diakinesis I, 18.42% participated in heterogenetic pairing, including 16.43% between the M- and P-genomes, 1.59% between the M- and T-genomes, and 0.39% in T- and P-genome pairing. Tripsazea is male sterile and partly female fertile. In comparison with previously synthesized trihybrids containing maize, and teosinte, tripsazea has a higher chromosome number, higher seed setting rate, and vegetative propagation ability of stand and stem. However, few trihybrids possess these valuable traits at the same time. The potential of tripsazea is discussed with respect to the deployment of the genetic bridge for maize improvement and forage breeding.
Topics: Chromosomes, Plant; Crosses, Genetic; Genome, Plant; Hybridization, Genetic; Karyotype; Phenotype; Plant Breeding; Poaceae; Polyploidy; Reproduction; Zea mays
PubMed: 31792004
DOI: 10.1534/g3.119.400942 -
PLoS Genetics May 2021Meiosis is a cell division process with complex chromosome events where various molecules must work in tandem. To find meiosis-related genes, we screened evolutionarily...
Meiosis is a cell division process with complex chromosome events where various molecules must work in tandem. To find meiosis-related genes, we screened evolutionarily conserved and reproductive tract-enriched genes using the CRISPR/Cas9 system and identified potassium channel tetramerization domain containing 19 (Kctd19) as an essential factor for meiosis. In prophase I, Kctd19 deficiency did not affect synapsis or the DNA damage response, and chiasma structures were also observed in metaphase I spermatocytes of Kctd19 KO mice. However, spermatocytes underwent apoptotic elimination during the metaphase-anaphase transition. We were able to rescue the Kctd19 KO phenotype with an epitope-tagged Kctd19 transgene. By immunoprecipitation-mass spectrometry, we confirmed the association of KCTD19 with zinc finger protein 541 (ZFP541) and histone deacetylase 1 (HDAC1). Phenotyping of Zfp541 KO spermatocytes demonstrated XY chromosome asynapsis and recurrent DNA damage in the late pachytene stage, leading to apoptosis. In summary, our study reveals that KCTD19 associates with ZFP541 and HDAC1, and that both KCTD19 and ZFP541 are essential for meiosis in male mice.
Topics: Anaphase; Animals; CRISPR-Cas Systems; Cell Cycle Proteins; Cell Nucleus; Chromosomal Proteins, Non-Histone; Chromosome Pairing; Conserved Sequence; DNA Damage; Evolution, Molecular; Fertility; Genes, Essential; Histone Deacetylase 1; Male; Meiosis; Meiotic Prophase I; Metaphase; Mice; Nuclear Proteins; Pachytene Stage; Phenotype; Spermatids; Spermatocytes; Testis; Transcription Factors; Transgenes
PubMed: 33961623
DOI: 10.1371/journal.pgen.1009412