Cell-fate transition and determination analysis of mouse male germ cells throughout development.
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ABSTRACT:
SUBMITTER: Zhao J
PROVIDER: S-EPMC8617176 | biostudies-literature |
REPOSITORIES: biostudies-literature
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Project description:Testicular development in the mouse is triggered in somatic cells by the function of Sry followed by the activation of fibroblast growth factor 9 (FGF9), which regulates testicular differentiation in both somatic and germ cells. However, the mechanism is unknown. We show here that the nodal/activin signaling pathway is activated in both male germ cells and somatic cells. Disruption of nodal/activin signaling drives male germ cells into meiosis and causes ectopic initiation of female-specific genes in somatic cells. Furthermore, we prove that nodal/activin-A works directly on male germ cells to induce the male-specific gene Nanos2 independently of FGF9. We conclude that nodal/activin signaling is required for testicular development and propose a model in which nodal/activin-A acts downstream of fibroblast growth factor signaling to promote male germ cell fate and protect somatic cells from initiating female differentiation.
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Project description:Continual spermatogenesis is the cornerstone of male fertility and relies on the actions of an undifferentiated spermatogonial population comprised of stem cells and progenitors. A foundational spermatogonial stem cell (SSC) pool is established during postnatal development that serves as a self-renewing reservoir from which progenitor spermatogonia arise that transiently amplify in number before committing to terminal differentiation. At present, the underlying molecular mechanisms governing these actions are undefined. Using conditional mutant mouse models, we investigated whether function of the undifferentiated spermatogonial population during postnatal life is influenced by the tumor suppressor protein RB1. Spermatogenesis initiates in mice with conditional inactivation of Rb1 in prospermatogonial precursors, but the germline is progressively lost upon aging due to impaired renewal of the undifferentiated spermatogonial population. In contrast, continual spermatogenesis is sustained following Rb1 inactivation in progenitor spermatogonia, but some cells transform into a carcinoma in situ-like state. Furthermore, knockdown of Rb1 abundance within primary cultures of wild-type undifferentiated spermatogonia impairs maintenance of the SSC pool, and some cells are invasive of the basement membrane after transplant into recipient testes, indicating acquisition of tumorigenic properties. Collectively, these findings indicate that RB1 plays an essential role in establishment of a self-renewing SSC pool and commitment to the spermatogenic lineage within progenitor spermatogonia.
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Project description:The ability of men to remain fertile throughout their lives depends upon establishment of a spermatogonial stem cell (SSC) pool from gonocyte progenitors, and thereafter balancing SSC renewal versus terminal differentiation. Here, we report that precise regulation of the cell cycle is crucial for this balance. Whereas cyclin-dependent kinase 2 (Cdk2) is not necessary for mouse viability or gametogenesis stages prior to meiotic prophase I, mice bearing a deregulated allele (Cdk2Y15S ) are severely deficient in spermatogonial differentiation. This allele disrupts an inhibitory phosphorylation site (Tyr15) for the kinase WEE1. Remarkably, Cdk2Y15S/Y15S mice possess abnormal clusters of mitotically active SSC-like cells, but these are eventually removed by apoptosis after failing to differentiate properly. Analyses of lineage markers, germ cell proliferation over time, and single cell RNA-seq data revealed delayed and defective differentiation of gonocytes into SSCs. Biochemical and genetic data demonstrated that Cdk2Y15S is a gain-of-function allele causing elevated kinase activity, which underlies these differentiation defects. Our results demonstrate that precise regulation of CDK2 kinase activity in male germ cell development is crucial for the gonocyte-to-spermatogonia transition and long-term spermatogenic homeostasis.
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Project description:BackgroundPartial loss of function of the transcription factor FOXL2 leads to premature ovarian failure in women. In animal models, Foxl2 is required for maintenance, and possibly induction, of female sex determination independently of other critical genes, e.g., Rspo1. Here we report expression profiling of mouse ovaries that lack Foxl2 alone or in combination with Wnt4 or Kit/c-Kit.ResultsFollowing Foxl2 loss, early testis genes (including Inhbb, Dhh, and Sox9) and several novel ovarian genes were consistently dysregulated during embryonic development. In the absence of Foxl2, expression changes affecting a large fraction of pathways were opposite those observed in Wnt4-null ovaries, reinforcing the notion that these genes have complementary actions in ovary development. Loss of one copy of Foxl2 revealed strong gene dosage sensitivity, with molecular anomalies that were milder but resembled ovaries lacking both Foxl2 alleles. Furthermore, a Foxl2 transgene disrupted embryonic testis differentiation and increased the levels of key female markers.ConclusionThe results, including a comprehensive principal component analysis, 1) support the proposal of dose-dependent Foxl2 function and anti-testis action throughout ovary differentiation; and 2) identify candidate genes for roles in sex determination independent of FOXL2 (e.g., the transcription factors IRX3 and ZBTB7C) and in the generation of the ovarian reserve downstream of FOXL2 (e.g., the cadherin-domain protein CLSTN2 and the sphingomyelin synthase SGMS2). The gene inventory is a first step toward the identification of the full range of pathways with partly autonomous roles in ovary development, and thus provides a framework to analyze the genetic bases of female fertility.
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Project description:Primordial germ cells (PGCs) are precursors of gametes that can generate new individuals throughout life in both males and females. Additionally, PGCs have been shown to differentiate into embryonic germ cells (EGCs) after in vitro culture. Most studies investigating germinative cells have been performed in rodents and humans but not dogs (Canis lupus familiaris). Here, we elucidated the dynamics of the expression of pluripotent (POU5F1 and NANOG), germline (DDX4, DAZL and DPPA3), and epigenetic (5mC, 5hmC, H3K27me3 and H3K9me2) markers that are important for the development of male canine germ cells during the early (22-30 days post-fertilization (dpf)), middle (35-40 dpf) and late (45-50 dpf) gestational periods. We performed sex genotype characterization, immunofluorescence, immunohistochemistry, and quantitative reverse transcriptase polymerase chain reaction (RT-qPCR) analyses. Furthermore, in a preliminary study, we evaluated the capacity of canine embryo PGCs (30 dpf) to differentiate into EGCs. To confirm the canine EGCs phenotype, we performed alkaline phosphatase detection, immunohistochemistry, electron and transmission scanning microscopy and RT-qPCR analyses. The PGCs were positive for POU5F1 and H3K27me3 during all assessed developmental periods, including all periods between the gonadal tissue stage and foetal testes development. The number of NANOG, DDX4, DAZL, DPPA3 and 5mC-positive cells increased along with the developing cords from 35-50 dpf. Moreover, our results demonstrate the feasibility of inducing canine PGCs into putative EGCs that present pluripotent markers, such as POU5F1 and the NANOG gene, and exhibit reduced expression of germinative genes and increased expression of H3K27me3. This study provides new insight into male germ cell development mechanisms in dogs.
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Project description:During mouse development, primordial germ cells (PGCs) that give rise to the entire germ line are first identified within the proximal epiblast. However, long-term tracing of the fate of the cells has not been done wherein all cells in and around the germ-cell lineage are identified. Also, quantitative estimates of the number of founder PGCs using different models have come up with various numbers. Here, we use tetrachimeric mice to show that the progenitor numbers for the entire germ line in adult testis, and for the initiating embryonic PGCs, are both 4 cells. Although they proliferate to form polyclonal germ-cell populations in fetal and neonatal testes, germ cells that actually contribute to adult spermatogenesis originate from a small number of secondary founder cells that originate in the fetal period. The rest of the "deciduous" germ cells are lost, most likely by apoptosis, before the reproductive period. The second "actual" founder germ cells generally form small numbers of large monoclonal areas in testes by the reproductive period. Our results also demonstrate that there is no contribution of somatic cells to the male germ cell pool during development or in adulthood. These results suggest a model of 2-step oligoclonal development of male germ cells in mice, the second step distinguishing the heritable germ line from cells selected not to participate in forming the next generation.
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Project description:The lymphatic vasculature, along with the blood vasculature, is a vascular system in our body that plays important functions in fluid homeostasis, dietary fat uptake, and immune responses. Defects in the lymphatic system are associated with various diseases such as lymphedema, atherosclerosis, fibrosis, obesity, and inflammation. The first step in lymphangiogenesis is determining the cell fate of lymphatic endothelial cells. Several genes involved in this commitment step have been identified using animal models, including genetically modified mice. This review provides an overview of these genes in the mammalian system and related human diseases.
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Project description:PIWI-interacting RNAs (piRNAs) are germ cell-specific small RNAs essential for retrotransposon gene silencing and male germ cell development. In piRNA biogenesis, the endonuclease MitoPLD/Zucchini cleaves long, single-stranded RNAs to generate 5' termini of precursor piRNAs (pre-piRNAs) that are consecutively loaded into PIWI-family proteins. Subsequently, these pre-piRNAs are trimmed at their 3'-end by an exonuclease called Trimmer. Recently, poly(A)-specific ribonuclease-like domain-containing 1 (PNLDC1) was identified as the pre-piRNA Trimmer in silkworms. However, the function of PNLDC1 in other species remains unknown. Here, we generate Pnldc1 mutant mice and analyze small RNAs in their testes. Our results demonstrate that mouse PNLDC1 functions in the trimming of both embryonic and post-natal pre-piRNAs. In addition, piRNA trimming defects in embryonic and post-natal testes cause impaired DNA methylation and reduced MIWI expression, respectively. Phenotypically, both meiotic and post-meiotic arrests are evident in the same individual Pnldc1 mutant mouse. The former and latter phenotypes are similar to those of MILI and MIWI mutant mice, respectively. Thus, PNLDC1-mediated piRNA trimming is indispensable for the function of piRNAs throughout mouse spermatogenesis.
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