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Genes Apr 2021Nuclear architecture undergoes an extensive remodeling during spermatogenesis, especially at levels of spermatocytes (SPC) and spermatids (SPT). Interestingly, typical... (Review)
Review
Nuclear architecture undergoes an extensive remodeling during spermatogenesis, especially at levels of spermatocytes (SPC) and spermatids (SPT). Interestingly, typical events of spermiogenesis, such as nuclear elongation, acrosome biogenesis, and flagellum formation, need a functional cooperation between proteins of the nuclear envelope and acroplaxome/manchette structures. In addition, nuclear envelope plays a key role in chromosome distribution. In this scenario, special attention has been focused on the LINC (linker of nucleoskeleton and cytoskeleton) complex, a nuclear envelope-bridge structure involved in the connection of the nucleoskeleton to the cytoskeleton, governing mechanotransduction. It includes two integral proteins: KASH- and SUN-domain proteins, on the outer (ONM) and inner (INM) nuclear membrane, respectively. The LINC complex is involved in several functions fundamental to the correct development of sperm cells such as head formation and head to tail connection, and, therefore, it seems to be important in determining male fertility. This review provides a global overview of the main LINC complex components, with a special attention to their subcellular localization in sperm cells, their roles in the regulation of sperm morphological maturation, and, lastly, LINC complex alterations associated to male infertility.
Topics: Animals; Cell Nucleus; Cytoskeleton; Humans; Infertility, Male; Male; Mechanotransduction, Cellular; Nuclear Envelope; Nuclear Matrix; Spermatids; Spermatocytes; Spermatozoa
PubMed: 33925685
DOI: 10.3390/genes12050658 -
Frontiers in Cell and Developmental... 2021Meiosis is a highly conserved and essential process in gametogenesis in sexually reproducing organisms. However, there are substantial sex-specific differences within... (Review)
Review
Meiosis is a highly conserved and essential process in gametogenesis in sexually reproducing organisms. However, there are substantial sex-specific differences within individual species with respect to meiosis-related chromatin reorganization, recombination, and tolerance for meiotic defects. A wide range of murine models have been developed over the past two decades to study the complex regulatory processes governing mammalian meiosis. The present review article thus provides a comprehensive overview of the knockout mice that have been employed to study meiosis, with a particular focus on gene- and gametogenesis-related sexual dimorphism observed in these model animals. In so doing, we aim to provide a firm foundation for the future study of sex-specific differences in meiosis at the molecular level.
PubMed: 34041246
DOI: 10.3389/fcell.2021.670599 -
Molecular Biology of the Cell Aug 2023Centrosomes are essential parts of diverse cellular processes, and precise regulation of the levels of their constituent proteins is critical for their function. One...
Centrosomes are essential parts of diverse cellular processes, and precise regulation of the levels of their constituent proteins is critical for their function. One such protein is Pericentrin (PCNT) in humans and Pericentrin-like protein (PLP) in . Increased PCNT expression and its protein accumulation are linked to clinical conditions including cancer, mental disorders, and ciliopathies. However, the mechanisms by which PCNT levels are regulated remain underexplored. Our previous study demonstrated that PLP levels are sharply down-regulated during early spermatogenesis and this regulation is essential to spatially position PLP on the proximal end of centrioles. We hypothesized that the sharp drop in PLP protein was a result of rapid protein degradation during the male germ line premeiotic G2 phase. Here, we show that PLP is subject to ubiquitin-mediated degradation and identify multiple proteins that promote the reduction of PLP levels in spermatocytes, including the UBR box containing E3 ligase Poe (UBR4), which we show binds to PLP. Although protein sequences governing posttranslational regulation of PLP are not restricted to a single region of the protein, we identify a region that is required for Poe-mediated degradation. Experimentally stabilizing PLP, via internal PLP deletions or loss of Poe, leads to PLP accumulation in spermatocytes, its mispositioning along centrioles, and defects in centriole docking in spermatids.
Topics: Male; Humans; Ubiquitin-Protein Ligases; Centrioles; Centrosome; Antigens
PubMed: 37342879
DOI: 10.1091/mbc.E22-11-0534 -
Development (Cambridge, England) Jul 2023Valosin-containing protein (VCP) binds and extracts ubiquitylated cargo to regulate protein homeostasis. VCP has been studied primarily in aging and disease contexts,...
Valosin-containing protein (VCP) binds and extracts ubiquitylated cargo to regulate protein homeostasis. VCP has been studied primarily in aging and disease contexts, but it also affects germline development. However, the precise molecular functions of VCP in the germline, particularly in males, are poorly understood. Using the Drosophila male germline as a model system, we find that VCP translocates from the cytosol to the nucleus as germ cells transition into the meiotic spermatocyte stage. Importantly, nuclear translocation of VCP appears to be one crucial event stimulated by testis-specific TBP-associated factors (tTAFs) to drive spermatocyte differentiation. VCP promotes the expression of several tTAF-target genes, and VCP knockdown, like tTAF loss of function, causes cells to arrest in early meiotic stages. At a molecular level, VCP activity supports spermatocyte gene expression by downregulating a repressive histone modification, mono-ubiquitylated H2A (H2Aub), during meiosis. Remarkably, experimentally blocking H2Aub in VCP-RNAi testes is sufficient to overcome the meiotic-arrest phenotype and to promote development through the spermatocyte stage. Collectively, our data highlight VCP as a downstream effector of tTAFs that downregulates H2Aub to facilitate meiotic progression.
Topics: Animals; Male; Spermatocytes; Valosin Containing Protein; Cell Differentiation; Drosophila; Testis; Gene Expression; Spermatogenesis; Meiosis
PubMed: 37401420
DOI: 10.1242/dev.201557 -
Fertility and Sterility Apr 2023To obtain de novo male gametes capable of inducing full preimplantation blastocyst development using a novel three-dimensional (3D) culture system.
OBJECTIVE
To obtain de novo male gametes capable of inducing full preimplantation blastocyst development using a novel three-dimensional (3D) culture system.
DESIGN
Mouse embryonic stem cells (mESCs) were spherified by plunging in sodium alginate followed by calcium chloride, delineating a 3D environment that simulates the seminiferous tubule. As a control, mESCs cultured on two-dimensional plates were used. Plates and spheres containing mESCs from both methods were exposed to Activin-A, bFGF, and KSR followed by exposure to BMP4, LIF, SCF, and EGF to promote differentiation into male germ-like cells.
MAIN OUTCOME MEASURES
Cells were assessed for VASA, DAZL, and BOULE on days 3 and 10. Cells were later injected into activated oocytes and monitored using time-lapse imaging on days 15, 22, 29, and 36. Control conceptuses generated using mature epididymal spermatozoa were also monitored via time-lapse imaging.
RESULTS
On day 3, cells differentiated on plates expressed VASA at 1% and DAZL at 29%. In spheres, VASA was expressed at a rate of 15% and DAZL at a rate of 45% (P<.001). On day 10, cells differentiated on plates had VASA expression of 7%, DAZL of 23%, and BOULE of only 0.5%. Cells differentiated into spheres expressed VASA at a rate of 20%, DAZL at 43%, and BOULE at 10% (P<.001). Subsequent differentiation in spheres on day 3 exhibited a DAZL (expressed in spermatogonia) expression of 43% and a VASA (further spermatogenesis progression) expression of 15%. On day 10, DAZL and VASA expressions were reassessed and increased to 45% and 18%, respectively. BOULE, a marker expressed solely in postmeiotic spermatocytes, was expressed at 8%, whereas acrosin was expressed in spermatids at 2%. On day 15, VASA expression plateaued at 17%, BOULE peaked at 10%, and acrosin reached 5%. On day 22, expression of VASA increased to 19%, BOULE decreased to 8%, and acrosin peaked at 7%. On day 29, VASA expression peaked at 20%, BOULE dropped to 2%, and acrosin remained stable at 7%. On day 36, VASA expression remained at 13%, whereas BOULE and acrosin expression decreased to 0% and 1%, respectively. The control cohort attained 88.4% fertilization and 76.9% blastocyst rates. De novo gametes achieved fertilization rates of 35.0%, 61.1%, 81.8%, and 50.0% on days 15, 22, 29, and 36, respectively. Neogametes-generated blastocyst rates were 5.0%, 16.7%, 36.4%, and 8.3% for days 15, 22, 29, and 36, respectively.
CONCLUSION
Our novel 3D differentiation model can generate functional gametes and is aimed at obviating the need for allogeneic/xenogeneic transplantation. The decreased overall marker expression and the reduced blastocyst development indicated that intrasphere germ cell differentiation correlated with the length of mouse spermatogenesis at approximately 30 days. Future experiments will be conducted to confirm the reproducibility of our findings and the eventual generation of offspring.
Topics: Male; Animals; Mice; Acrosin; Haploidy; Reproducibility of Results; Spermatozoa; Spermatogenesis; Spermatocytes
PubMed: 36706828
DOI: 10.1016/j.fertnstert.2023.01.021 -
Communications Biology Oct 2023Caseinolytic protease proteolytic subunit (ClpP) and caseinolytic protease X (ClpX) are mitochondrial matrix peptidases that activate mitochondrial unfolded protein...
Caseinolytic protease proteolytic subunit (ClpP) and caseinolytic protease X (ClpX) are mitochondrial matrix peptidases that activate mitochondrial unfolded protein response to maintain protein homeostasis in the mitochondria. However, the role of ClpP and ClpX in spermatogenesis remains largely unknown. In this study, we demonstrated the importance of ClpP/ClpX for meiosis and spermatogenesis with two conditional knockout (cKO) mouse models. We found that ClpP/ClpX deficiency reduced mitochondrial functions and quantity in spermatocytes, affected energy supply during meiosis and attenuated zygotene-pachytene transformation of the male germ cells. The dysregulated spermatocytes finally underwent apoptosis resulting in decreased testicular size and vacuolar structures within the seminiferous tubules. We found mTORC1 pathway was over-activated after deletion of ClpP/ClpX in spermatocytes. Long-term inhibition of the mTORC1 signaling via rapamycin treatment in vivo partially rescue spermatogenesis. The data reveal the critical roles of ClpP and ClpX in regulating meiosis and spermatogenesis.
Topics: Animals; Male; Mice; Mitochondria; Peptide Hydrolases; Serine Endopeptidases; Spermatocytes; Spermatogenesis; Endopeptidase Clp
PubMed: 37798322
DOI: 10.1038/s42003-023-05372-2 -
Andrology Jul 2023The germline perpetuates genetic information across generations. To maintain the integrity of the germline, transposable elements in the genome must be silenced, as... (Review)
Review
BACKGROUND
The germline perpetuates genetic information across generations. To maintain the integrity of the germline, transposable elements in the genome must be silenced, as these mobile elements would otherwise engender widespread mutations passed on to subsequent generations. There are several well-established mechanisms that are dedicated to providing defense against transposable elements, including DNA methylation, RNA interference, and the PIWI-interacting RNA pathway.
OBJECTIVES
Recently, several studies have provided evidence that transposon defense is not only provided by factors dedicated to this purpose but also factors with other roles, including in germline development. Many of these are transcription factors. Our objective is to summarize what is known about these "bi-functional" transcriptional regulators.
MATERIALS AND METHODS
Literature search.
RESULTS AND CONCLUSION
We summarize the evidence that six transcriptional regulators-GLIS3, MYBL1, RB1, RHOX10, SETDB1, and ZBTB16-are both developmental regulators and transposable element-defense factors. These factors act at different stages of germ cell development, including in pro-spermatogonia, spermatogonial stem cells, and spermatocytes. Collectively, the data suggest a model in which specific key transcriptional regulators have acquired multiple functions over evolutionary time to influence developmental decisions and safeguard transgenerational genetic information. It remains to be determined whether their developmental roles were primordial and their transposon defense roles were co-opted, or vice versa.
Topics: Male; Humans; DNA Transposable Elements; Gene Expression Regulation, Developmental; Spermatogonia; Transcription Factors; Spermatocytes; RNA, Small Interfering; Germ Cells
PubMed: 36895139
DOI: 10.1111/andr.13427 -
The Journal of Clinical Investigation Oct 2023Maturation arrest (MA) is a subtype of non-obstructive azoospermia, and male infertility is a known risk factor for testicular tumors. However, the genetic basis for...
Maturation arrest (MA) is a subtype of non-obstructive azoospermia, and male infertility is a known risk factor for testicular tumors. However, the genetic basis for many affected individuals remains unknown. Here, we identified a deleterious hemizygous variant of X-linked retinoblastoma-binding protein 7 (RBBP7) as a potential key cause of MA, which was also found to be associated with the development of Leydig cell tumors. This mutation resulted in premature protein translation termination, affecting the sixth WD40 domain of the RBBP7 and the interaction of the mutated RBBP7 with histone H4. Decreased BRCA1 and increased γH2AX were observed in the proband. In mouse spermatogonial and pachytene spermatocyte-derived cells, deprivation of rbbp7 led to cell cycle arrest and apoptosis. In Drosophila, knockdown of RBBP7/Caf1-55 in germ cells resulted in complete absence of germ cells and reduced testis size, whereas knockdown of RBBP7/Caf1-55 in cyst cells resulted in hyperproliferative testicular cells. Interestingly, male infertility caused by Caf1-55 deficiency was rescued by ectopic expression of wild-type human RBBP7 but not mutant variants, suggesting the importance of RBBP7 in spermatogenesis. Our study provides insights into the mechanisms underlying the co-occurrence of MA and testicular tumors and may pave the way for innovative genetic diagnostics of these 2 diseases.
Topics: Animals; Humans; Male; Mice; Azoospermia; Infertility, Male; Mutation; Retinoblastoma-Binding Protein 7; Spermatogenesis; Testicular Neoplasms; Testis
PubMed: 37843278
DOI: 10.1172/JCI171541 -
Biology of Reproduction Sep 2019Mammalian spermatogenesis is a complex developmental program that transforms mitotic testicular germ cells (spermatogonia) into mature male gametes (sperm) for... (Review)
Review
Mammalian spermatogenesis is a complex developmental program that transforms mitotic testicular germ cells (spermatogonia) into mature male gametes (sperm) for production of offspring. For decades, it has been known that this several-weeks-long process involves a series of highly ordered and morphologically recognizable cellular changes as spermatogonia proliferate, spermatocytes undertake meiosis, and spermatids develop condensed nuclei, acrosomes, and flagella. Yet, much of the underlying molecular logic driving these processes has remained opaque because conventional characterization strategies often aggregated groups of cells to meet technical requirements or due to limited capability for cell selection. Recently, a cornucopia of single-cell transcriptome studies has begun to lift the veil on the full compendium of gene expression phenotypes and changes underlying spermatogenic development. These datasets have revealed the previously obscured molecular heterogeneity among and between varied spermatogenic cell types and are reinvigorating investigation of testicular biology. This review describes the extent of available single-cell RNA-seq profiles of spermatogenic and testicular somatic cells, how those data were produced and evaluated, their present value for advancing knowledge of spermatogenesis, and their potential future utility at both the benchtop and bedside.
Topics: Animals; Humans; Male; Mammals; RNA-Seq; Single-Cell Analysis; Spermatogenesis; Transcriptome; Translational Research, Biomedical
PubMed: 31077285
DOI: 10.1093/biolre/ioz088 -
Vavilovskii Zhurnal Genetiki I Selektsii Oct 2023Germline-restricted chromosomes (GRCs) are present in the genomes of germline cells and absent from somatic cells. A GRC is found in all species of the songbirds...
Germline-restricted chromosomes (GRCs) are present in the genomes of germline cells and absent from somatic cells. A GRC is found in all species of the songbirds (Passeri) and in none of the other bird orders studied to date. This indicates that GRC originated in the common ancestor of the songbirds. The germline-restricted chromosome is permanently absent from somatic cells of the songbird, while female germline cells usually contain two copies of GRC and male ones have one copy. In females, GRCs undergo synapsis and restricted recombination in their terminal regions during meiotic prophase. In males, it is almost always eliminated from spermatocytes. Thus, GRC is inherited almost exclusively through the maternal lineage. The germline-restricted chromosome is a necessary genomic element in the germline cells of songbirds. To date, the GRC genetic composition has been studied in four species only. Some GRC genes are actively expressed in female and male gonads, controlling the development of germline cells and synthesis of the proteins involved in the organization of meiotic chromosomes. Songbird species vary in GRC size and genetic composition. The GRC of each bird species consists of amplified and modified copies of genes from the basic genome of that species. The level of homology between GRCs of different species is relatively low, indicating a high rate of genetic evolution of this chromosome. Transmission through the maternal lineage and suppression of the recombination contribute significantly to the accelerated evolution of GRCs. One may suggest that the rapid coordinated evolution between the GRC genes and the genes of the basic genome in the songbirds might be responsible for the explosive speciation and adaptive radiation of this most species-rich and diverse infraorder of birds.
PubMed: 38023808
DOI: 10.18699/VJGB-23-75