scholarly journals Human NANOS1 Represses Apoptosis by Downregulating Pro-Apoptotic Genes in the Male Germ Cell Line

2020 ◽  
Vol 21 (8) ◽  
pp. 3009
Author(s):  
Damian M. Janecki ◽  
Erkut Ilaslan ◽  
Maciej J. Smialek ◽  
Marcin P. Sajek ◽  
Maciej Kotecki ◽  
...  
Keyword(s):  
Cell Cycle ◽  
Germ Cell ◽  
Cell Line ◽  
Germ Cells ◽  
Male Germ Cell ◽  
Seminiferous Tubules ◽  
Sequencing Analysis ◽  
Germ Cell Line ◽  
Apoptotic Genes

While two mouse NANOS paralogues, NANOS2 and NANOS3, are crucial for maintenance of germ cells by suppression of apoptosis, the mouse NANOS1 paralogue does not seem to regulate these processes. Previously, we described a human NANOS1 p.[(Pro34Thr);(Ser83del)] mutation associated with the absence of germ cells in seminiferous tubules of infertile patients, which might suggest an anti-apoptotic role of human NANOS1. In this study, we aimed to determine a potential influence of human NANOS1 on the maintenance of TCam-2 model germ cells by investigating proliferation, cell cycle, and apoptosis. Constructs encoding wild-type or mutated human NANOS1 were used for transfection of TCam-2 cells, in order to investigate the effect of NANOS1 on cell proliferation, which was studied using a colorimetric assay, as well as apoptosis and the cell cycle, which were measured by flow cytometry. RNA-Seq (RNA sequencing) analysis followed by RT-qPCR (reverse transcription and quantitative polymerase chain reaction) was conducted for identifying pro-apoptotic genes repressed by NANOS1. Here, we show that overexpression of NANOS1 downregulates apoptosis in TCam-2 cells. Moreover, we found that NANOS1 represses a set of pro-apoptotic genes at the mRNA level. We also found that the infertility-associated p.[(Pro34Thr);(Ser83del)] mutation causes NANOS1 to functionally switch from being anti-apoptotic to pro-apoptotic in the human male germ cell line. Thus, this report is the first to show an anti-apoptotic role of NANOS1 exerted by negative regulation of mRNAs of pro-apoptotic genes.

Male germ cell transplantation: promise and problems

2001 ◽  
Vol 13 (8) ◽  
pp. 609 ◽  
Author(s):  
Fang-Xu Jiang
Keyword(s):  
Germ Cell ◽  
Cell Transplantation ◽  
Germ Cells ◽  
Cell Biology ◽  
Direct Injection ◽  
Primordial Germ Cells ◽  
Rete Testis ◽  
Male Germ Cell ◽  
Seminiferous Tubules ◽  
Germ Cell Transplantation

Male germ cell transplantation is a novel technique in which donor male stem germ cells are surgically transferred to the seminiferous tubules of a recipient testis by direct injection or via the rete testis or efferent duct. All germ cells that are destined to become stem spermatogonia are defined as male stem germ cells, including primordial germ cells from the gonadal ridges, and gonocytes and stem spermatogonia from the testis, all of which are transplantable and capable of undergoing normal spermatogenesis. Xenotransplantation of male germ cells from one species into the testis of another species, including human testicular cells in the mouse, has so far proved to be unsuccessful. However, the immunodeficient mouse testis can support rat spermatogenesis and produce apparently normal rat spermatozoa. The underlying mechanisms remain elusive. The present mini-review will focus on the importance of stem spermatogonial transplantation for testicular stem cell biology and discuss the likelihood of immune rejection after transplantation, which may limit the success of all male germ cell transplantation.

scholarly journals TRIM71 Deficiency Causes Germ Cell Loss During Mouse Embryogenesis and Is Associated With Human Male Infertility

2021 ◽  
Vol 9 ◽  
Author(s):  
Lucia A. Torres-Fernández ◽  
Jana Emich ◽  
Yasmine Port ◽  
Sibylle Mitschka ◽  
Marius Wöste ◽  
...  
Keyword(s):  
Male Infertility ◽  
Germ Cell ◽  
Cell Line ◽  
Rna Binding ◽  
Seminiferous Tubules ◽  
Obstructive Azoospermia ◽  
Sequencing Analysis ◽  
Mouse Embryogenesis ◽  
Human Male

Mutations affecting the germline can result in infertility or the generation of germ cell tumors (GCT), highlighting the need to identify and characterize the genes controlling germ cell development. The RNA-binding protein and E3 ubiquitin ligase TRIM71 is essential for embryogenesis, and its expression has been reported in GCT and adult mouse testes. To investigate the role of TRIM71 in mammalian germ cell embryonic development, we generated a germline-specific conditional Trim71 knockout mouse (cKO) using the early primordial germ cell (PGC) marker Nanos3 as a Cre-recombinase driver. cKO mice are infertile, with male mice displaying a Sertoli cell-only (SCO) phenotype which in humans is defined as a specific subtype of non-obstructive azoospermia characterized by the absence of germ cells in the seminiferous tubules. Infertility in male Trim71 cKO mice originates during embryogenesis, as the SCO phenotype was already apparent in neonatal mice. The in vitro differentiation of mouse embryonic stem cells (ESCs) into PGC-like cells (PGCLCs) revealed reduced numbers of PGCLCs in Trim71-deficient cells. Furthermore, TCam-2 cells, a human GCT-derived seminoma cell line which was used as an in vitro model for PGCs, showed proliferation defects upon TRIM71 knockdown. Additionally, in vitro growth competition assays, as well as proliferation assays with wild type and CRISPR/Cas9-generated TRIM71 mutant NCCIT cells showed that TRIM71 also promotes proliferation in this malignant GCT-derived non-seminoma cell line. Importantly, the PGC-specific markers BLIMP1 and NANOS3 were consistently downregulated in Trim71 KO PGCLCs, TRIM71 knockdown TCam-2 cells and TRIM71 mutant NCCIT cells. These data collectively support a role for TRIM71 in PGC development. Last, via exome sequencing analysis, we identified several TRIM71 variants in a cohort of infertile men, including a loss-of-function variant in a patient with an SCO phenotype. Altogether, our work reveals for the first time an association of TRIM71 deficiency with human male infertility, and uncovers further developmental roles for TRIM71 in the germline during mouse embryogenesis.

Migration of gonocytes into the mammalian gonad and their differentiation

1970 ◽  
Vol 259 (828) ◽  
pp. 91-101 ◽  
Keyword(s):  
Alkaline Phosphatase ◽  
Germ Cell ◽  
Cell Line ◽  
Germ Cells ◽  
Primordial Germ Cells ◽  
Histological Investigation ◽  
Controversial Subject ◽  
Germ Cell Line ◽  
The Subject ◽  
Chemical Techniques

The migration of the germ cells into the mammalian gonad and their subsequent differentiation has been the subject of many investigations and controversies. However, sufficient facts have accumulated which led Witschi (1962) to state that ‘the formerly controversial subject of the origin and unbroken continuity of the germ cell lines can now be considered settled’. Histo-chemical techniques for alkaline phosphatase have made it possible to selectively stain and identify primordial germ cells (McKay, Hertig, Adams & Danziger 1953; Chiquoine 1954; Mintz 1959). The path and development of the germ cells has been described in detail in histological investigation (for review see Brambell 1956; Mintz 1960; Franchi, Mandl & Zuckerman 1962) and recently labelling techniques have made the continuity of the germ cell line ‘visible’ (Rudkin & Griech 1962; Peters, Levy & Crone 1962; Kennelly & Foote 1966; Borum 1966; Peters & Crone 1967).

scholarly journals A male infertility mutation reverts NANOS1 activity from anti-apoptotic to pro-apoptotic by disrupting repression of GADD45A, GADD45B, GADD45G and RHOB genes

2019 ◽  
Author(s):  
Damian M. Janecki ◽  
Erkut Ilaslan ◽  
Maciej J. Smialek ◽  
Marcin P. Sajek ◽  
Maciej Kotecki ◽  
...  
Keyword(s):  
Cell Line ◽  
Germ Cells ◽  
Rna Seq ◽  
Wild Type ◽  
Rt Pcr ◽  
Germ Cell Line ◽  
Culture Influence ◽  
Human Reproductive ◽  
Apoptotic Genes ◽  
Infertile Patients

AbstractBackgroundWhile Nanos-mediated maintenance of germ cells in Drosophila and mice has been related to regulation of apoptosis, the relevance of these findings to human physiology is uncertain. Previously we have described the p.[(Pro34Thr);(Ser83del)] double NANOS1 mutation as associated with an absence of germ cells in the testes of infertile patients. The aim of this study was to identify the mechanism underlying infertility phenotype of patients bearing the NANOS1 mutation.MethodsConstructs encoding a wild-type or mutated NANOS1 protein were used for transfection of TCam-2 cell line, representing male germ cells in culture. Influence of this mutation on cell proliferation was performed using MTS assay while apoptosis and cell cycle were measured by flow cytometry. RNA-Seq analysis including quantitative RT-PCR was conducted for selecting pro-apoptotic genes, repressed by the wild-type NANOS1. Influence of the p.[(Pro34Thr);(Ser83del)] NANOS1 mutation on that repression was investigated by quantitative RT-PCR.ResultsWe show here that the p.[(Pro34Thr);(Ser83del)] double NANOS1 mutation causes NANOS1 to functionally switch from being anti-apoptotic to pro-apoptotic in the human male germ cell line TCam-2. This mutation disrupts repression of mRNAs encoding pro-apoptotic GADG45A, GADD45B, GADD45G and RHOB factors, which could contribute to an increase in apoptosis.ConclusionsThis report underscores the conservation of Nanos from flies to humans as a repressor of pro-apoptotic mRNAs in germ cells, and provides a basis for understanding NANOS1 functions in human reproductive health.

scholarly journals Essential Role of Sox2 for the Establishment and Maintenance of the Germ Cell Line

Stem Cells ◽  
2013 ◽  
Vol 31 (7) ◽  
pp. 1408-1421 ◽  
Author(s):  
Federica Campolo ◽  
Manuele Gori ◽  
Rebecca Favaro ◽  
Silvia Nicolis ◽  
Manuela Pellegrini ◽  
...  
Keyword(s):  
Germ Cell ◽  
Cell Line ◽  
Essential Role ◽  
Germ Cell Line

scholarly journals TRIM71 deficiency causes germ cell loss during mouse embryogenesis and promotes human male infertility

2021 ◽  
Author(s):  
Lucia A. Torres-Fernández ◽  
Jana Emich ◽  
Yasmine Port ◽  
Sibylle Mitschka ◽  
Marius Wöste ◽  
...  
Keyword(s):  
Male Infertility ◽  
Germ Cell ◽  
Germ Cells ◽  
Cell Loss ◽  
Seminiferous Tubules ◽  
Obstructive Azoospermia ◽  
Sequencing Analysis ◽  
Mouse Embryogenesis ◽  
Human Male

AbstractMutations affecting the germline can result in infertility or the generation of germ cell tumors (GCT), highlighting the need to identify and characterize the genes controlling the complex molecular network orchestrating germ cell development. TRIM71 is a stem cell-specific factor essential for embryogenesis, and its expression has been reported in GCT and adult mouse testes. To investigate the role of TRIM71 in mammalian germ cell embryonic development, we generated a germline-specific conditional Trim71 knockout mouse (cKO) using the early primordial germ cell (PGC) marker Nanos3 as a Cre-recombinase driver. cKO mice are infertile, with male mice displaying a Sertoli cell-only (SCO) phenotype, which in humans is defined as a specific subtype of non-obstructive azoospermia characterized by the absence of developing germ cells in the testes’ seminiferous tubules. Infertility originates during embryogenesis, as the SCO phenotype was already apparent in neonatal mice. The in vitro differentiation of mouse embryonic stem cells (ESCs) into PGC-like cells (PGCLCs) revealed reduced numbers of PGCLCs in Trim71-deficient cells. Furthermore, in vitro growth competition assays with wild type and CRISPR/Cas9-generated TRIM71 mutant NCCIT cells, a human GCT-derived cell line which we used as a surrogate model for proliferating PGCs, showed that TRIM71 promotes NCCIT cell proliferation and survival. Our data collectively suggest that germ cell loss in cKO mice results from combined defects during the specification and maintenance of PGCs prior to their sex determination in the genital ridges. Last, via exome sequencing analysis, we identified several TRIM71 variants in a cohort of infertile men, including a loss-of-function variant in a patient with SCO phenotype. Our work reveals for the first time an association of TRIM71 variants with human male infertility, and uncovers further developmental roles for TRIM71 in the generation and maintenance of germ cells during mouse embryogenesis.

scholarly journals TEX13B is important for germ cell development and male fertility

2022 ◽  
Author(s):  
Umesh Kumar ◽  
Digumarthi V S Sudhakar ◽  
Nithyapriya Kumar ◽  
Hanuman T Kale ◽  
Rajan Kumar Jha ◽  
...  
Keyword(s):  
Male Infertility ◽  
Germ Cell ◽  
Cell Line ◽  
Germ Cells ◽  
Male Partner ◽  
Epidemiological Studies ◽  
Reproductive Age ◽  
Sequencing Data ◽  
Knock Out

AbstractThe recent epidemiological studies suggest that nearly one out of every 7 reproductive age couples face problem to conceive a child after trying for at least one year. Impaired fertility of the male partner is causative in approximately 50% of the infertile couples. However, the etiologies of large proportion of male infertility are still unclear. Our unpublished exome sequencing data identified several novel genes including TEX13B, which motivated us to further explore the role of TEX13B in male infertility in large infertile case control cohort. Hence in this study, we have examined the role of TEX13B in male infertility by whole gene sequencing 628 infertile and 427 control men and have demonstrated the functional role of Tex13b in spermatogonia GC1spg (GC1) cells. We identified 2 variants on TEX13B which are tightly associated with male infertility. TEX13B gene exclusively expressed in germ cells, but its molecular functions in germ cells are still unknown. Hence, we demonstrated the functional importance of Tex13b in GC1 cell line by genomic manipulation via CRISPR-Cas9 and mass spectrometry-based whole cell proteomics. The gene knock out in GC1 cell line clearly shows that Tex13b play an important role in germ cell growth and morphology. We demonstrate that Tex13b knockout or conditional overexpression in GC1 cells reprograms the metabolic status from an oxidative phosphorylation to glycolysis state and vice versa. In conclusion, our study clearly showed the importance of Tex13b in germ cells development and Its association with male infertility.

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