scholarly journals NANOS2 suppresses the cell cycle by repressing mTORC1 activators in embryonic male germ cells

2020 ◽  
Author(s):  
Ryuki Shimada ◽  
Hiroko Koike ◽  
Takamasa Hirano ◽  
Yumiko Saga
Keyword(s):  
Cell Cycle ◽  
Germ Cell ◽  
Germ Cells ◽  
Rna Binding ◽  
Initial Step ◽  
Sexually Dimorphic ◽  
Wild Type ◽  
Male Germ Cells ◽  
Cycle Arrest ◽  
Male Specific

AbstractDuring murine germ cell development, male germ cells enter the mitotically arrested G0 stage, which is an initial step of sexually dimorphic differentiation. The male specific RNA-binding protein NANOS2 has a key role in suppressing the cell cycle in germ cells. However, the detailed mechanism of how NANOS2 regulates the cell cycle remains unclear. Using single-cell RNA sequencing (scRNA-seq), we extracted the cell cycle state of each germ cell in wild-type and Nanos2-KO testes, and revealed that Nanos2 expression starts in mitotic cells and induces mitotic arrest. We also found that NANOS2 and p38 MAPK work in parallel to regulate the cell cycle, suggesting that several different cascades are involved in the induction of cell cycle arrest. Furthermore, we identified Rheb, a regulator of mTORC1, and Ptma as possible targets of NANOS2. We propose that the repression of the cell cycle is a primary function of NANOS2 and that it is mediated via the suppression of mTORC1 activity by repressing Rheb in a post-transcriptional manner.

Apoptosis in male germ cells in response to cyclin A1-deficiency and cell cycle arrest

2003 ◽  
Vol 66 (8) ◽  
pp. 1571-1579 ◽  
Author(s):  
Glicella Salazar ◽  
Dong Liu ◽  
Ching Liao ◽  
Leah Batkiewicz ◽  
Rachel Arbing ◽  
...  
Keyword(s):  
Cell Cycle ◽  
Cell Cycle Arrest ◽  
Germ Cells ◽  
Cyclin A1 ◽  
Male Germ Cells ◽  
Cycle Arrest

scholarly journals The Golgi Glycoprotein MGAT4D is an Intrinsic Protector of Testicular Germ Cells From Mild Heat Stress

2019 ◽  
Author(s):  
Ayodele Akintayo ◽  
Meng Liang ◽  
Boris Bartholdy ◽  
Frank Batista ◽  
Jennifer Aguilan ◽  
...  
Keyword(s):  
Heat Stress ◽  
Germ Cell ◽  
Germ Cells ◽  
The Body ◽  
Wild Type ◽  
Tgfβ Signaling ◽  
Male Germ Cells ◽  
Mild Heat ◽  
Intrinsic Mechanism ◽  
Shock Response

AbstractMale germ cells are sensitive to heat stress and testes must be maintained outside the body for optimal fertility. However, no germ cell intrinsic mechanism that protects from heat has been reported. Here, we identify the germ cell specific Golgi glycoprotein MGAT4D as a protector of male germ cells from heat stress. Mgat4d is highly expressed in spermatocytes and spermatids. Unexpectedly, when the Mgat4d gene was inactivated globally or conditionally in spermatogonia, or mis-expressed in spermatogonia, spermatocytes or spermatids, neither spermatogenesis nor fertility were affected. On the other hand, when males were subjected to mild heat stress of the testis (43°C for 25 min), germ cells with inactivated Mgat4d were markedly more sensitive to the effects of heat stress, and transgenic mice expressing Mgat4d were partially protected from heat stress. Germ cells lacking Mgat4d generally mounted a similar heat shock response to control germ cells, but could not maintain that response. Several pathways activated by heat stress in wild type were induced to a lesser extent in Mgat4d[−/−] heat-stressed germ cells (NFκB response, TNF and TGFβ signaling, Hif1α and Myc genes). Thus, the Golgi glycoprotein MGAT4D is a novel, intrinsic protector of male germ cells from heat stress.

002. REGULATION OF PLURIPOTENCY AND CELL CYCLE IN FETAL GERM CELLS

2009 ◽  
Vol 21 (9) ◽  
pp. 2
Author(s):  
P. Western ◽  
J. Van Den Bergen ◽  
D. Miles ◽  
R. Ralli ◽  
A. Sinclair
Keyword(s):  
Cell Cycle ◽  
Stem Cells ◽  
Transcriptional Regulation ◽  
Germ Cell ◽  
Germ Cells ◽  
Germ Line ◽  
Cell Lineage ◽  
Mitotic Arrest ◽  
Male Germ Cells ◽  
Developmental Potential

The germ cell lineage is unique in that it must ensure that the genome retains the complete developmental potential (totipotency) that supports development in the following generation. This is achieved through a number of mechanisms that prevent the early germ cell lineage from somatic differentiation and promote the capactity for functional totipotency. Part of this process involves the retained germ line expression of key genes that regulate pluripotency in embryonic stem cells, embryonic germ cells and some embryonal carcinoma cells, the stem cells of testicular tumours. Despite this, germ cells are not intrinsically pluripotent and must differentiate along the male or female pathways, a process which requires commitment of the bi-potential primordial germ cells to the spermatogenic (male) pathway and their entry into mitotic arrest, or to the oogenic pathway (females) and entry into meiosis. This involves robust regulation of regulatory networks controlling pluripotency, cell cycle and sex specific differentiation. Our work aims to further understand the mechanisms controlling differentiation, pluripotency and cell cycle in early male and female germ cells. Our data shows that mitotic arrest of male germ cells involves strict regulation of the G1-S phase check-point through the retinoblastoma protein. In addition, suppression of pluripotency in differentiating male germ cells involves post-transcriptional regulation of OCT4, transcriptional regulation of Sox2 and Nanog and methylation of the Sox2 and Nanog promoters. Further understanding of these processes promises to lead to a greater understanding of the molecular mechanisms underlying control of pluripotency, cell cycle and differentiation in the germ line and the initiation of germ cell derived testis tumours.

scholarly journals Genetic and structural analysis of the in vivo functional redundancy between murine NANOS2 and NANOS3

Development ◽  
2020 ◽  
pp. dev.191916
Author(s):  
Danelle Wright ◽  
Makoto Kiso ◽  
Yumiko Saga
Keyword(s):  
Germ Cell ◽  
Germ Cells ◽  
Rna Binding ◽  
Rna Binding Proteins ◽  
Double Knockout ◽  
Germ Cell Differentiation ◽  
Evolutionarily Conserved ◽  
Male Specific

NANOS2 and NANOS3 are evolutionarily conserved RNA-binding proteins involved in murine germ cell development. NANOS3 is required for protection from apoptosis during migration and gonadal colonization in both sexes, whereas NANOS2 is male-specific and required for the male-type differentiation of germ cells. Ectopic NANOS2 rescues the functions of NANOS3, but NANOS3 cannot rescue NANOS2 function even though its expression is up-regulated in Nanos2-null conditions. It is unknown why NANOS3 cannot rescue NANOS2 function and it is unclear whether NANOS3 plays any role in male germ cell differentiation. To address these questions, we made conditional Nanos3/Nanos2 knockout mice and chimeric mice expressing chimeric NANOS proteins. Conditional double knockout of Nanos2 and 3 led to the rapid loss of germ cells, and in vivo and in vitro experiments revealed that DND1 and NANOS2 binding is dependent on the specific NANOS2 zinc finger structure. Moreover, murine NANOS3 failed to bind CNOT1, an interactor of NANOS2 at its N-terminal. Collectively, our study suggests that the inability of NANOS3 to rescue NANOS2 function is due to poor DND1 recruitment and CNOT1 binding.

scholarly journals Inactivation of CUG-BP1/CELF1 Causes Growth, Viability, and Spermatogenesis Defects in Mice

2006 ◽  
Vol 27 (3) ◽  
pp. 1146-1157 ◽  
Author(s):  
Chantal Kress ◽  
Carole Gautier-Courteille ◽  
H. Beverley Osborne ◽  
Charles Babinet ◽  
Luc Paillard
Keyword(s):  
Germ Cell ◽  
Germ Cells ◽  
Leydig Cells ◽  
Rna Binding ◽  
Rna Binding Protein ◽  
Cell Number ◽  
Pcr Analysis ◽  
Wild Type ◽  
Mammalian Development ◽  
Males And Females

ABSTRACT CUG-BP1/CELF1 is a multifunctional RNA-binding protein involved in the regulation of alternative splicing and translation. To elucidate its role in mammalian development, we produced mice in which the Cugbp1 gene was inactivated by homologous recombination. These Cugbp1 − / − mice were viable, although a significant portion of them did not survive after the first few days of life. They displayed growth retardation, and most Cugbp1 − / − males and females exhibited impaired fertility. Male infertility was more thoroughly investigated. Histological examination of testes from Cugbp1 − / − males showed an arrest of spermatogenesis that occurred at step 7 of spermiogenesis, before spermatid elongation begins, and an increased apoptosis. A quantitative reverse transcriptase PCR analysis showed a decrease of all the germ cell markers tested but not of Sertoli and Leydig markers, suggesting a general decrease in germ cell number. In wild-type testes, CUG-BP1 is expressed in germ cells from spermatogonia to round spermatids and also in Sertoli and Leydig cells. These findings demonstrate that CUG-BP1 is required for completion of spermatogenesis.

scholarly journals Kinesin KIF18A is a novel PUM regulated target promoting mitotic progression and survival of human male germ cell line - TCam-2

2019 ◽  
Author(s):  
Maciej Jerzy Smialek ◽  
Bogna Kuczynska ◽  
Erkut Ilaslan ◽  
Damian Mikolaj Janecki ◽  
Marcin Piotr Sajek ◽  
...  
Keyword(s):  
Cell Cycle ◽  
Germ Cell ◽  
Germ Cells ◽  
Posttranscriptional Regulation ◽  
Rna Binding ◽  
Rna Binding Proteins ◽  
Luciferase Reporter ◽  
Specific Gene ◽  
Transcriptional Level ◽  
Human Male

ABSTRACTRegulation of proliferation, apoptosis and cell cycle is crucial for the physiology of germ cells. Their malfunction contributes to infertility and germ cell tumours. Kinesin KIF18A is an important germ cell specific regulator which downregulates apoptosis while promoting cell proliferation in animal models. Whereas regulation of KIF18A expression was studied at the transcriptional level, its posttranscriptional regulation has not been extensively explored. Due to the presence of two PUM Binding Elements (PBEs) within 3’UTR, KIF18A mRNA is a potential target of PUMs, well known RNA-binding proteins involved in posttranscriptional gene regulation (PTGR). We investigated that possibility in TCam-2 cells originating from seminoma, representing human male germ cells. We conducted RNA co-immunoprecipitation combined with RT-qPCR, as well as luciferase reporter assay by applying appropriate luciferase construct encoding the wild type KIF18A 3’UTR, upon PUM1 and PUM2 overexpression or knockdown. We found that KIF18A is repressed by PUM1 and PUM2. To study how this regulation influences KIF18A function in TCam-2 cells, MTS proliferation assay, apoptosis and cell cycle, analysis using flow cytometry was performed upon KIF18A siRNA knockdown. We uncovered that KIF18A significantly influences proliferation, apoptosis and cell cycle, these effects being opposite to PUM effects in TCam-2 cells. We propose that repression by PUM proteins may represent one of mechanisms influencing KIF18A level in controlling proliferation, cell cycle and apoptosis in TCam-2 cells. To the best of our knowledge, this paper identifies the first mammalian functionally germ cell specific gene that is regulated by Pum proteins via 3’UTR.

scholarly journals The RNA-Binding Protein DND1 Acts Sequentially as a Negative Regulator of Pluripotency and a Positive Regulator of Epigenetic Modifiers Required for Germ Cell Reprogramming

2018 ◽  
Author(s):  
Victor A. Ruthig ◽  
Matthew B. Friedersdorf ◽  
Jason A. Garness ◽  
Steve C. Munger ◽  
Corey Bunce ◽  
...  
Keyword(s):  
Cell Cycle ◽  
Germ Cells ◽  
Rna Binding ◽  
Binding Protein ◽  
Rna Binding Protein ◽  
Embryonic Cell ◽  
Negative Regulator ◽  
Stem Cell Population ◽  
Male Germ Cells ◽  
Mitotic Proliferation

AbstractThe adult spermatogonial stem cell population arises from pluripotent primordial germ cells (PGCs) that enter the fetal testis around embryonic day 10.5 (E10.5). These cells undergo rapid mitotic proliferation, then enter a prolonged period of cell cycle arrest (G1/G0) during which they transition to pro-spermatogonia. In mice homozygous for the Ter mutation in the RNA-binding protein DND1 (DND1Ter/Ter), many germ cells fail to enter G1/G0, and give rise to teratomas, tumors in which many embryonic cell types are represented. To investigate the origin of these tumors, we sequenced the transcriptome of male germ cells in DND1Ter/Ter mutants at E12.5, E13.5, and E14.5, just prior to the formation of teratomas, and correlated this information with direct targets of DND1 identified by DO-RIP-Seq. Consistent with previous results, we found that DND1 controls the down regulation of many genes associated with pluripotency and active cell cycle, including elements of the mTor, Hippo and Bmp/Nodal signaling pathways. However, DND1 targets also include genes associated with male differentiation including a large group of chromatin regulators activated in wild type but not mutant germ cells during the transition between E13.5 and E14.5. These results suggest multiple functions of DND1, and link DND1 to the initiation of epigenetic modifications in male germ cells.

scholarly journals MCG10, a Novel p53 Target Gene That Encodes a KH Domain RNA-Binding Protein, Is Capable of Inducing Apoptosis and Cell Cycle Arrest in G2-M

2000 ◽  
Vol 20 (15) ◽  
pp. 5602-5618 ◽  
Author(s):  
Jianhui Zhu ◽  
Xinbin Chen
Keyword(s):  
Cell Cycle ◽  
Cell Proliferation ◽  
Cell Cycle Arrest ◽  
Target Gene ◽  
Rna Binding ◽  
Rna Binding Protein ◽  
Wild Type ◽  
Cycle Arrest ◽  
Kh Domain ◽  
P53 Target Gene

ABSTRACT p53, a tumor suppressor, inhibits cell proliferation by inducing cellular genes involved in the regulation of the cell cycle.MCG10, a novel cellular p53 target gene, was identified in a cDNA subtraction assay with mRNA isolated from a p53-producing cell line. MCG10 can be induced by wild-type but not mutant p53 and by DNA damage via two potential p53-responsive elements in the promoter of the MCG10 gene. The MCG10 gene contains 10 exons and is located at chromosome 3p21, a region highly susceptible to aberrant chromosomal rearrangements and deletions in human neoplasia. The MCG10 gene locus encodes at least two alternatively spliced transcripts, MCG10 and MCG10as. The MCG10 and MCG10as proteins contain two domains homologous to the heterogeneous nuclear ribonucleoprotein K homology (KH) domain. By generating cell lines that inducibly express either wild-type or mutated forms of MCG10 and MCG10as, we found that MCG10 and MCG10as can suppress cell proliferation by inducing apoptosis and cell cycle arrest in G2-M. In addition, we found that MCG10 and MCG10as, through their KH domains, can bind poly(C) and that their RNA-binding activity is necessary for inducing apoptosis and cell cycle arrest. Furthermore, we found that the level of the poly(C) binding MCG10 protein is increased in cells treated with the DNA-damaging agent camptothecin in a p53-dependent manner. These results suggest that the MCG10 RNA-binding protein is a potential mediator of p53 tumor suppression.

scholarly journals Multi-modal regulation of C. elegans hermaphrodite spermatogenesis by the GLD-1-FOG-2 complex

2018 ◽  
Author(s):  
Shuang Hu ◽  
Lauren E. Ryan ◽  
Ebru Kaymak ◽  
Lindsay Freeberg ◽  
Te-Wen Lo ◽  
...  
Keyword(s):  
Sex Determination ◽  
Germ Cell ◽  
Germ Cells ◽  
Translational Control ◽  
Rna Binding ◽  
Rna Binding Proteins ◽  
Wild Type ◽  
C Elegans ◽  
Target Mrnas ◽  
Mrna Targets

AbstractProper germ cell sex determination in Caenorhabditis nematodes requires a network of RNA-binding proteins (RBPs) and their target mRNAs. In some species, changes in this network enabled limited XX spermatogenesis, and thus self-fertility. In C. elegans, one of these selfing species, the global sex-determining gene tra-2 is regulated in germ cells by a conserved RBP, GLD-1, via the 3’ untranslated region (3’UTR) of its transcript. A C. elegans-specific GLD-1 cofactor, FOG-2, is also required for hermaphrodite sperm fate, but how it modifies GLD-1 function is unknown. Germline feminization in gld-1 and fog-2 null mutants has been interpreted as due to cell-autonomous elevation of TRA-2 translation. Consistent with the proposed role of FOG-2 in translational control, the abundance of nearly all GLD-1 target mRNAs (including tra-2) is unchanged in fog-2 mutants. Epitope tagging reveals abundant TRA-2 expression in somatic tissues, but an undetectably low level in wild-type germ cells. Loss of gld-1 function elevates germline TRA-2 expression to detectable levels, but loss of fog-2 function does not. A simple quantitative model of tra-2 activity constrained by these results can successfully sort genotypes into normal or feminized groups. Surprisingly, fog-2 and gld-1 activity enable the sperm fate even when GLD-1 cannot bind to the tra-2 3’ UTR. This suggests the GLD-1-FOG-2 complex regulates uncharacterized sites within tra-2, or other mRNA targets. Finally, we quantify the RNA-binding capacities of dominant missense alleles of GLD-1 that act genetically as “hyperrepressors” of tra-2 activity. These variants bind RNA more weakly in vitro than does wild-type GLD-1. These results indicate that gld-1 and fog-2 regulate germline sex via multiple interactions, and that our understanding of the control and evolution of germ cell sex determination in the C. elegans hermaphrodite is far from complete.

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