Embryonic Testis Research Articles

Two redundant transcription factor binding sites in a single enhancer are essential for mammalian sex determination.

Male development in mammals depends on the activity of the two SOX gene: Sry and Sox9, in the embryonic testis. As deletion of Enhancer 13 (Enh13) of the Sox9 gene results in XY male-to-female sex reversal, we explored the critical elements necessary for its function and hence, for testis and male development. Here, we demonstrate that while microdeletions of individual transcription factor binding sites (TFBS) in Enh13 lead to normal testicular development, combined microdeletions of just two SRY/SOX binding motifs can alone fully abolish Enh13 activity leading to XY male-to-female sex reversal. This suggests that for proper male development to occur, these few nucleotides of non-coding DNA must be intact. Interestingly, we show that depending on the nature of these TFBS mutations, dramatically different phenotypic outcomes can occur, providing a molecular explanation for the distinct clinical outcomes observed in patients harboring different variants in the same enhancer.

Rhox8 homeobox gene ablation leads to rete testis abnormality and male subfertility in mice†.

The reproductive homeobox X-linked (Rhox) genes encode transcription factors that are expressed selectively in reproductive tissues including the testis, epididymis, ovary, and placenta. While many Rhox genes are expressed in germ cells in the mouse testis, only Rhox8 is expressed exclusively in the Sertoli cells during embryonic and postnatal development, suggesting a possible role of Rhox8 in embryonic gonad development. Previously, Sertoli cell-specific knockdown of RHOX8 resulted in male subfertility due to germ cell defects. However, this knockdown model was limited in examining the functions of Rhox8 as RHOX8 knockdown occurred only postnatally, and there was still residual RHOX8 in the testis. In this study, we generated new Rhox8 knockout (KO) mice using the CRISPR/Cas9 system. Sex determination and fetal testis development were apparently normal in mutant mice. Fertility analysis showed a low fecundity in Rhox8 KO adult males, with disrupted spermatogenic cycles, increased germ cell apoptosis, and reduced sperm count and motility. Interestingly, Rhox8 KO testes showed an increase in testis size with dilated seminiferous tubules and rete testis, which might be affected by efferent duct (ED) Rhox8 ablation dysregulating the expression of metabolism and transport genes in the EDs. Taken together, the data presented in this study suggest that Rhox8 in the Sertoli cells is not essential for sex determination and embryonic testis differentiation but has an important role in complete spermatogenesis and optimal male fertility.

Loss of Dead end1 induces testicular teratomas from primordial germ cells that failed to undergo sexual differentiation in embryonic testes

Spontaneous testicular teratomas (STTs) are tumours comprising a diverse array of cell and tissue types, which are derived from pluripotent stem-like cells called embryonal carcinoma cells (ECCs). Although mouse ECCs originate from primordial germ cells (PGCs) in embryonic testes, the molecular basis underlying ECC development remains unclear. This study shows that the conditional deletion of mouse Dead end1 (Dnd1) from migrating PGCs leads to STT development. In Dnd1-conditional knockout (Dnd1-cKO) embryos, PGCs colonise the embryonic testes but fail to undergo sexual differentiation; subsequently, ECCs develop from a portion of the PGCs. Transcriptomic analyses reveal that PGCs not only fail to undergo sexual differentiation but are also prone to transformation into ECCs by upregulating the expression of marker genes for primed pluripotency in the testes of Dnd1-cKO embryos. Thus, our results clarify the role of Dnd1 in developing STTs and developmental process of ECC from PGC, providing novel insights into pathogenic mechanisms of STTs.

Turnover of mammal sex chromosomes in the Sry-deficient Amami spiny rat is due to male-specific upregulation of Sox9

Mammalian sex chromosomes are highly conserved, and sex is determined by SRY on the Y chromosome. Two exceptional rodent groups in which some species lack a Y chromosome and Sry offer insights into how novel sex genes can arise and replace Sry, leading to sex chromosome turnover. However, intensive study over three decades has failed to reveal the identity of novel sex genes in either of these lineages. We here report our discovery of a male-specific duplication of an enhancer of Sox9 in the Amami spiny rat Tokudaia osimensis, in which males and females have only a single X chromosome (XO/XO) and the Y chromosome and Sry are completely lost. We performed a comprehensive survey to detect sex-specific genomic regions in the spiny rat. Sex-related genomic differences were limited to a male-specific duplication of a 17-kb unit located 430 kb upstream of Sox9 on an autosome. Hi-C analysis using male spiny rat cells showed the duplicated region has potential chromatin interaction with Sox9. The duplicated unit harbored a 1,262-bp element homologous to mouse enhancer 14 (Enh14), a candidate Sox9 enhancer that is functionally redundant in mice. Transgenic reporter mice showed that the spiny rat Enh14 can function as an embryonic testis enhancer in mice. Embryonic gonads of XX mice in which Enh14 was replaced by the duplicated spiny rat Enh14 showed increased Sox9 expression and decreased Foxl2 expression. We propose that male-specific duplication of this Sox9 enhancer substituted for Sry function, defining a novel Y chromosome in the spiny rat.

Early development of the human embryonic testis

Human gonadal development culminating in testicular differentiation is described through analysis of histologic sections derived from 33-day to 20-week human embryos/fetuses, focusing on early development (4–8 weeks of gestation). Our study updates the comprehensive studies of Felix (1912), van Wagenen and Simpson (1965), and Juric-Lekic et al. (2013), which were published in books and thus are unsearchable via PubMed. Human gonads develop from the germinal ridge, a thickening of coelomic epithelium on the medial side of the urogenital ridge. The bilateral urogenital ridges contain elements of the mesonephric kidney, namely the mesonephric duct, mesonephric tubules, and mesonephric glomeruli. The germinal ridge, into which primordial germ cells migrate, is initially recognized as a thickening of coelomic epithelium on the urogenital ridge late in the 4th week of gestation. Subsequently, in the 5th week of gestation, a dense mesenchyme develops sub-adjacent to the epithelium of the germinal ridge, and together these elements bulge into the coelomic cavity forming bilateral longitudinal ridges attached to the urogenital ridges. During development, primordial cells migrate into the germinal ridge and subsequently into testicular cords that form within the featureless dense mesenchyme of the germinal ridge at 6–8 weeks of gestation. The initial low density of testicular cords seen at 8 weeks remodels into a dense array of testicular cords surrounded by α-actin-positive myoid cells during the second trimester. Human testicular development shares many features with that of mice being derived from 4 elements: coelomic epithelium, sub-adjacent mesenchyme, primordial germ cells, and the mesonephros.

Unraveling mitochondrial piRNAs in mouse embryonic gonadal cells

Although mitochondria are widely studied organelles, the recent interest in the role of mitochondrial small noncoding RNAs (sncRNAs), miRNAs, and more recently, piRNAs, is providing new functional perspectives in germ cell development and differentiation. piRNAs (PIWI-interacting RNAs) are single-stranded sncRNAs of mostly about 20–35 nucleotides, generated from the processing of pre-piRNAs. We leverage next-generation sequencing data obtained from mouse primordial germ cells and somatic cells purified from early-differentiating embryonic ovaries and testis from 11.5 to 13.5 days postcoitum. Using bioinformatic tools, we elucidate (i) the origins of piRNAs as transcribed from mitochondrial DNA fragments inserted in the nucleus or from the mitochondrial genome; (ii) their levels of expression; and (iii) their potential roles, as well as their association with genomic regions encoding other sncRNAs (such as tRNAs and rRNAs) and the mitochondrial regulatory region (D-loop). Finally, our results suggest how nucleo-mitochondrial communication, both anterograde and retrograde signaling, may be mediated by mitochondria-associated piRNAs.

CD34 is Expressed in Endothelial Cells in Embryonic Testes and is Additionally Expressed in Non-Endothelial Cells in Postnatal Mouse Testes.

CD34 is expressed in various cell types in various tissues/organs, and has been regarded as being expressed in progenitors in various differentiation pathways. On the other hand, morphological studies have reported the presence of a special type of interstitial cells, telocytes, which generally express CD34, and have extremely long moniliform prolongations in various tissues/organs in vertebrates. We have recently reported the successful reconstruction of testicular structures by 3-D re-aggregation culture of dissociated prepubertal mouse testicular cells, and the roles of CD34 + cells in the reconstruction. However, it was unknown whether CD34 is expressed in embryonic through adult testes, and if so, in what cell type it is expressed. In order to clarify the expression of CD34 and behavior of CD34 + cells during development of mouse testes, we performed immunohistochemical studies. The results show that CD34 is expressed in two cell types in testes; one is endothelial cells which co-express CD31, VE-cadherin, and integrin β1, but barely express PDGFRα and integrin α4 and α9, throughout development, while the other one is non-endothelial cells in which CD34 expression is initiated after birth, and which co-express PDGFRα and integrin α4, α9, and β1. The latter corresponds to telocytes. The present findings will lead to clarifying the roles of these two types of CD34 + cells in spermatogenesis.

The Expression Dynamics of piRNAs Derived From Male Germline piRNA Clusters and Retrotransposons.

In mammals, germ cells produce a class of small regulatory RNAs called PIWI-interacting RNAs or piRNAs, which are 25–32 nucleotides in length. The profile of testicular piRNAs changes during development. The piRNAs detected in fetal testes at embryonic day 13.5 and later are called fetal piRNAs. The piRNAs detected in testes in a period where germ cells do not yet enter the pachytene stage of meiotic prophase I are called pre-pachytene piRNAs, whereas those in testes at later postnatal days are called pachytene piRNAs. Here, to elucidate the exact expression dynamics of these piRNAs during development, we compared piRNAs present in male germ cells at different stages, which were purified by fluorescence-activated cell sorting, and those in embryonic testes. The analysis identified three distinct groups of piRNA clusters: prospermatogonial, early, and late clusters. piRNA length was largely correlated with the repertoire of PIWI-like proteins in respective germ cells; however, the late piRNA clusters tended to generate longer (PIWIL1-type) piRNAs, whereas the early clusters tended to generate shorter (PIWIL2-type) piRNAs, suggesting a cluster- or sequence-dependent mechanism for loading onto PIWI-like proteins. Retrotransposon-derived piRNAs, particularly evolutionary young retrotransposons, were abundantly produced in prospermatogonia, however, their abundance declined as development proceeded. Thus, in later stages, retrotransposon-derived piRNAs were not enriched with those from evolutionary young elements. The results revealed that, depending on the piRNA clusters from which they are derived, longer PIWIL1-type piRNAs are produced earlier, and shorter PIWIL2-type piRNAs remain in a longer period, than previously thought.

Advances in research of TGF-Β1 in human testis

Transforming growth factor β1 (TGF-β1) is a polypeptide growth factor with various biological activities. Recent studies have shown that this growth factor and they receptors are expressed in human Leydig cells, Sertoli cells, spermatogonia, spermatocytes and sperm cells after birth, and are involved in the endocrine regulation of spermatogenesis. However, there is no report on the expression of TGF-β in human embryonic testis. TGF-β1 is a peptide growth factor widely distributed in germ cells at various levels in the testis, regulating the function of the testes and maintaining the stability of the testis environment by acting on various cells. This review summarizes the research progress of TGF-β1 in human testis.

Generation and mutational analysis of a transgenic mouse model of human SRY.

SRY is the Y-chromosomal gene that determines male sex development in humans and most other mammals. After three decades of study, we still lack a detailed understanding of which domains of the SRY protein are required to engage the pathway of gene activity leading to testis development. Some insight has been gained from the study of genetic variations underlying differences/disorders of sex determination (DSD), but the lack of a system of experimentally generating SRY mutations and studying their consequences in vivo has limited progress in the field. To address this issue, we generated a mouse model carrying a human SRY transgene able to drive testis determination in XX mice. Using CRISPR-Cas9 gene editing, we generated novel genetic modifications in each of SRY's three domains (N-terminal, HMG box, and C-terminal) and performed a detailed analysis of their molecular and cellular effects on embryonic testis development. Our results provide new functional insights unique to human SRYand present a versatile and powerful system in which to functionally analyze variations of SRY including known and novel pathogenic variants found in DSD.

MORC3, a novel MIWI2 association partner, as an epigenetic regulator of piRNA dependent transposon silencing in male germ cells

The PIWI (P-element-induced wimpy testis)-interacting-RNA (piRNA) pathway plays a crucial role in the repression of TE (transposable element) expression via de novo DNA methylation in mouse embryonic male germ cells. Various proteins, including MIWI2 are involved in the process. TE silencing is ensured by piRNA-guided MIWI2 that recruits some effector proteins of the DNA methylation machinery to TE regions. However, the molecular mechanism underlying the methylation is complex and has not been fully elucidated. Here, we identified MORC3 as a novel associating partner of MIWI2 and also a nuclear effector of retrotransposon silencing via piRNA-dependent de novo DNA methylation in embryonic testis. Moreover, we show that MORC3 is important for transcription of piRNA precursors and subsequently affects piRNA production. Thus, we provide the first mechanistic insights into the role of this effector protein in the first stage of piRNA biogenesis in embryonic TE silencing mechanism.

COMPARATIVE STUDY BETWEEN ROTA VIRUSES ISOLATED FROM DIARRHEATIC INFANT BABIES AND NEWLY BORN CALVES

Rota viruses were isolated in vero continuous cell line from infant babies and newly borne calves affected by diarrhea after treatment in every passage with 10 mg/ml of trypsin and adding 0.5 mg/ml of trypsin in the maintenance media. The isolated viruses induced distinctive and progressive type of cytopatheic effect in infected cells and highest titer was 2x10 6. TCID,50/0.1 for bovine viral isolate. The isolated 50/0.1 for human viral isolate and 2x10 » viruses were identified by indirect fluorescent antibody technique by using reference calf rotavirus antisera. Comparative study were conducted on both human and bovine viral isolate including growth in different cell culture, vero cell line was very sensitive to support growth of both viruses than secondary embryonic calf kidney cell culture, lamb testes and primary embryonic chicken fibroblast cell culture. Both viruses induced morphologically similar kind or plaques in vero cell line but plaqes formed by human isolate were larger in size about 1.5 - 2.5 mm in diameter than those of bovine isolate 1.5 - 2 mm. Cross reactive viral antigens were detected between the isolated viruses in indirect fluorescent antibody technique but not in serum neutralization test by using reference bovine rotavirus antisera.

Testicular germ cell tumors arise in the absence of sex-specific differentiation.

In response to signals from the embryonic testis, the germ cell intrinsic factor NANOS2 coordinates a transcriptional program necessary for the differentiation of pluripotent-like primordial germ cells toward a unipotent spermatogonial stem cell fate. Emerging evidence indicates that genetic risk factors contribute to testicular germ cell tumor initiation by disrupting sex-specific differentiation. Here, using the 129.MOLF-Chr19 mouse model of testicular teratomas and a NANOS2 reporter allele, we report that the developmental phenotypes required for tumorigenesis, including failure to enter mitotic arrest, retention of pluripotency and delayed sex-specific differentiation, were exclusive to a subpopulation of germ cells failing to express NANOS2. Single-cell RNA sequencing revealed that embryonic day 15.5 NANOS2-deficient germ cells and embryonal carcinoma cells developed a transcriptional profile enriched for MYC signaling, NODAL signaling and primed pluripotency. Moreover, lineage-tracing experiments demonstrated that embryonal carcinoma cells arose exclusively from germ cells failing to express NANOS2. Our results indicate that NANOS2 is the nexus through which several genetic risk factors influence tumor susceptibility. We propose that, in the absence of sex specification, signals native to the developing testis drive germ cell transformation.

Mouse dead end1 acts with Nanos2 and Nanos3 to regulate testicular teratoma incidence.

Spontaneous testicular teratomas (STTs) derived from primordial germ cells (PGCs) in the mouse embryonic testes predominantly develop in the 129 family inbred strain. Ter (spontaneous mutation) is a single nucleotide polymorphism that generates a premature stop codon of Dead end1 (Dnd1) and increases the incidence of STTs in the 129 genetic background. We previously found that DND1 interacts with NANOS2 or NANOS3 and that these complexes play a vital role in male embryonic germ cells and adult spermatogonia. However, the following are unclear: (a) whether DND1 works with NANOS2 or NANOS3 to regulate teratoma incidence, and (b) whether Ter simply causes Dnd1 loss or produces a short mutant DND1 protein. In the current study, we newly established a conventional Dnd1-knockout mouse line and found that these mice showed phenotypes similar to those of Ter mutant mice in spermatogenesis, oogenesis, and teratoma incidence, with a slight difference in spermiogenesis. In addition, we found that the amount of DND1 in Dnd1+/Ter embryos decreased to half of that in wild-type embryos, while the expression of the short mutant DND1 was not detected. We also found that double mutants for Dnd1 and Nanos2 or Nanos3 showed synergistic increase in the incidence of STTs. These data support the idea that Ter causes Dnd1 loss, leading to an increase in STT incidence, and that DND1 acts with NANOS2 and NANOS3 to regulate the development of teratoma from PGCs in the 129 genetic background. Thus, our results clarify the role of Dnd1 in the development of STTs and provide a novel insight into its pathogenic mechanism.

GPAT2 is required for piRNA biogenesis, transposon silencing, and maintenance of spermatogonia in mice†.

PIWI-interacting RNAs (piRNAs), a subclass of germ cell-specific noncoding small RNAs, are essential for de novo DNA methylation of retrotransposon genes in embryonic testes. PIWIL2/MILI, one of three mouse PIWI family members, is indispensable for piRNA production, DNA methylation of retrotransposons presumably via piRNA, and normal spermatogenesis. In vitro analysis using germline stem cells (GS cells) revealed that glycerol-3-phosphate acyltransferase 2 (GPAT2), which is a mitochondrial outer membrane protein involved in generation of lysophosphatidic acid (LPA) and highly expressed in testes, plays important roles in spermatogenesis. Namely, GPAT2 binds to PIWIL2 and is closely involved in the biogenesis of piRNAs; this process is independent of its enzymatic activity on LPA. However, GS cells recapitulate only a limited phase of spermatogenesis and the biological functions of GPAT2 remain largely unknown. In this study, we generated GPAT2-deficient mice and conducted comprehensive analyses. The deficient mice showed defective piRNA production and subsequent de-silencing of IAP and Line-1 retrotransposons in fetal testes. In addition, apoptosis of pachytene spermatocytes was observed. These abnormalities were all common to the phenotype of PIWIL2-deficient mice, in which piRNA production was impaired. GPAT2-deficient mice exhibited apoptosis in spermatogonia at the neonatal stage, which was not observed in PIWIL2-deficient mice. These data show that GPAT2 plays a critical role in preventing apoptosis in spermatogonia.

The Sertoli cell marker FOXD1 regulates testis development and function in the chicken.

FOXD1, one of the transcription factors of the FOX family, has been shown to be important for mammalian reproduction but little is known about its function in avian species. In the present study, we identified the expression pattern and location of FOXD1 in chicken tissues and testis by performing quantitative polymerase chain reaction, immunohistochemistry and immunofluorescence, and further investigated the regulatory relationship of FOXD1 with genes involved in testis development by RNA interference. Our results showed that FOXD1 is confirmed to be significantly male-biased expressed in the brain, kidney and testis of adults as well as in embryonic gonads, and it is localised in the testicular Sertoli cell in chicken, consistent with its localisation in mammals. After knock-down of FOXD1 in chicken Sertoli cells, the expression of anti-Müllerian hormone (AMH), sex-determining region Y-box 9 (SOX9) and PKA regulatory subunits type I α (RIα) was significantly downregulated, expression of androgen receptor (AR) was notably increased whereas double-sex and MAB-3-related transcription factor 1 (DMRT1) showed no obvious change in expression. These results suggest that FOXD1 is an essential marker for Sertoli cells upstream of SOX9 expression and a potential regulator of embryonic testis differentiation and development and of normal testis function in the chicken.

Tumor Suppressor PTEN Regulates Negatively Sertoli Cell Proliferation, Testis Size, and Sperm Production In Vivo.

The IGFs are the major intratesticular factors regulating immature Sertoli cell proliferation and are, therefore, critical to establish the magnitude of sperm production. However, the intratesticular source of IGF production and the downstream signaling pathway mediating IGF-dependent Sertoli cell proliferation remain unclear. Single-cell RNA sequencing on mouse embryonic testis revealed a robust expression of Igf1 and Igf2 in interstitial steroidogenic progenitors, suggesting that IGFs exert paracrine actions on immature Sertoli cells. To elucidate the intracellular signaling mechanism that underlies the proliferative effects of IGFs on immature Sertoli cells, we have generated mice with Sertoli cell-specific deletion of the Pten gene, a negative regulator of the phosphatidylinositol-3 kinase (PI3K)/AKT pathway, alone or together with the insulin receptor (Insr) and the IGF1 receptor (Igf1r). Although ablation of Pten appears dispensable for Sertoli cell proliferation and spermatogenesis, inactivation of Pten in the absence of Insr and Igf1r rescued the Sertoli cell proliferation rate during late fetal development, testis size, and sperm production. Overall, these findings suggest that IGFs secreted by interstitial progenitor cells act in a paracrine fashion to promote the proliferation of immature Sertoli cells through the IGF/PTEN/PI3K pathway.

Derivation of pluripotent stem cells from nascent undifferentiated teratoma

Teratomas are tumors consisting of components of the three germ layers that differentiate from pluripotent stem cells derived from germ cells. In the normal mouse testis, teratomas rarely form, but a deficiency in Dead-end1 (Dnd1) in mice with a 129/Sv genetic background greatly enhances teratoma formation. Thus, DND1 is crucial for suppression of teratoma development from germ cells. In the Dnd1 mutant testis, nascent teratoma cells emerge at E15.5. To understand the nature of early teratoma cells, we established cell lines in the presence of serum and leukemia inhibitory factor (LIF) from teratoma-forming cells in neonatal Dnd1 mutant testis. These cells, which we designated cultured Dnd1 mutant germ cells (CDGCs), were morphologically similar to embryonic stem cells (ESCs) and could be maintained in the naïve pluripotent condition. In addition, the cells expressed pluripotency genes including Oct4, Nanog, and Sox2; differentiated into cells of the three germ layers in culture; and contributed to chimeric mice. The expression levels of pluripotency genes and global transcriptomes in CDGCs as well as these cells’ adaption to culture conditions for primed pluripotency suggested that their pluripotent status is intermediate between naïve and primed pluripotency. In addition, the teratoma-forming cells in the neonatal testis from which CDGCs were derived also showed gene expression profiles intermediate between naïve and primed pluripotency. The results suggested that germ cells in embryonic testes of Dnd1 mutants acquire the intermediate pluripotent status during the course of conversion into teratoma cells.

Transgenerational impaired spermatogenesis with sperm H19 and Gtl2 hypomethylation induced by the endocrine disruptor p,p’-DDE

The hypothesis of fetal origins unraveled that environmental exposures in early life could alter epigenetic modifications in the male germ-line, increasing the susceptibility to adult-onset diseases for generations. In our previous study, p,p’-dichlorodiphenoxydichloroethylene (p,p’-DDE), a representative endocrine disruptor, was found to induce transgenerational impaired male fertility with sperm Igf2 hypomethylation. However, whether other imprinted genes involved remains uncovered. In the present study, we administered p,p’-DDE (100 mg/kg body weight) to pregnant Sprague-Dawley rats from gestational day (GD) 8 to GD15. Male F1 offspring were mated with the females from the same group to obtain F2 progeny. F3 progeny was obtained by inter-crossing the control and exposed F2 generation. Gestational exposure to p,p’-DDE impaired the testis histology and decreased the sperm fertility with H19 and Gtl2 hypomethylation in F1 offspring. Interestingly, this impaired spermatogenesis and DNA hypomethylation were maintained through F2 and F3 generations. DNA methyltransferase DNMT1 and 3a were significantly decreased in the embryonic testis of p,p’-DDE-treated F1 and F2 generation and returned to control levels by the F3 generation. p,p’-DDE induced a transgenerational impaired spermatogenesis with altered epigenetic modification, posing significant implications for disease etiology.

Embryonic lethality and defective male germ cell development in mice lacking UTF1

The germ cell lineage is specified early in embryogenesis and undergoes complex developmental programs to generate gametes. Here, we conducted genetic studies to investigate the role of Utf1 (Undifferentiated embryonic cell transcription factor 1) in mouse germ cell development. Utf1 is expressed in pluripotent embryonic stem (ES) cells and regulates ES cell differentiation. In a proteomics screen, we identified UTF1 among 38 proteins including DNMT3L and DND1 that associate with chromatin in embryonic testes. We find that UTF1 is expressed in embryonic and newborn gonocytes and in a subset of early spermatogonia. Ubiquitous inactivation of Utf1 causes embryonic lethality in mice with a hybrid genetic background. Male mice with a germline-specific deletion of Utf1 resulting from Prdm1-Cre mediated recombination are born with significantly fewer gonocytes and exhibit defective spermatogenesis and reduced sperm count as young adults. These defects are ameliorated in older animals. These results demonstrate that UTF1 is required for embryonic development and regulates male germ cell development.