Biology subjects

Estermann, M. A.

Publications and source records attributed to Estermann, M. A..

9 recordsLinked to original sources

bioRxiv2025

Glycogen metabolism in mouse embryonic Sertoli cells sustains the germ line through the lactate shuttle

Metabolites are key regulators of cell fate decisions, epigenetic remodeling, and lineage commitment. While genetic pathways governing testis differentiation are well studied, the role of metabolism remains poorly understood. In this study, we investigated a transient, male-specific accumulation of glycogen in supporting (Sertoli) cells of embryonic testis in mice. Although glycogen metabolism was dispensable for Sertoli cell differentiation, its disruption led to reduced lactate production and impaired PGCs ability to colonize the newly forming testis cords. Inhibiting lactate transport further revealed a critical metabolic coupling between Sertoli and germ cells during early testis development. Surprisingly, external lactate or glucose supplementation failed to rescue the germ cell phenotype. These findings suggest that glycogen accumulation supports a critical developmental window in which both Sertoli and germ cells are metabolically constrained and unable to rely on external carbon sources. This highlights a sensitive period during testicular development where nutrient scarcity could have long-term consequences for fertility.

developmental biology↗

bioRxiv2025

Calcium signals shape metabolic control of H3K27ac and H3K18la to regulate EGA

The use of assisted reproductive technologies (ART) has enabled the birth of over 9 million babies; but it is associated with increased risks of negative metabolic outcomes in offspring. Yet, the underlying mechanism remains unknown. Calcium (Ca2+) signals, which initiate embryo development at fertilization, are frequently disrupted in human ART. In mice, abnormal Ca2+ signals at fertilization impair embryo development and adult offspring metabolism. Changes in intracellular Ca2+ drive mitochondrial activity and production of metabolites used by the epigenetic machinery. For example, acetyl-CoA (derived mainly from pyruvate) and lactyl-CoA (derived from lactate) are used for writing H3K27ac and H3K18la marks that orchestrate initiation of development. Using both a genetic mouse model and treatment with ionomycin to raise intracellular Ca2+ of wild-type fertilized eggs, we found that excess Ca2+ at fertilization changes metabolic substrate availability, causing epigenetic changes that impact embryo development and offspring health. Specifically, increased Ca2+ exposure at fertilization led to increased H3K27ac levels and decreased H3K18la levels at the 1-cell (1C) stage, that persisted until the 2-cell (2C) stage. Ultralow input CUT&Tag revealed significant differences in H3K27ac and H3K18la genomic profiles between control and ionomycin groups. In addition, increased Ca2+ exposure resulted in a marked reduction in global transcription at the 1C stage that persisted through the 2C stage due to diminished activity of RNA polymerase I. Excess Ca2+ following fertilization increased pyruvate dehydrogenase activity (enzyme that converts pyruvate to acetyl-CoA) and decreased total lactate levels. Provision of exogenous lactyl-CoA before ionomycin treatment restored H3K18la levels at the 1C and 2C stages and rescued global transcription to control levels. Our findings demonstrate conclusively that Ca2+ dynamics drive metabolic regulation of epigenetic reprogramming at fertilization and alter EGA.

developmental biology↗

bioRxiv2025

Single-nucleus multiomics of murine gonadal cells reveals transcriptional regulatory network underlying supporting lineage differentiation

Sex determination of mammalian gonads hinges upon sex-specific differentiation of gonadal supporting cells: Sertoli cells in the testis and granulosa cells in the ovary. To gain insights into how supporting cells acquire their identities, we performed joint single-nucleus transcriptomics and chromatin accessibility assays on murine gonadal cells across sex determination. By contrasting sex-specific gene expression and corresponding chromatin accessibility among progenitor and differentiated cells, we discovered that sex-specific chromatin regions in supporting cells are established shortly after sex determination, accompanied by the acquisition of active histone marks. The presence of potential transcription factor binding motifs in the open chromatin regions revealed regulatory networks underlying ovary-enriched factors LEF1 and MSX1, which promote granulosa fate by inducing granulosa-specific genes such as Foxl2 and Fst. Our results not only identify the gene regulatory framework underlying supporting cell sex differentiation, but also provide invaluable resources for the field. TeaserSex-specific changes in chromatin and gene expression underly the divergent developmental fates of the bipotential gonadal primordium.

developmental biology↗

bioRxiv2025

Generation of a Nr2f2-driven inducible Cre mouse to target interstitial cells in the reproductive system

The orphan nuclear receptor Nr2f2, also known as COUP-TFII, play important roles in development and function of multiple organs, including the reproductive system. NR2F2 is expressed in the interstitial cells of both embryonic and adult testes, ovaries, and reproductive tracts. Taking advantage of such unique expression pattern, we have developed a tamoxifen inducible Cre model, Nr2f2-iCreERT2, which specifically and efficiently targets interstitial cells in both male and female reproductive organs across embryonic and adult stages. This model offers a powerful tool for gene knockout studies specifically in the interstitial compartment, without affecting the supporting or germ cells. Additionally, it enables interstitial cell lineage tracing, facilitating the assessment of the interstitials contribution to non-interstitial cell types during development and differentiation.

developmental biology↗

bioRxiv2024

NR2F2 regulation of interstitial to fetal Leydig cell differentiation in the testis: insights into differences of sex development

Testicular fetal Leydig cells are a specialized cell type responsible for embryo masculinization. Fetal Leydig cells produce androgens, that induce the differentiation of male reproductive system and sexual characteristics. Deficiencies in Leydig cell differentiation leads to various disorders of sex development and male reproductive defects such as ambiguous genitalia, hypospadias, cryptorchidism, and infertility. Fetal Leydig cells are thought to originate from proliferating progenitor cells in the testis interstitium, marked by genes like Arx, Pdgfra, Tcf21 and Wnt5a. However, the precise mechanisms governing the transition from interstitial cells to fetal Leydig cells remain elusive. Through integrated approaches involving mouse models and single-nucleus multiomic analyses, we discovered that fetal Leydig cells originate from a Nr2f2-positive non-steroidogenic interstitial cell population. Embryonic deletion of Nr2f2 in mouse testes resulted in disorders of sex development, including dysgenic testes, Leydig cell hypoplasia, cryptorchidism, and hypospadias. We found that NR2F2 promotes the progenitor cell fate while suppresses Leydig cell differentiation by directly and indirectly controlling a cohort of transcription factors and downstream genes. Bioinformatic analyses of single-nucleus ATAC-seq and NR2F2 ChIP-seq data revealed putative transcription factors co-regulating the process of interstitial to Leydig cell differentiation. Collectively, our findings not only highlight the critical role of Nr2f2 in orchestrating the transition from interstitial cells to fetal Leydig cells, but also provide molecular insight into the disorders of sex development as a result of Nr2f2 mutations.

developmental biology↗

bioRxiv2024

Single-nucleus multiomics reveals the gene-regulatory networks underlying sex determination of murine primordial germ cells

Accurate specification of female and male germ cells during embryonic development is critical for sexual reproduction. Primordial germ cells (PGCs) are the bipotential precursors of mature gametes that commit to an oogenic or spermatogenic fate in response to sex-determining cues from the fetal gonad. The critical processes required for PGCs to integrate and respond to signals from the somatic environment in gonads are not understood. In this study, we developed the first single-nucleus multiomics map of chromatin accessibility and gene expression during murine PGC development in both XX and XY embryos. Profiling of cell-type specific transcriptomes and regions of open chromatin from the same cell captured the molecular signatures and gene networks underlying PGC sex determination. Joint RNA and ATAC data for single PGCs resolved previously unreported PGC subpopulations and cataloged a multimodal reference atlas of differentiating PGC clusters. We discovered that regulatory element accessibility precedes gene expression during PGC development, suggesting that changes in chromatin accessibility may prime PGC lineage commitment prior to differentiation. Similarly, we found that sexual dimorphism in chromatin accessibility and gene expression increased temporally in PGCs. Combining single-nucleus sequencing data, we computationally mapped the cohort of transcription factors that regulate the expression of sexually dimorphic genes in PGCs. For example, the gene regulatory networks of XX PGCs are enriched for the transcription factors, TFAP2c, TCFL5, GATA2, MGA, NR6A1, TBX4, and ZFX. Sex-specific enrichment of the forkhead-box and POU6 families of transcription factors was also observed in XY PGCs. Finally, we determined the temporal expression patterns of WNT, BMP, and RA signaling during PGC sex determination, and our discovery analyses identified potentially new cell communication pathways between supporting cells and PGCs. Our results illustrate the diversity of factors involved in programming PGCs towards a sex-specific fate.

developmental biology↗

bioRxiv2022

Fadrozole-mediated sex reversal in the embryonic chicken gonad involves a PAX2 positive undifferentiated supporting cell state

Gonadal sex differentiation among vertebrates involves divergent fates of a common groups of progenitor cells present in both presumptive ovaries and testes. The first cell type to differentiate gives rise to pre-Sertoli cells in the testis, and pre-follicular cells in the ovary. These cells derive form a common lineage of so-called "supporting cells". In birds and other egg-laying vertebrates, locally synthesised estrogen has a central role in ovarian development and influence the fate of these supporting cells. Manipulation of estrogen levels during embryonic development induces gonadal sex reversal, providing an experimental setting to evaluate the process of gonadal sex differentiation. Recently, we identified PAX2 as a novel marker of the undifferentiated supporting cell lineage in the chicken embryo, expressed in both sexes prior to overt gonadal sex differentiation. PAX2 expression is downregulated at the onset of gonadal sex differentiation in both males and females. The analysis of this undifferentiated supporting cell marker, together with Sertoli (male) and pre-granulosa (female) will enhance our understanding of supporting cell differentiation. Here we characterized the supporting cells differentiation process and identified undifferentiated supporting cells in estrogen-mediated sex reversal experiments. Female embryos treated with the aromatase inhibitor fadrozole developed ovotestis, containing pre-granulosa cells, Sertoli cells and PAX2 positive undifferentiated supporting cells. In contrast, male embryos treated with 17{beta}-estradiol showed no PAX2+ undifferentiated gonadal supporting cells. Fadrozole time-course as well as multiple dose analysis suggests that supporting cell transdifferentiation involves a dedifferentiation event into a PAX2+ undifferentiated supporting cell state, followed by a redifferentiation towards the opposite sex lineage.

developmental biology↗

bioRxiv2022

DMRT1 regulation of TOX3 modulates expansion of the gonadal steroidogenic cell lineage

Vertebrate gonads comprise three primary cell types, germ cells, steroidogenic cells, and supporting cells. The latter are the first cell type to differentiate in the embryonic gonad and direct the formation of other somatic lineages. During gonadal sex determination, the supporting cell lineage differentiates into Sertoli cells in males and pre-granulosa cells in females. In the chicken embryo, the molecular trigger for Sertoli cell differentiation is the Z-linked gene DMRT1. Recently, single cell RNA-seq data indicate that that chicken steroidogenic cells, derive from differentiated supporting cells. This differentiation process is achieved by a sequential upregulation of steroidogenic genes and down-regulation of supporting cell markers. The exact mechanism regulating this differentiation process remains unknown. We identified the gene TOX3 as a novel transcription factor expressed in embryonic Sertoli cells of the chicken testis. TOX3 knockdown in males resulted in increased CYP17A1 positive Leydig cells. TOX3 over-expression in male and female gonads resulted in a significant decline in CYP17A1 positive steroidogenic cells. TOX3 expression is negatively regulated by estrogens in vivo, but not induced during masculinization induced by estrogen inhibition. In ovo knock-down of the testis determinant, DMRT1, in male gonads resulted in a down-regulation of TOX3 expression. Conversely, DMRT1 over-expression caused an increase in TOX3 expression. Taken together, this data indicates that DMRT1 regulation of TOX3 modulates expansion of the steroidogenic lineage, either directly, via cell lineage allocation, or indirectly via signaling from the supporting to steroidogenic cell populations.

developmental biology↗

bioRxiv2021

TGIF1 is required for chicken ovarian cortical development and generation of the juxtacortical medulla

During early embryogenesis in amniotic vertebrates, the gonads differentiate into either ovaries or testes. The first cell lineage to differentiate gives rise to the supporting cells; Sertoli cells in males and pre-granulosa cells in females. These key cell types direct the differentiation of the other cell types in the gonad, including steroidogenic cells. The gonadal surface epithelium and the interstitial cell populations are less well studied, and little is known about their sexual differentiation programs. Here, we show the requirement of the transcription factor gene TGIF1 for ovarian development in the chicken embryo. TGIF1 is expressed in the two principal ovarian somatic cell populations, the cortex and the pre-granulosa cells of the medulla. TGIF1 expression is associated with an ovarian phenotype in sex reversal experiments. In addition, targeted over-expression and gene knockdown experiments indicate that TGIF1 is required for proper ovarian cortical formation. TGIF1 is identified as the first known regulator of juxtacortical medulla formation. These findings provide new insights into chicken ovarian differentiation and development, specifically in the process of cortical and juxtacortical medulla formation, a poorly understood area. SUMMARY STATEMENTThe transcription factor TGIF1 is required for proper ovarian sex differentiation in chicken embryos, regulating development of the cortical and juxtacortical medulla, independently of the supporting cell sex lineage.

developmental biology↗