Biology subjects

Yao, H. H.-C.

Publications and source records attributed to Yao, H. H.-C..

10 recordsLinked to original sources

bioRxiv2026

Intermittent exposure to high ambient heat during the second half of gestation in mice causes mild alterations of reproductive endpoints in male embryos

Periods of elevated ambient temperature challenge the bodys ability to maintain internal homeostasis, and heat stress poses particular risks during pregnancy. Epidemiological studies associate gestational heat exposure with higher rates of congenital anomalies such as hypospadias, yet the direct link between gestational heat exposure and reproductive anomalies remains unknown. In this study, we examined the effects of intermittent heat exposure on reproductive development in male mouse offspring. Pregnant dams either remained at constant temperature of 22{degrees}C (control) or were exposed to 38{degrees}C for 2 hours daily (experimental) from embryonic day (E)10 to E18, modeling intermittent heat exposure during mid-to-late gestation. Embryos were collected at E18 for analysis. While heat exposure did not affect pregnancy outcomes, including placental development, litter size, sex ratio, or fetal growth, male embryos exhibited significantly reduced anogenital distance and increased hypospadias scores, which are both markers of disrupted androgen signaling. Despite these phenotypic changes, expression of genes involved in androgen synthesis in the fetal testis, as well as gene expression in external genitalia, remained unchanged. Instead, transcriptomic analysis revealed significant alterations in testicular pathways related to RNA splicing and mRNA processing. Together, these findings reveal that maternal heat stress disrupts reproductive development of male offspring, with altered gene regulatory processes being a potential driver.

developmental biology↗

bioRxiv2026

Specification of the interstitial cell lineage in the fetal mouse ovary requires balanced Notch signaling

The ovary contains two major somatic lineages, granulosa cells and interstitial cells, that arise from progenitors within the coelomic epithelium. However, how these two lineages diverge during ovarian development remains unclear. By analyzing joint single-nucleus transcriptomic and chromatin accessibility profiles of murine ovarian cells at the onset of ovary formation, we identified two somatic progenitor populations from the coelomic epithelium distinguished by expression of the nuclear receptors Nr5a1 and Nr2f2. Based on their transcriptomic trajectories, the Nr5a1+ epithelial cells preferentially transitioned toward the granulosa lineage whereas the Nr2f2+ epithelial cells differentiated into mesenchymal populations. This lineage relationship was supported by Nr2f2 lineage tracing experiments that fetal Nr2f2+ progenitors contribute to ovarian interstitial cells postnatally. To define the molecular features underlying this divergence, we performed differential gene expression and chromatin accessibility analyses and found that Nr2f2+ epithelial cells, but not Nr5a1+ cells, were enriched for Notch pathway components and Notch effector motifs. Consistently, lineage tracing of Notch-responsive cells marked Nr2f2+ interstitial cells in postnatal ovaries, whereas ectopic Notch activation in Nr5a1+ cells promoted expansion of the interstitial population accompanied by reduced granulosa cells. By integrating motif analysis with accessible chromatin-gene linkage, we also identified downstream targets regulated by Notch effectors in Nr2f2+ cells, which showed concordant changes upon ectopic Notch activation. These findings demonstrate that somatic cell fate is established early during ovarian development, with active Notch signaling specifying the interstitial lineage and a balanced Notch activity required for proper somatic lineage establishment. Significance StatementProper differentiation of somatic cell types in the fetal ovary lays the foundation for future ovarian function in adulthood. Understanding how each cell type is formed is essential for developing methods to intervene in ovarian diseases caused by cellular dysfunction. Given that common somatic progenitors give rise to both supporting and interstitial lineages, a main unanswered question is how these two lineages diverge apart from each other during ovarian development. By integrating joint single-nucleus transcriptomic and chromatin accessibility assays with lineage tracing, single cell RNA-sequencing, and mouse genetic models, we demonstrate the role of Notch signaling in specifying the interstitial lineage and separating it from the supporting cell fate.

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↗

bioRxiv2023

An extra-genital cell population contributes to urethra closure during mouse penis development

Hypospadias, or incomplete closure of the urethra along the penis, is the second most common birth defect in the United States. We discovered a population of extra- genital mesenchymal cells that are essential for proper penile urethra closure in mouse embryos. This extra-genital population first appeared in the mesenchyme posterior to the hindlimb of the fetus after the onset of penis formation. These extra-genital cells, which transiently express a lineage marker Nr5a1, migrated centrally and colonized the penis bilateral to the urethra epithelium. Removal of the Nr5a1+ extra-genital cells, using a cell-type specific ablation model, resulted in severe hypospadias. The absence of extra-genital cells had the most significant impacts on another mesenchymal cells, the peri-urethra that were immediately adjacent to the Nr5a1+ extra-genital cells. Single cell mRNA sequencing revealed that the extra-genital cells extensively interact with the peri-urethra, particularly through Neuregulin 1, an epidermal Growth Factor (EGF) ligand. Disruption of Neuregulin 1 signaling in the ex-vivo slice culture system led to failure of urethra closure, recapitulating the phenotypes of extra-genital cell ablation. These results demonstrate that the Nr5a1+ extra-genital mesenchymal cells from outside of the fetal penis are indispensable for urethra closure through their interaction with the peri-urethra mesenchymal cells. This discovery provides a new entry point to understand the biology of penis formation and potential causes of hypospadias in humans.

developmental biology↗

bioRxiv2022

Contribution of the Wolffian duct mesenchyme to the formation of the female reproductive tract

The female reproductive tract develops from its embryonic precursor, the Mullerian duct. In close proximity to the Mullerian duct lies the precursor for the male reproductive tract, the Wolffian duct, which is eliminated in the female embryo during sexual differentiation. We discovered that a component of the Wolffian duct, its mesenchyme, is not eliminated after sexual differentiation. Instead, the Wolffian duct mesenchyme underwent changes in transcriptome and chromatin accessibility from male tract to female tract identity, and became a unique mesenchymal population in the female reproductive tract with localization and transcriptome distinct from the mesenchyme derived from the Mullerian duct. Partial ablation of the Wolffian duct mesenchyme stunted the growth of the fetal female reproductive tract. These findings reveal a new fetal origin of mesenchymal tissues for female reproductive tract formation and reshape our understanding of sexual differentiation of reproductive tracts. Significance statementA female embryo initially possesses both the primitive female and male reproductive tracts, also known as the Mullerian and Wolffian duct. During sexual differentiation, the female eliminates the Wolffian duct and maintains the Mullerian duct that eventually differentiates into the female reproductive tract organs. However, in this paper, we show the female embryo retains mesenchymal cells surrounding the Wolffian duct for its reproductive tract formation. When incorporated into the female reproductive tract organs, the Wolffian duct mesenchyme shows unique anatomical localization and transcriptome and plays critical roles in female reproductive tract growth. This discovery provides new insights into female reproductive tract development and advances our understanding of sexual differentiation of reproductive tracts.

developmental biology↗

bioRxiv2021

Progesterone receptor-A isoform interaction with RUNX transcription factors controls chromatin remodelling at promoters during ovulation

Progesterone receptor (PGR) plays diverse roles in reproductive tissues and thus coordinates mammalian fertility. In the ovary, acutely induced PGR is the key determinant of ovulation through transcriptional control of a unique set of genes that culminates in follicle rupture. However, the molecular mechanisms for PGRs specialised function in ovulation is poorly understood. To address this, we assembled a detailed genomic profile of PGR action through combined ATAC-seq, RNA-seq and ChIP-seq analysis in wildtype and isoform-specific PGR null mice. We demonstrated the unique action of PGR-A isoform in the ovary through a transcriptional complex involving physical interaction with RUNX and JUN/FOS transcription factors. The assembly of this unique complex directs targeting of PGR binding to proximal promoter regions and enables chromatin accessibility, leading to ovulatory gene induction. This PGR signalling mechanism is specific to ovulation and provides potential targets for infertility treatments as well as new contraceptives that block ovulation.

molecular biology↗

bioRxiv2020

Increased FOXL2 Expression Alters Uterine Structures and Functions

Transcription factor FOXL2 exhibits an increase in mRNA levels in eutopic endometrial biopsy in endometriosis patients. While FOXL2 is known of regulating sex differentiation and reproductive function, the impact of elevated FOXL2 expression on uterine physiology remains unknown. To answer this question, we generated mice with over expression of FOXL2 (FOXL2OE) in the female reproductive tract by crossing Foxl2LsL/+ with the Pgrcre model. FOXL2OE uterus showed severe morphological abnormality including abnormal epithelial stratification, blunted adenogenesis, increased endometrial fibrosis and disrupted myometrial morphology. In contrast, increasing FOXL2 levels specifically in uterine epithelium by crossing the Foxl2LsL/+ with the Ltficre mice resulted in the eFOXL2OE mice with uterine epithelial stratification but without defects in endometrial fibrosis and adenogenesis, demonstrating a role of the endometrial stroma in the uterine abnormalities of the FOXL2OE mice. Transcriptomic analysis of 12 weeks old Pgrcre and FOXL2OE uterus at diestrus stage showed a positive correlation of FOXL2OE uterine transcriptome with human endometrium of endometriosis patients. Furthermore, we found FOXL2OE mice were sterile. The infertility was caused in part by a disruption of the hypophyseal ovarian axis resulting in an anovulatory phenotype. The FOXL2OE mice failed to show decidual responses during artificial decidualization in ovariectomized mice which demonstrates the uterine contribution to the infertility phenotype. These data supported that aberrantly increased FOXL2 expressions in the female reproductive tract can disrupt ovarian and uterine functions, particularly, may be involved in the progressions of endometriosis.

developmental biology↗

bioRxiv2019

Patterns, Profiles, and Parsimony: dissecting transcriptional signatures from minimal single-cell RNA-seq output with SALSA

Single-cell RNA sequencing (scRNA-seq) technologies have precipitated the development of bioinformatic tools to reconstruct cell lineage specification and differentiation processes with single-cell precision. However, start-up costs and data volumes currently required for statistically reproducible insight remain prohibitively expensive, preventing scRNA-seq technologies from becoming mainstream. Here, we introduce single-cell amalgamation by latent semantic analysis (SALSA), a versatile workflow to address those issues from a data science perspective. SALSA is an integrative and systematic methodology that introduces matrix focusing, a parametric frequentist approach to identify fractions of statistically significant and robust data within single-cell expression matrices. SALSA then transforms the focused matrix into an imputable mix of data-positive and data-missing information, projects it into a latent variable space using generalized linear modelling, and extracts patterns of enrichment. Last, SALSA leverages multivariate analyses, adjusted for rates of library-wise transcript detection and cluster-wise gene representation across latent patterns, to assign individual cells under distinct transcriptional profiles via unsupervised hierarchical clustering. In SALSA, cell type assignment relies exclusively on genes expressed both robustly, relative to sequencing noise, and differentially, among latent patterns, which represent best-candidates for confirmatory validation assays. To benchmark how SALSA performs in experimental settings, we used the publicly available 10X Genomics PBMC 3K dataset, a pre-curated silver standard comprising 2,700 single-cell barcodes from human frozen peripheral blood with transcripts aligned to 16,634 genes. SALSA identified at least 7 distinct transcriptional profiles in PBMC 3K based on <500 differentially expressed Profiler genes determined agnostically, which matched expected frequencies of dominant cell types in peripheral blood. We confirmed that each transcriptional profile inferred by SALSA matched known expression signatures of blood cell types based on surveys of 15 landmark genes and other supplemental markers. SALSA was able to resolve transcriptional profiles from only [~]9% of the total count data accrued, spread across <0.5% of the PBMC 3K expression matrix real estate (16,634 genes x 2,700 cells). In conclusion, SALSA amalgamates scRNA-seq data in favor of reproducible findings. Furthermore, by extracting statistical insight at lower experimental costs and computational workloads than previously reported, SALSA represents an alternative bioinformatics strategy to make single-cell technologies affordable and widespread.

bioinformatics↗