Biology subjects

Gonen, N.

Publications and source records attributed to Gonen, N..

5 recordsLinked to original sources

bioRxiv2026

Male fertility is independent of Enh13 control of Sox9 testicular expression

Testis development relies on precise temporal control of Sox9 expression, which is rapidly activated to initiate testis development and subsequently maintained to preserve Sertoli cell identity and male fertility. The distal enhancer, Enh13, is essential for Sox9 upregulation as its constitutive deletion results in complete XY male-to-female sex reversal. Enh13s requirement across distinct developmental stages remains unexplored. We show that early gonadal deletion of Enh13 fully recapitulates XY sex reversal, demonstrating that Enh13 activity is strictly required to initiate the male pathway. In contrast, Sertoli cell-specific deletion of Enh13 after sex determination has no effect on Sox9 expression, testis architecture, or male fertility, revealing that Enh13 is dispensable for Sox9 maintenance in the differentiated testis. Chromatin accessibility identifies candidate enhancers gaining activity after sex determination, suggesting a regulatory handoff within the Sox9 locus. Finally, we show that duplicating Enh13 in the endogenous mouse locus fails to recapitulate the human XX sex reversal, suggesting that it may not be solely Enh13 sequence duplication that led to XX sex reversal in humans. To the best of our knowledge, this study represents the first in vivo conditional knockout of a developmental enhancer, allowing for temporally controlled interrogation of regulatory logic.

developmental biology↗

bioRxiv2024

Divergent regulatory element programs steer sex-specific supporting cell differentiation along mouse gonadal development

Gonadal sex determination relies on tipping a delicate balance involving the activation and repression of several transcription factors and signalling pathways. This is likely mediated by numerous non-coding regulatory elements that shape sex-specific transcriptomic programs. To explore the dynamics of these in detail, we performed paired time-series of transcriptomic and chromatin accessibility assays on pre-granulosa and Sertoli cells throughout their development in the embryo, making use of new and existing mouse reporter lines. Regulatory elements were associated with their putative target genes by linkage analysis, and this was complemented and verified experimentally using promoter capture Hi-C. We identified the transcription factor motifs enriched in these regulatory elements along with their occupancy, pinpointing LHX9/EMX2 as potentially critical regulators of ovarian development. Variations in the DNA sequence of these regulatory elements are likely to be responsible for many of the unexplained cases of individuals with Differences of Sex Development. TeaserMultiomics analysis revealed the regulatory elements and transcription factors responsible for gonadal sex determination.

developmental biology↗

bioRxiv2023

Transposable elements acquire time- and sex-specific transcriptional and epigenetic signatures along mouse fetal gonad development

Gonadal sex determination in mice is a complex and dynamic process, crucial for the development of functional reproductive organs. The expression of genes involved in this process is regulated by a variety of genetic and epigenetic mechanisms. Recently, there has been increasing evidence that transposable elements (TEs), which are a class of mobile genetic elements, play a significant role in regulating gene expression during embryogenesis and organ development. In this study, we aimed to investigate the involvement of TEs in the regulation of gene expression during mouse embryonic gonadal development. Through bioinformatic analysis, we aimed to identify and characterize specific TEs acting as regulatory elements for sex-specific genes, as well as their potential mechanisms of regulation. We identified TE loci expressed in a time- and sex-specific manner along fetal gonad development that correlate positively and negatively with nearby gene expression, suggesting that their expression is integrated to the gonadal regulatory network. Moreover, chromatin accessibility and histone post-transcriptional modification analyses in the differentiating supporting cells revealed that TEs are acquiring sex-specific signature for promoter-, enhancer-, and silencer-like elements with some of them being proximal to critical sex determining genes. Altogether, our study introduces TEs as new potential players of the gene regulatory network controlling gonadal development in mammals.

developmental biology↗

bioRxiv2021

In-vitro cellular reprogramming to model gonad development and its disorders

During embryonic development, mutually antagonistic signaling cascades determine the fate of the bipotential gonad towards a testicular or ovarian identity. Errors in this process result in human Disorders of Sex Development (DSDs), where there is discordance between chromosomal, gonadal, and anatomical sex. The absence of an appropriate, accessible in-vitro system is a major obstacle in understanding mechanisms of sex-determination/DSDs. Here, we describe protocols for differentiation of mouse and human pluripotent cells towards gonadal progenitors. Transcriptomic analysis reveals that the in-vitro-derived murine gonadal cells are equivalent to E11.5 in-vivo progenitors. Using similar conditions, Sertoli-like cells derived from 46,XY human induced pluripotent stem cells (hiPSCs) exhibit sustained expression of testis-specific genes, secrete AMH, migrate and form tubular structures. The cells derived from a 46,XY DSD female hiPSCs, carrying a NR5A1 variant, show aberrant gene expression and absence of tubule formation. CRISPR/Cas9-mediated correction of the variant rescued the phenotype. This is a robust tool to understand mechanisms of sex-determination and model DSDs.

developmental biology↗

bioRxiv2018

Widespread PERK-dependent repression of ER targets in response to ER stress

The UPR (Unfolded Protein Response) is a well-orchestrated response to ER protein folding and processing overload, integrating both transcriptional and translational outputs. Its three arms in mammalian cells, the PERK translational response arm, together with the ATF6 and IRE1-XBP1-mediated transcriptional arms, have been thoroughly investigated.\n\nUsing ribosome footprint profiling, we performed a deep characterization of gene expression programs involved in the early and late ER stress responses, within WT or PERK -/- Mouse Embryonic Fibroblasts (MEFs). We found that both repression and activation gene expression programs, affecting hundreds of genes, are significantly hampered in the absence of PERK. Specifically, PERK -/- cells do not show global translational inhibition, nor do they specifically activate early gene expression programs upon short exposure to ER stress. Furthermore, while PERK -/- cells do activate/repress late ER-stress response genes, the response is substantially weaker. Importantly, we highlight a widespread PERK-dependent repression gene expression program, consisting of ER targeted proteins, including transmembrane proteins, glycoproteins, and proteins with disulfide bonds. This phenomenon occurs in various different cell types, and has a major translational regulatory component. Moreover, we revealed a novel interplay between PERK and the XBP1-ATF6 arms of the UPR, whereby PERK attenuates the expression of a specific subset of XBP1-ATF6 targets, further illuminating the complexity of the integrated ER stress response.

molecular biology↗