A single-cell atlas of the cycling murine ovary

Morris, Mary E.; Meinsohn, Marie-Charlotte; Chauvin, Maëva; Saatcioglu, Hatice D.; Kashiwagi, A; Sicher, Natalie A; Nguyen, Ngoc Minh; Yuan, Selena; Stavely, Rhian; Hyun, Minsuk; Donahoe, Patricia K.; Sabatini, Bernardo L.; Pépin, David

doi:10.7554/elife.77239

Cited by 38 publications

(76 citation statements)

References 84 publications

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“…This valuable genetic tool allowed us to generate age-matched transcriptome and chromatin landscape datasets to integrate with our ChIP-SICAP data on the genomic binding sites of FOXL2 and be able to draw conclusions as to which genes may be repressed or activated by this factor. FOXL2 is also expressed in theca cells [59], therefore, we first assessed the expression values of known theca and granulosa cell markers [60]. We found an enrichment of granulosa over theca markers, confirming the suitability of this dataset for the exploration of granulosa-specific gene regulatory networks.…”

Section: Discussionsupporting

confidence: 57%

“…Granulosa cells and the steroidogenic cell lineage including theca cells both express FOXL2 [59]. We therefore checked expression of proven markers [60] expressed exclusively in either theca or granulosa cells (Figure 4 G). This analysis indicated enrichment of granulosa cells over theca cells in cells isolated using the Foxl2 EGFP transgene.…”

Section: Identification Of Foxl2 On-chromatin Protein Interactorsmentioning

confidence: 99%

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FOXL2 interaction with different binding partners regulates the dynamics of ovarian development

Migale

Neumann

Mitter

et al. 2023

Preprint

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FOXL2 is a transcription factor essential for female fertility, expressed in somatic cells of the ovary, notably granulosa cells. In the mouse, Foxl2 deletion leads to partial sex reversal postnatally, with mutants developing dysgenic ovaries devoid of oocytes. However, deletion of the gene in 8-week-old females leads to granulosa to Sertoli cell transdifferentiation and gonadal sex reversal. We hypothesise that different outcomes of Foxl2 deletion in embryonic versus adult ovary may derive from a different role played by FOXL2 across ovarian development. Therefore, in this study, we take a multi-omics approach to characterise the dynamics of gene expression and chromatin accessibility changes in purified murine granulosa cells across key developmental stages (E14.5, 1 and 8 weeks). We coupled these analyses with genome wide identification of FOXL2 target genes and on-chromatin interacting partners by ChIP-SICAP to reconstruct the gene regulatory networks underpinned by this essential transcription factor. In the embryonic ovary, FOXL2 interacts with factors important for early stages of gonadal development, such as GATA4 and WT1, whilst postnatally it interacts with factors regulating primordial follicle activation, such as NR5A2, and with factors regulating steroidogenesis including AR and ESR2. Integration of chromatin landscape dynamics with gene expression changes and FOXL2 binding sites analysis revealed that its critical role in ovarian cell fate maintenance goes beyond repression of the Sertoli-specific gene Sox9. Our chromatome analysis revealed also that FOXL2 interacts with several proteins involved in chromatin remodelling, DNA repair, splicing and gene repression. In summary, in this study we identified target genes dynamically regulated by FOXL2 across ovarian development including known and newly identified FOXL2 targets with a role in embryonic ovarian development and folliculogenesis, as well as cofactors that point towards additional roles played by FOXL2 besides transcriptional regulation. This work constitutes a comprehensive resource for exploration of the molecular mechanisms of ovarian development and causes of female infertility.

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Section: Discussionsupporting

confidence: 57%

Section: Identification Of Foxl2 On-chromatin Protein Interactorsmentioning

confidence: 99%

FOXL2 interaction with different binding partners regulates the dynamics of ovarian development

Migale

Neumann

Mitter

et al. 2023

Preprint

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“…The fascinating biology of stem cells that reside in the OSE gets altogether missed during scRNAseq studies (Table 1). Morris et al in 2022 [6] describe OSE as a simple mesothelial cell layer, covering the ovarian stroma, that gets disrupted during ovulation. But OSE is much more dynamic as it houses the stem cells that result in regular GCNs formation, meiosis, neo-oogenesis and primordial follicle assembly during adult life [17].…”

Section: Main Textmentioning

confidence: 99%

VSELs and OSCs together sustain oogenesis in adult ovaries and their dysfunction results in age-related senescence, PCOS, POI and cancer

Bhartiya

Sharma

2023

J Ovarian Res

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Multiple studies using single-cell RNA sequencing (scRNAseq) have failed to detect stem cells in adult ovaries. We have maintained that two populations of ovarian stem cells including pluripotent, very small embryonic-like stem cells (VSELs) and tissue-committed ‘progenitors’ termed ovarian stem cells (OSCs) can easily be detected in Hematoxylin and Eosin-stained ovary surface epithelial (OSE) cells smears prepared from both mice and human ovaries. Most likely the stem cells never get subjected to scRNAseq since they pellet down only by centrifuging cells suspension at 1000 g while cells for scRNAseq were invariably prepared by centrifuging at 200-400 g. A recent article provided further explanation for the failure of scRNAseq to detect ovarian stem cells. Extensive reanalysis of data (generated by scRNAseq) using an advanced software successfully detected OSCs and meiotic markers supporting neo-oogenesis in adult human ovaries. But this article remained critical on the biological relevance of VSELs and their relationship with OSCs. By carefully studying the OSE cells smears (which hold VSELs, OSCs and germ cell nests GCNs), prepared by partial trypsin digestion of intact mice ovaries during different stages of estrus cycle, we have successfully delineated novel functions of VSELs/OSCs in vivo under physiological conditions. VSELs undergo asymmetrical divisions to self-renew and give rise to slightly bigger OSCs which in turn undergo symmetrical divisions and clonal expansion to form GCNs, regular neo-oogenesis and follicle assembly. GCNs have been earlier described in fetal ovaries and during OSE cells culture (from adult ovaries) in response to FSH treatment. Dysfunction of VSELs/OSCs (which express ERα, ERβ, FSHR) due to neonatal exposure to endocrine disruption results in ovarian insufficiency and polycystic ovaries. VSELs have also been implicated in ovarian cancer. Age-related ovarian senescence/menopause is also due to dysfunction and blocked differentiation of VSELs/OSCs. These novel findings in vivo along with abundant in vitro and lineage tracing studies data in published literature provides huge scope for further research, offers novel avenues to manage ovarian pathologies and calls for re-writing of textbooks.

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“…Where the focus is usually on the oocyte, the transcriptomic datasets we provide here highlight the importance and role of somatic cells in follicle activation. Combining this dataset with previously published single cell RNA-sequencing studies of the developing ovary could establish a time course of gene expression across the processes of germ cell nest breakdown, primordial follicle formation and activation, particularly since each study covers different timepoints (6, 7, [23][24][25][26]38). Exciting opportunities also exist to correlate this data with human ovary single cell RNAsequencing studies and those from other species to uncover how well the process is conserved (39).…”

Section: Discussionmentioning

confidence: 99%

“…The emergence of single-cell RNA sequencing (scRNAseq) permits examination of heterogenous populations such as follicles within the mouse ovary (21). Dissociation protocols suitable for scRNAseq on neonatal mouse ovary have been developed by our lab and others (22,23) and used to investigate the mechanisms underpinning germ cell nest breakdown and the first wave of primordial follicle activation in mice (6, [23][24][25][26]. These datasets provide the transcriptomes of oocytes and granulosa cells during ovarian development, with significant overlap of the genes identified in each study.…”

Section: Introductionmentioning

confidence: 99%

Pregranulosa cells upregulate cardiac troponin I Tnni3 upon cell cycle resumption during primordial follicle activation.

Frost

Taylor

Boeing

et al. 2022

Preprint

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Female fertility is dictated by the number of oocyte-containing primordial follicles within the ovary. These follicles are established early in development and their selective activation represents the definitive first step towards oocyte maturation and ovulation. The intrinsic molecular mechanisms that regulate ovarian follicle activation are largely uncharacterised. In this study, we report a single-cell RNA sequencing (scRNAseq) dataset to examine mouse embryonic and neonatal ovaries across the developmental window of primordial follicle formation and activation. We identified five distinct clusters of granulosa cells and bioinformatic analysis revealed a population of pregranulosa cells that appeared to be undergoing follicle activation. This cluster was uniquely differentiated by the expression of Tnni3, Slc18a2, Fam13a and Klf2, all of which are novel to follicle activation. Finally, we examined the transcriptome of a mouse model where all primordial follicles enter folliculogenesis simultaneously (Cdkn1b-/-) and showed that the signature genes of activating pregranulosa cells was observed precociously in Cdkn1b-/- ovaries, further demonstrating that p27kip1 acts as an important repressor of follicle activation. Combined, this data provides insight into how primordial follicle activation is regulated in mammals and could be utilised to identify pathways to target to improve fertility preservation and infertility conditions in the future.

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A single-cell atlas of the cycling murine ovary

Cited by 38 publications

References 84 publications

FOXL2 interaction with different binding partners regulates the dynamics of ovarian development

FOXL2 interaction with different binding partners regulates the dynamics of ovarian development

VSELs and OSCs together sustain oogenesis in adult ovaries and their dysfunction results in age-related senescence, PCOS, POI and cancer

Pregranulosa cells upregulate cardiac troponin I Tnni3 upon cell cycle resumption during primordial follicle activation.

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