Single-cell transcriptome landscape of ovarian cells during primordial follicle assembly in mice

Abstract

Primordial follicle assembly in the mouse occurs during perinatal ages and largely determines the ovarian reserve that will be available to support the reproductive life span. The development of primordial follicles is controlled by a complex network of interactions between oocytes and ovarian somatic cells that remain poorly understood. In the present research, using single-cell RNA sequencing performed over a time series on murine ovaries, coupled with several bioinformatics analyses, the complete dynamic genetic programs of germ and granulosa cells from E16.5 to postnatal day (PD) 3 were reported. Along with confirming the previously reported expression of genes by germ cells and granulosa cells, our analyses identified 5 distinct cell clusters associated with germ cells and 6 with granulosa cells. Consequently, several new genes expressed at significant levels at each investigated stage were assigned. By building single-cell pseudotemporal trajectories, 3 states and 1 branch point of fate transition for the germ cells were revealed, as well as for the granulosa cells. Moreover, Gene Ontology (GO) term enrichment enabled identification of the biological process most represented in germ cells and granulosa cells or common to both cell types at each specific stage, and the interactions of germ cells and granulosa cells basing on known and novel pathway were presented. Finally, by using single-cell regulatory network inference and clustering (SCENIC) algorithm, we were able to establish a network of regulons that can be postulated as likely candidates for sustaining germ cell-specific transcription programs throughout the period of investigation. Above all, this study provides the whole transcriptome landscape of ovarian cells and unearths new insights during primordial follicle assembly in mice.

Germ–Somatic Cell Interactions Are Involved in Establishing the Follicle Reserve in Mammals

“…Deleting both factors in mice caused defects in both oocyte and granulosa cells during primordial follicle assembly (Fu et al, 2018). These results show that communication between all the cells involved is essential for proper germ cell cluster breakdown and PF assembly (Fu et al, 2018;Wang et al, 2020).…”

Establishing and maintaining fertility: the importance of cell cycle arrest

“…Using single-cell sequencing, individual cells are clustered together into related populations based on transcriptomic data. In the ovary, the pregranulosa cell lineages and subclusters have been reconstructed using single-cell data sets (Niu and Spradling 2020;Wang et al 2020). Comprehensive data sets covering the stages of germ cell nest breakdown, primordial follicle formation, and primordial follicle activation were generated using single-cell sequencing on whole ovaries from E18.5, PND1, and PND5 (Niu and Spradling 2020).…”

“…Importantly, in the Niu and Spradling (2020) data set, all three Cip/Kip family members Cdkn1a, Cdkn1b, and Cdkn1c are all expressed in both pregranulosa populations, whether the cells are of a bipotential cell origin or an epithelial cell origin. A subsequent study from Wang et al (2020) performed a similar study at E16.5, PND0, and PND3 to better understand the dynamics of primordial follicle assembly. This study confirmed the presence of multiple populations of pregranulosa cells and went one step further to examine the transcriptional networks that are involved in the interactions between oocytes and pregranulosa cells (Wang et al 2020).…”

“…A subsequent study from Wang et al (2020) performed a similar study at E16.5, PND0, and PND3 to better understand the dynamics of primordial follicle assembly. This study confirmed the presence of multiple populations of pregranulosa cells and went one step further to examine the transcriptional networks that are involved in the interactions between oocytes and pregranulosa cells (Wang et al 2020). In agreement with the Niu and Spradling (2020) study, Wang et al (2020) comment on the expression of Cdkn1a and Cdkn1b in the pregranulosa cells and hypothesize an interaction with the FOXO pathways in oocytes (Wang et al 2020).…”

Two alternative methods for the retrieval of somatic cell populations from the mouse ovary

“…1 ) ( Fu et al , 2018 ; Lv et al , 2019 ; Stevant and Nef, 2019 ). To support these numerous developmental pathways, there is heterogeneity within the granulosa cell population ( Rastetter et al , 2014 ; Niu and Spradling, 2020 ; Wang et al , 2020 ). For example, two populations of primordial follicles are formed within the ovary, distinguished by their location in the ovary and the expression of either Forkhead box L2 (FOXL2) or Leucine Rich Repeat Containing G Protein-Coupled Receptor 5 (LGR5) in the granulosa cells ( Rastetter et al , 2014 ; Niu and Spradling, 2020 ).…”