# Characterizing the Human Fetal Perimeiotic 45,X Ovary at Single-Cell Resolution

## TL;DR

This study uses single-cell sequencing to explore why ovaries in Turner syndrome develop poorly, revealing differences in gene activity and cell counts.

## Contribution

The study provides the first single-cell resolution characterization of the human fetal 45,X ovary in Turner syndrome.

## Key findings

- 45,X ovaries have fewer germ cells in all subpopulations compared to 46,XX ovaries.
- Disrupted X-chromosome inactivation and reactivation are observed in 45,X ovaries.
- Genes related to proteostasis, cell cycle, and energy production are underexpressed in 45,X ovaries.

## Abstract

Turner syndrome (TS) is the most common genetic cause of premature (primary) ovarian insufficiency (POI). Human fetal 45,X ovaries demonstrate marked apoptosis by 15 to 20 weeks post conception (wpc), likely partly driven by X-chromosome haploinsufficiency. However, the genomic drivers of ovarian insufficiency in TS remain largely unexplored.

We used single-nuclei sequencing (snRNA-seq) and bulk RNA sequencing (RNA-seq) technologies to profile the transcriptome of ovarian insufficiency in TS.

Using snRNA-seq, we profiled 2 perimeiotic 46,XX and 2 45,X (TS) human fetal ovaries (12-13 wpc). Using bulk RNA-seq, we conducted a time-series analysis of human fetal tissue across 4 developmental time points (19 fetal ovary, 20 fetal testis, 8 fetal control tissue (n = 47 total samples; Carnegie stage 22-16 wpc)).

Germ and somatic cell subpopulations were mostly shared across 46,XX and 45,X ovaries, aside from an oogonia cluster depleted in 45,X ovaries containing genes with functions relating to sex chromosome synapsis. snRNA-seq enabled accurate cell counting across individual cell clusters and revealed that the 45,X ovary has fewer germ cells than the 46,XX ovary in every germ cell subpopulation, confirmed by histopathological analysis. The normal sequence of X-chromosome inactivation and reactivation is disrupted in 45,X ovaries. The 45,X ovary has a globally abnormal transcriptome, with lower expression of genes with proteostasis functions (RSP4X); cell cycle progression (BUB1B); and OXPHOS energy production (COX6C, ATP11C).

We characterize the human fetal perimeiotic 45,X ovary at single-cell resolution and offer insights into the genomic mechanisms of the ovarian insufficiency phenotype in TS.