Parental haplotype-specific single-cell transcriptomics reveal incomplete epigenetic reprogramming in human female germ cells

Abstract

In contrast to mouse, human female germ cells develop asynchronously. Germ cells transition to meiosis, erase genomic imprints, and reactivate the X chromosome. It is unknown if these events all appear asynchronously, and how they relate to each other. Here we combine exome sequencing of human fetal and maternal tissues with single-cell RNA-sequencing of five donors. We reconstruct full parental haplotypes and quantify changes in parental allele-specific expression, genome-wide. First we distinguish primordial germ cells (PGC), pre-meiotic, and meiotic transcriptional stages. Next we demonstrate that germ cells from various stages monoallelically express imprinted genes and confirm this by methylation patterns. Finally, we show that roughly 30% of the PGCs are still reactivating their inactive X chromosome and that this is related to transcriptional stage rather than fetal age. Altogether, we uncover the complexity and cell-to-cell heterogeneity of transcriptional and epigenetic remodeling in female human germ cells.

Specification and epigenetic resetting of the pig germline exhibit conservation with the human lineage

“…showing loss of the H3K27me3 spot at Wk4 (Tang et al, 2015), and biallelic expression of X-linked genes at Wk7-8 PGCs; however data from earlier stages is not available (Sangrithi et al, 2017;Vértesy et al, 2018). Even though it is not currently possible to conclude whether human XCR occurs as early as shown in pPGCs, our evidence of XCR in pig premigratory pPGCs contrasts with observations in mouse PGCs, where there is limited loss of H3K27me3 (<10%) and Xist (<15%) expression in pre-migratory PGCs; the increase in X:A ratio is first detected in E11.…”

Applying Single-Cell Analysis to Gonadogenesis and DSDs (Disorders/Differences of Sex Development)

“…In addition, Leydig or steroidogenic cells, macrophages and T cells were also detected [ 119 ]. As expected, germ cells were also sexually dimorphic, being arrested in mitosis in males and meiotically arrested in females [ 123 ]. Novel surface markers were identified for the different types of germ cells, useful for isolating fetal germ cells in meiotic prophase (IL13RA2 + ) and oogenesis (PECAM1 + ) [ 119 ].…”

X-Chromosome Inactivation during Preimplantation Development and in Pluripotent Stem Cells

“…However, more recently, it has been shown that about 30% of hPGCs at 4-9 weeks of development still exhibit incomplete XCR, as suggested by the presence of faint perinuclear spots of H3K27me3, a marker of XCI. XCR appears to be more related to the transcriptional signature of the cells rather than to the fetal age [Vértesy et al, 2018]. These observations suggest that, in hPGCs, XCR is heterogeneous and asynchronous, starting from 4 weeks of development onward [Guo et al, 2015;Tang et al, 2015].…”

“…During postimplantation development, once randomly established, the epigenetic memory determining Xlinked gene silencing is stably inherited through cellular generations. However, in vivo, Xi is reactivated in different cell types like mouse Epi cells [Mak et al, 2004;Borensztein et al, 2017], mouse [Sugimoto and Abe, 2007;Chuva de Sousa Lopes et al, 2008;Mallol et al, 2019] and human [Von Meyenn and Reik, 2015;Vértesy et al, 2018] primordial germ cells (PGCs) and spermatids, during their differentiation into spermatozoa [Ernst et al, 2019].…”

“…In hPGCs, XIST is expressed regardless of the XCR status [Gkountela et al, 2015;Vértesy et al, 2018]. Its expression is not associated with H3K27me3 gathering [Tang et al, 2015], and it is unknown whether it accumulates on the X-chromosomes.…”