Gender Differences in Global but Not Targeted Demethylation in iPSC Reprogramming

Abstract

SummaryGlobal DNA demethylation is an integral part of reprogramming processes in vivo and in vitro, but whether it occurs in the derivation of induced pluripotent stem cells (iPSCs) is not known. Here, we show that iPSC reprogramming involves both global and targeted demethylation, which are separable mechanistically and by their biological outcomes. Cells at intermediate-late stages of reprogramming undergo transient genome-wide demethylation, which is more pronounced in female cells. Global demethylation requires activation-induced cytidine deaminase (AID)-mediated downregulation of UHRF1 protein, and abolishing demethylation leaves thousands of hypermethylated regions in the iPSC genome. Independently of AID and global demethylation, regulatory regions, particularly ESC enhancers and super-enhancers, are specifically targeted for hypomethylation in association with transcription of the pluripotency network. Our results show that global and targeted DNA demethylation are conserved and distinct reprogramming processes, presumably because of their respective roles in epigenetic memory erasure and in the establishment of cell identity.

X Chromosome Dosage Modulates Multiple Molecular and Cellular Properties of Mouse Pluripotent Stem Cells Independently of Global DNA Methylation Levels

“…One outcome of our study is that the number of active X-chromosomes correlates with differences in the transcriptome and in pluripotency exit in iPSCs (Fig 1), in addition to differences in DNA methylation [22,23]. Reprogramming somatic cells to iPSCs is an important system to study erasure of epigenetic memory and pluripotency.…”

“…Work over the past decade showed that XX female ESCs exhibit global DNA hypomethylation affecting most genomic features including imprint control regions [13][14][15][16][17][18][19][20][21]. Recent work showed that XX female iPSCs also display global hypomethylation [22,23]. Differences in global DNA methylation have been attributed to X-dosage since female XO cells display male-like DNA methylation levels [13,15,17].…”

“…While several advances have been made, the molecular pathways by which XaXa modulate pluripotency remain incompletely understood [10]. At the mechanistic level, two active X-chromosomes inhibit MAPK and GSK3 signaling [12,15], and global DNA hypomethylation has been attributed to reduced expression of DNMT3A and DNMT3B [13], or DNMT3L [14], or UHRF1 [17,19,22] in XX female ESCs/iPSCs. More recently, it was discovered that increased dosage of the X-linked MAPK inhibitor Dusp9 (dual-specificity phosphatase 9) is in part responsible for inhibiting DNMT3A/B/L and global DNA methylation in XX female ESCs [17].…”

Induced pluripotent stem cells (iPSCs) as a new source of bone in reconstructive surgery: A systematic review and meta-analysis of preclinical studies

“…iPSCs can differentiate into nearly all types of cells in the body. A major principle in iPSCs was the cellular reprogramming technology using various methods as somatic cell nuclear transfer, cell fusion, and currently by means of reprogramming factors, which helped to establish and maintain cellular identity during development by the stepwise activation of fundamental pluripotency genes and the silencing of somatic cell‐of‐origin genes (Ma, Xie, Laurent, & Ding, ; Milagre et al, ; Szablowska‐Gadomska, Gorska, & Malecki, ; Zhang, ). In our review, the most commonly used reprogramming factors were either Yamanaka (OSKM) or Thomson reprogramming factors (OSNL; Zhang, ).…”

Xmas ESC: A new female embryonic stem cell system that reveals the BAF complex as a key regulator of the establishment of X chromosome inactivation

“…Female ESCs are also karyotypically unstable, with XO cells rapidly dominating cultures (Choi et al, 2017b;Yagi et al, 2017;Zvetkova et al, 2005), which is also observed in human female embryonic stem cells (Di Stefano et al, 2018). These complications additionally arise when somatic female cells are reprogrammed to induced pluripotent stem cells (iPSCs) (Milagre et al, 2017;Pasque et al, 2018;Song et al, 2019), suggesting that these are features of female pluripotency in culture that confound both research requiring such cells and future medical applications.…”