scATAC-Seq reveals epigenetic heterogeneity associated with an EMT-like process in male germline stem cells and its regulation by G9a

Abstract

Epithelial-mesenchymal transition (EMT) is a phenomenon in which epithelial cells acquire mesenchymal traits. It contributes to organogenesis and tissue homeostasis, as well as stem cell differentiation. Emerging evidence indicates that heterogeneous expression of EMT gene markers presents in sub-populations of germline stem cells (GSCs). However, the functional implications of such heterogeneity are largely elusive. Here, we unravelled an EMT-like process in GSCs by in vitro extracellular matrix (ECM) model and single-cell genomics approaches. Through modulating ECM, we demonstrated that GSCs exist in interconvertible epithelial-like and mesenchymal-like cell states. GSCs gained higher migratory ability after transition to a mesenchymal-like cell state, which was largely mediated by the TGF-β signaling pathway. Dynamics of epigenetic regulation at the single-cell level was also found to align with the EMT-like process. Chromatin accessibility profiles generated by single-cell sequencing assay for transposase-accessible chromatin (scATAC-seq) clustered GSCs into epithelial-like and mesenchymal-like states, which were associated with differentiation status. The high-resolution data revealed novel regulators in the EMT-like process, including transcription factors Zeb1 and Hes1. We further identified putative enhancer-promoter interactions and cis-co-accessibility networks at loci such as Tgfb1, Notch1 and Lin28a. Importantly, we demonstrated that histone methyltransferase G9a promoted the GSC EMT-like process in vitro and contributed to neonatal germ cell migration in vivo. Our work thus provides the foundation for understanding the EMT-like process and a comprehensive resource for future investigation of epigenetic regulatory networks in GSCs.

The single-cell chromatin accessibility landscape in mouse perinatal testis development

“…Assessment of enriched TFs and their cognate motifs identified several known cell type-specific regulators – including the nuclear receptors (NR4A1 and NR5A1) in Sertoli cells and Leydig cells, ESR2 in Leydig cells, MYOG in PTMs, and previously uncharacterized TFs as potential cell type-specific regulators ( Figure 2C ). For example, we found that ZEB1 and SNAI2 motifs were enriched in germ cells, indicating they may undergo mesenchymal-like transition in perinatal development ( Hammoud et al, 2015 ; Liao et al, 2020 ). DNA bound by TFs is protected from transposition by Tn5, which can be visualized by plotting the ‘footprint’ pattern of each TF as the local chromatin accessibility surrounding the motif midpoint.…”

Loss of SDHB Induces a Metabolic Switch in the hPheo1 Cell Line toward Enhanced OXPHOS

“…The interaction with ECM molecules may induce epithelial-to-mesenchymal transition (EMT) [ 37 , 38 ], which is a crucial cellular program that enables polarized epithelial cells to transition toward a mesenchymal phenotype with increased cellular motility [ 39 ]. EMT can be initiated by external signals, such as transforming growth factor (TGF)-β [ 40 , 41 , 42 ].…”

Regulating epithelial-mesenchymal plasticity from 3D genome organization

“…While single cell transcriptomic profiling has enabled us to look at EMT from an individualistic perspective, the chromatin regulation of the transcriptional activity of EMT genes in concert with TFs is still largely uncharted at a single cell resolution. By using Assay for Transposase-Accessible Chromatin using sequencing (ATACseq) in single cells, germline stem cells (GSCs) have been clustered into epithelial-like and mesenchymal-like states that constitute into an EMT spectrum 101 .…”