Unique Epigenetic Programming Distinguishes Regenerative Spermatogonial Stem Cells in the Developing Mouse Testis

Abstract

Summary Spermatogonial stem cells (SSCs) both self-renew and give rise to progenitors that initiate spermatogenic differentiation in the mammalian testis. Questions remain regarding the extent to which the SSC and progenitor states are functionally distinct. Here we provide the first multiparametric integrative analysis of mammalian germ cell epigenomes comparable with that done for >100 somatic cell types by the ENCODE Project. Differentially expressed genes distinguishing SSC- and progenitor-enriched spermatogonia showed distinct histone modification patterns, particularly for H3K27ac and H3K27me3. Motif analysis predicted transcription factors that may regulate spermatogonial subtype-specific fate, and immunohistochemistry and gene-specific chromatin immunoprecipitation analyses confirmed subtype-specific differences in target gene binding of a subset of these factors. Taken together, these results show that SSCs and progenitors display distinct epigenetic profiling consistent with these spermatogonial subtypes being differentially programmed to either self-renew and maintain regenerative capacity as SSCs or lose regenerative capacity and initiate lineage commitment as progenitors.

An mTORC1-dependent switch orchestrates the transition between mouse spermatogonial stem cells and clones of progenitor spermatogonia

“…The advent of technologies to survey the entire mRNA transcriptome with single-cell resolution (single-cell RNA sequencing [scRNA-seq]) reveals transcriptome heterogeneity among otherwise homogeneous cells to resolve potential subtypes ( Lafzi et al, 2018 ). Using scRNA-seq datasets from testicular germ cells, we and others ordered spermatogenic cells in pseudotime to distinguish cell states (e.g., SSC versus progenitor and subdivisions thereof) and to infer cell trajectories reflective of state transitions and the responsible regulatory framework ( Cheng et al, 2020 ; Green et al, 2018 ; Hermann et al, 2018 ; La et al, 2018b ; Li et al, 2017 ; Lukassen et al, 2018 ; Song et al, 2016 ; Suzuki et al, 2019 ). Although these analyses have uncovered many cell states and pathways, they are limited by computational assumptions and the static nature of steady-state mRNA levels.…”

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

“…To select links connecting distal regulatory element and promoter, two peaks in each pair were annotated by ChIPseeker separately and the links with only one peak falling in the promoter region were retained. To identify enhancers, datasets of H3K4me3, H3K27me3, H3K4me1, H3K4me2 and H3K27ac modifications in Id4-GFP-bright SSC were downloaded from GSE131656 56 . ChromHMM was used to train a 12 state model on all histone marks assayed.…”

“…Chromatin states of DARs among different clusters, topic regions and distal element-promoter links were annotated using annotatr package 110 , and candidate regions overlap “Active enhancer” states were considered as putative enhancers. ROSE was used for the identification of super-enhancers with H3K27ac dataset (GSE131656) 56 . Overlapping regions of the resulting sets of superenhancers and CCANs were evaluated using makeVennDiagram in ChIPpeakAnno.…”

“…All data in this study have been deposited in the Gene Expression Omnibus (GEO). Histone from ref 56 are available from GSE131656. Hi-C data from ref 64 is available from GSE147536.…”

“…First, we identified the top 500 differentially accessible regions (DARs) within each cluster. Then, we located the enhancer regions by considering chromatin state revealed by chromHMM using recent ChIP-seq profiling of Id4-GFP spermatogonia [64] . Following this, we uncovered potential stage-specific enhancers of each cluster (Supplementary Figure S10).…”