Global and gene-specific histone modification profiles of mouse multipotent adult germline stem cells

Abstract

abstract:We previously reported the generation of multipotent adult germline stem cells (maGSCs) from spermatogonial stem cells (SSCs) isolated from adult mouse testis. In a later study, we substantiated the pluripotency of maGSCs by demonstrating their close similarity to pluripotent male embryonic stem cells (ESCs) at the epigenetic level of global and gene-specific DNA methylation. Here, we extended the comparative epigenetic analysis of maGSCs and male ESCs by investigating the second main epigenetic modification in mammals, i.e. global and gene-specific modifications of histones (H3K4 trimethylation, H3K9 acetylation, H3K9 trimethylation and H3K27 trimethylation). Using immunofluorescence staining, flow cytometry and western blot analysis, we show that maGSCs are very similar to male ESCs with regard to global levels and nuclear distribution patterns of these modifications. Chromatin immunoprecipitation real-time PCR analysis of these modifications at the gene-specific level further revealed modification patterns of the pluripotency marker genes Oct4, Sox2 and Nanog in maGSCs that are nearly identical to those of male ESCs. These genes were enriched for activating histone modifications including H3K4me3 and H3K9ac and depleted of repressive histone modifications including H3K27me3 and H3K9me3. In addition, Hoxa11, a key regulator of early embryonic development showed the ESC-typical bivalent chromatin conformation with enrichment of both the activating H3K4me3 and the repressive H3K27me3 modification also in maGSCs. Collectively, our results demonstrate that maGSCs also closely resemble ESCs with regard to their chromatin state and further evidence their pluripotent nature.

Kinetics of HIV-1 Latency Reversal Quantified on the Single-Cell Level Using a Novel Flow-Based Technique

“…RMD stimulation resulted in significant cell toxicity at later time points, not allowing an assessment at later time points following stimulation (data not shown). RMD has been shown to affect the acetylation status of H3K9 in treated cells (32,33). Indeed, RMD treatment resulted in acetylated H3K9 (H3K9ac) upregulation at 24 h poststimulation in double-negative cells compared to unstimulated J89 cells, which was increased in cells that reactivated HIV-1 (Fig.…”

H3K9 histone acetylation predicts pluripotency and reprogramming capacity of ES cells

“…overall mild, increased histone acetylation may support pervasive low-level transcription across the entire genome as we previously observed in R1 ESCs. 35 Whether pervasive transcription keeps an open chromatin conformation or whether it is a byproduct of open chromatin remains to be seen, 42 but it may well go hand in hand with increased H3K9 acetylation level as observed in germ cells 43 and human ESCs as well. 44 Taken together, our data support the idea that pluripotency is associated with H3K9ac levels, and that HDACi may restore reprogramming efficiency and stem cell potency by promoting ECM activity.…”

Endometriosis Is Characterized by a Distinct Pattern of Histone 3 and Histone 4 Lysine Modifications

“…Positive and negative controls were processed using anti-RNA polymerase II and normal mouse immunoglobulin G, respectively. The immune complexes and input controls were incubated with proteinase K at 65 C, transferred to the column, washed with 70% and 90% ethanol, and finally the purified DNA was eluted.…”