Selective de-repression of germ cell-specific genes in mouse embryonic fibroblasts in a permissive epigenetic environment.

Tamotsu Sekinaka,Yohei Hayashi,Hitoshi Niwa,Yasuhisa Matsui,Toshiaki Noce

doi:10.1038/srep32932

Abstract

Epigenetic modifications play crucial roles on establishment of tissue-specific transcription profiles and cellular characteristics. Direct conversions of fibroblasts into differentiated tissue cells by over-expression of critical transcription factors have been reported, but the epigenetic mechanisms underlying these conversions are still not fully understood. In addition, conversion of somatic cells into germ cells has not yet been achieved. To understand epigenetic mechanisms that underlie germ cell characteristics, we attempted to use defined epigenetic factors to directly convert mouse embryonic fibroblasts (MEFs) into germ cells. Here, we successfully induced germ cell-specific genes by inhibiting repressive epigenetic modifications via RNAi or small-molecule compounds. Under these conditions, some tissue-specific genes and stimulus-inducible genes were also induced. Meanwhile, the treatments did not result in genome-wide transcriptional activation. These results suggested that a permissive epigenetic environment resulted in selective de-repression of stimulus- and differentiation-inducible genes including germ cell-specific genes in MEFs.

Highlights

embryonic stem cells (ESCs) and primordial germ cells (PGCs) both express pluripotency-associated genes, but germ cell-specific genes are repressed in ESCs, and we suggested that H3K9me[2], which can be generated via Max-associating G9a and GLP, is involved in their repression
We simultaneously used RNAi to knockdown Max and thereby induce germ cell-specific genes (Supplementary Fig. S1); again, our previous findings indicate that the Max transcription factor globally represses germ-cell specific genes in mouse embryonic stem cells, and that Max knockdown (Max-KD) in mESCs results in upregulation of those genes[40]
We found that 1234 genes were upregulated in both PGCs and OS-treated Mouse embryonic fibroblasts (MEFs), and 1427 genes were upregulated in both PGCs and OCKS-treated MEFs; notably, meiosis-related GO terms and genes including Tex19.1, Dazl, Sycp[1] and Sycp[3] were highly enriched in both sets of up-regulated genes (Fig. 4e; Supplementary Fig. S8e; Supplementary Fig. S9; Supplementary Table S3; Supplementary Table S4)

Summary

Introduction

After undergoing complex differentiation processes that include the above-mentioned epigenetic reprogramming, germ cells acquire totipotency through fertilization and can go on to generate an entire organism, but somatic cells derived from the zygote do not normally have this potential. Direct reprogramming of MEFs into cells of particular somatic tissues has been reported[35,36,37,38,39] One such reprogramming strategy involves the overexpression of transcription factors that regulate differentiation into particular cell lineages including hepatocytes, neurons, or Sertoli cells. Direct reprogramming of MEFs into germ-cell lineages, including PGCs, has not yet been reported. ESCs and PGCs both express pluripotency-associated genes, but germ cell-specific genes are repressed in ESCs, and we suggested that H3K9me[2], which can be generated via Max-associating G9a and GLP, is involved in their repression. We tested several conditions to alter the epigenetic state of MEFs into that of PGCs

Methods

Results

Conclusion