Keiji Hirabayashi

Progesterone is a cell death suppressor that downregulates Fas expression in rat corpus luteum

In female rats, apoptotic cell death in the corpus luteum is induced by the prolactin (PRL) surge occurring in the proestrous afternoon during the estrous cycle. We have previously shown that this luteolytic action of PRL is mediated by the Fas/Fas ligand (FasL) system. During pregnancy or pseudopregnancy, apoptosis does not occur in the corpus luteum. Progesterone (P4), a steroid hormone secreted from luteal steroidogenic cells, attenuated PRL‐induced apoptosis in cultured luteal cells in a dose‐dependent manner. P4 significantly decreased the expression of mRNA of Fas, but not FasL, in cultured luteal cells prepared from both proestrous and mid‐pseudopregnant rats. These data indicate that P4 suppresses PRL‐induced luteal cell apoptosis via reduction of the expression level of Fas mRNA in the corpus luteum, suggesting that P4 acts as an important factor that can change the sensitivity of corpus luteum to PRL.

DNA methylation status of nuclear-encoded mitochondrial genes underlies the tissue-dependent mitochondrial functions

BackgroundMitochondria are semi-autonomous, semi-self-replicating organelles harboring their own DNA (mitochondrial DNA, mtDNA), and their dysregulation is involved in the development of various diseases. While mtDNA does not generally undergo epigenetic modifications, almost all mitochondrial proteins are encoded by nuclear DNA. However, the epigenetic regulation of nuclear-encoded mitochondrial genes (nuclear mt genes) has not been comprehensively analyzed.ResultsWe analyzed the DNA methylation status of 899 nuclear mt genes in the liver, brain, and heart tissues of mouse, and identified 636 nuclear mt genes carrying tissue-dependent and differentially methylated regions (T-DMRs). These nuclar mt genes are involved in various mitochondrial functions and they also include genes related to human diseases. T-DMRs regulate the expression of nuclear mt genes. Nuclear mt genes with tissue-specific hypomethylated T-DMRs were characterized by enrichment of the target genes of specific transcription factors such as FOXA2 in the liver, and CEBPA and STAT1 in the brain.ConclusionsA substantial proportion of nuclear mt genes contained T-DMRs, and the DNA methylation status of numerous T-DMRs should underlie tissue-dependent mitochondrial functions.

Genome-wide DNA methylation profile of tissue-dependent and differentially methylated regions (T-DMRs) residing in mouse pluripotent stem cells

DNA methylation profile, consisting of tissue‐dependent and differentially methylated regions (T‐DMRs), has elucidated tissue‐specific gene function in mouse tissues. Here, we identified and profiled thousands of T‐DMRs in embryonic stem cells (ESCs), embryonic germ cells (EGCs) and induced pluripotent stem cells (iPSCs). T‐DMRs of ESCs compared with somatic tissues well illustrated gene function of ESCs, by hypomethylation at genes associated with CpG islands and nuclear events including transcriptional regulation network of ESCs, and by hypermethylation at genes for tissue‐specific function. These T‐DMRs in EGCs and iPSCs showed DNA methylation similar to ESCs. iPSCs, however, showed hypomethylation at a considerable number of T‐DMRs that were hypermethylated in ESCs, suggesting existence of traceable progenitor epigenetic information. Thus, DNA methylation profile of T‐DMRs contributes to the mechanism of pluripotency, and can be a feasible solution for identification and evaluation of the pluripotent cells.

Enrichment of short interspersed transposable elements to embryonic stem cell‐specific hypomethylated gene regions

Embryonic stem cells (ESCs) have a distinctive epigenome, which includes their genome‐wide DNA methylation modification status, as represented by the ESC‐specific hypomethylation of tissue‐dependent and differentially methylated regions (T‐DMRs) of Pou5f1 and Nanog. Here, we conducted a genome‐wide investigation of sequence characteristics associated with T‐DMRs that were differentially methylated between ESCs and somatic cells, by focusing on transposable elements including short interspersed elements (SINEs), long interspersed elements (LINEs) and long terminal repeats (LTRs). We found that hypomethylated T‐DMRs were predominantly present in SINE‐rich/LINE‐poor genomic loci. The enrichment for SINEs spread over 300 kb in cis and there existed SINE‐rich genomic domains spreading continuously over 1 Mb, which contained multiple hypomethylated T‐DMRs. The characterization of sequence information showed that the enriched SINEs were relatively CpG rich and belonged to specific subfamilies. A subset of the enriched SINEs were hypomethylated T‐DMRs in ESCs at Dppa3 gene locus, although SINEs are overall methylated in both ESCs and the liver. In conclusion, we propose that SINE enrichment is the genomic property of regions harboring hypomethylated T‐DMRs in ESCs, which is a novel aspect of the ESC‐specific epigenomic information.

DNA methylation profile of Aire-deficient mouse medullary thymic epithelial cells.

BackgroundMedullary thymic epithelial cells (mTECs) are characterized by ectopic expression of self-antigens during the establishment of central tolerance. The autoimmune regulator (Aire), which is specifically expressed in mTECs, is responsible for the expression of a large repertoire of tissue-restricted antigens (TRAs) and plays a role in the development of mTECs. However, Aire-deficient mTECs still express TRAs. Moreover, a subset of mTECs, which are considered to be at a stage of terminal differentiation, exists in the Aire-deficient thymus. The phenotype of a specific cell type in a multicellular organism is governed by the epigenetic regulation system. DNA methylation modification is an important component of this system. Every cell or tissue type displays a DNA methylation profile, consisting of tissue-dependent and differentially methylated regions (T-DMRs), and this profile is involved in cell-type-specific genome usage. The aim of this study was to examine the DNA methylation profile of mTECs by using Aire-deficient mTECs as a model.ResultsWe identified the T-DMRs of mTECs (mTEC-T-DMRs) via genome-wide DNA methylation analysis of Aire−/− mTECs by comparison with the liver, brain, thymus, and embryonic stem cells. The hypomethylated mTEC-T-DMRs in Aire−/− mTECs were associated with mTEC-specific genes, including Aire, CD80, and Trp63, as well as other genes involved in the RANK signaling pathway. While these mTEC-T-DMRs were also hypomethylated in Aire+/+ mTECs, they were hypermethylated in control thymic stromal cells. We compared the pattern of DNA methylation levels at a total of 55 mTEC-T-DMRs and adjacent regions and found that the DNA methylation status was similar for Aire+/+ and Aire−/− mTECs but distinct from that of athymic cells and tissues.ConclusionsThese results indicate a unique DNA methylation profile that is independent of Aire in mTECs. This profile is distinct from other cell types in the thymic microenvironment and is indicated to be involved in the differentiation of the mTEC lineage.

Novel sex-dependent differentially methylated regions are demethylated in adult male mouse livers.

In mammalian livers, sexual dimorphisms are observed in tissue-specific functions and diseases such as hepatocellular carcinoma. We identified sex-dependent differentially methylated regions (S-DMRs) which had been previously been characterized as growth hormone- STAT5 dependent. In this study, we performed genome-wide screening and identified ten additional hypomethylated S-DMR gene regions in male livers. Of these S-DMRs, Uggt2 and Sarnp were hypomethylated in both male and female livers compared to brain and embryonic stem (ES) cells. Similarly, Adam2, Uggt2, and Scp2 were hypomethylated in female embryonic germ (EG) cells and not in male EG cells, indicating that these S-DMRs are liver-specific male hypo-S-DMRs. Interestingly, the five S-DMRs were free from STAT5 chromatin immunoprecipitation (ChIP) signals, suggesting that S-DMRs are independent of the growth hormone-STAT5-pathway. Instead, the DNA methylation statuses of the S-DMRs of Adam2, Snx29, Uggt2, Sarnp, and Rnpc3 genes were under the control of testosterone. Importantly, the hypomethylated S-DMRs of the Adam2 and Snx29 regions showed chromatin decondensation. Epigenetic factors could be responsible for the sexual dimorphisms in DNA methylation status and chromatin structure, as the expression of Dnmt1, Dnmt3b, and Tet2 genes was lower in male mice compared to female mice and TET2 expression recovered following orchidectomy by testosterone treatment. In conclusion, we identified novel male-specific hypomethylated S-DMRs that contribute to chromatin decondensation in the liver. S-DMRs were tissue-specific and the hypomethylation is testosterone-dependent.