The role of maternal-specific H3K9me3 modification in establishing imprinted X-chromosome inactivation and embryogenesis in mice

Maintaining a single active X-chromosome by repressing Xist is crucial for embryonic development in mice. Although the Xist activator RNF12/RLIM is present as a maternal factor, maternal Xist (Xm-Xist) is repressed during preimplantation phases to establish imprinted X-chromosome inactivation (XCI). Here we show, using a highly reproducible chromatin immunoprecipitation method that facilitates chromatin analysis of preimplantation embryos, that H3K9me3 is enriched at the Xist promoter region, preventing Xm-Xist activation by RNF12. The high levels of H3K9me3 at the Xist promoter region are lost in embryonic stem (ES) cells, and ES-cloned embryos show RNF12-dependent Xist expression. Moreover, lack of Xm-XCI in the trophectoderm, rather than loss of paternally expressed imprinted genes, is the primary cause of embryonic lethality in 70–80% of parthenogenotes immediately after implantation. This study reveals that H3K9me3 is involved in the imprinting that silences Xm-Xist. Our findings highlight the role of maternal-specific H3K9me3 modification in embryo development

Gene expression signatures associated with chronic endometritis revealed by RNA sequencing

IntroductionChronic endometritis (CE) is a persistent inflammatory condition of the endometrium characterized by the infiltration of plasma cells in the endometrial stroma. CD138 immunohistochemistry is considered to improve the CE diagnosis rate.MethodsUsing the number of CD138-positive cells equal or greater than five as a diagnostic criterion for CE, we identified 24 CE and 33 non-CE cases among women with infertility. We conducted RNA-sequencing analysis for these 57 cases in total as an attempt to elucidate the molecular pathogenesis of CE and to search for new biomarkers for CE.Results and DiscussionBy comparing CE and non-CE groups, we identified 20 genes upregulated in the endometria of CE patients, including 12 immunoglobulin-related genes and eight non-immunoglobulin genes as differentially expressed genes. The eight genes were MUC5AC, LTF, CAPN9, MESP1, ACSM1, TVP23A, ALOX15, and MZB1. By analyzing samples in the proliferative and secretory phases of the menstrual cycle separately, we also identified four additional non-immunoglobulin genes upregulated in CE endometria: CCDC13 by comparing the samples in the proliferative phase, and OVGP1, MTUS2, and CLIC6 by comparing the samples in the secretory phase. Although the genes upregulated in CE may serve as novel diagnostic markers of CE, many of them were upregulated only in a limited number of CE cases showing an extremely high number of CD138-positive cells near or over one hundred. Exceptionally, TVP23A was upregulated in the majority of CE cases regardless of the number of CD138-positive cells. The upregulation of TVP23A in the endometria of CE cases may reflect the pathophysiology of a cell-type or cell-types intrinsic to the endometrium rather than the accumulation of plasma cells. Our data, consisting of clinical and transcriptomic information for CE and non-CE cases, helped us identify gene expression signatures associated with CE

Data from: Reciprocal changes of H3K27ac and H3K27me3 at the promoter regions of the critical genes for endometrial decidualization

Aim: Decidualization is essential for embryo implantation and placental development. We aimed to obtain transcriptome and epigenome profiles for primary endometrial stromal cells (ESCs) and in vitro decidualized cells. Materials & methods: ESCs isolated from human endometrial tissues remained untreated (D0), or decidualized for 4 days (D4) and 8 days (D8) in the presence of 8-bromo-cAMP and progesterone. Results: Among the epigenetic modifications examined (DNA methylation, H3K27ac, H3K9me3 and H3K27me3), the H3K27ac patterns changed most dramatically, with a moderate correlation with gene expression changes, upon decidualization. Subsets of up- and down-regulated genes upon decidualization were associated with reciprocal changes of H3K27ac and H3K27me3 modifications at their promoter region, and were enriched with genes essential for decidualization such as WNT4, ZBTB16, PROK1 and GREB1. Conclusion: Our dataset is useful to further elucidate the molecular mechanisms underlying decidualization

Tables for fpkm values

Tables for fpkm values for RNA-seq data calculated using Cufflinks:genes.fpkm_tracking, isoforms.fpkm_tracking, and tss_groups.fpkm_trackin

Peak call files for ChIP-seq data

Peak call files for CHIP-seq data produced by MACS2:EM0409D0_K27ac_macs2_peaks.narrowPeak EM0409D0_K27me3_macs2_peaks.broadPeak EM0409D0_K9me3_macs2_peaks.broadPeak EM0409D4_K27ac_macs2_peaks.narrowPeak EM0409D4_K27me3_macs2_peaks.broadPeak EM0409D4_K9me3_macs2_peaks.broadPeak EM0409D8_K27ac_macs2_peaks.narrowPeak EM0409D8_K27me3_macs2_peaks.broadPeak EM0409D8_K9me3_macs2_peaks.broadPeak EM0519D0_K27ac_macs2_peaks.narrowPeak EM0519D0_K27me3_macs2_peaks.broadPeak EM0519D0_K9me3_macs2_peaks.broadPeak EM0519D4_K27ac_macs2_peaks.narrowPeak EM0519D4_K27me3_macs2_peaks.broadPeak EM0519D4_K9me3_macs2_peaks.broadPeak EM0519D8_K27ac_macs2_peaks.narrowPeak EM0519D8_K27me3_macs2_peaks.broadPeak EM0519D8_K9me3_macs2_peaks.broadPea

bigwig_files

"bigwig_files.zip" (md5 checksum: a7cee3f55c6c3d53dd562475aa7fc1f3). The "bigwig_files.zip" file contains the following 25 bigwig files (19 files for ChIP-seq data and 6 files for RNA-sea data). EM0409_D0_H3K27ac.hg19.bw EM0409_D0_H3K27me3.hg19.bw EM0409_D0_H3K9me3.hg19.bw EM0409_D4_H3K27ac.hg19.bw EM0409_D4_H3K27me3.hg19.bw EM0409_D4_H3K9me3.hg19.bw EM0409_D4_input.hg19.bw EM0409_D8_H3K27ac.hg19.bw EM0409_D8_H3K27me3.hg19.bw EM0409_D8_H3K9me3.hg19.bw EM0519_D0_H3K27ac.hg19.bw EM0519_D0_H3K27me3.hg19.bw EM0519_D0_H3K9me3.hg19.bw EM0519_D4_H3K27ac.hg19.bw EM0519_D4_H3K27me3.hg19.bw EM0519_D4_H3K9me3.hg19.bw EM0519_D4_input.hg19.bw EM0519_D8_H3K27me3.hg19.bw EM0519_D8_H3K9me3.hg19.bw RNAseq_EM0409_D0.hg19.bw RNAseq_EM0409_D4.hg19.bw RNAseq_EM0409_D8.hg19.bw RNAseq_EM0519_D0.hg19.bw RNAseq_EM0519_D4.hg19.bw RNAseq_EM0519_D8.hg19.b

An Insulator Element Located at the Cyclin B1 Interacting Protein 1 Gene Locus Is Highly Conserved among Mammalian Species

<div><p>Insulators are <i>cis</i>-elements that control the direction of enhancer and silencer activities (enhancer-blocking) and protect genes from silencing by heterochromatinization (barrier activity). Understanding insulators is critical to elucidate gene regulatory mechanisms at chromosomal domain levels. Here, we focused on a genomic region upstream of the mouse <i>Ccnb1ip1</i> (cyclin B1 interacting protein 1) gene that was methylated in E9.5 embryos of the C57BL/6 strain, but unmethylated in those of the 129X1/SvJ and JF1/Ms strains. We hypothesized the existence of an insulator-type element that prevents the spread of DNA methylation within the 1.8 kbp segment, and actually identified a 242-bp and a 185-bp fragments that were located adjacent to each other and showed insulator and enhancer activities, respectively, in reporter assays. We designated these genomic regions as the <i>Ccnb1ip1</i> insulator and the <i>Ccnb1ip1</i> enhancer. The <i>Ccnb1ip1</i> insulator showed enhancer-blocking activity in the luciferase assays and barrier activity in the colony formation assays. Further examination of the <i>Ccnb1ip1</i> locus in other mammalian species revealed that the insulator and enhancer are highly conserved among a wide variety of species, and are located immediately upstream of the transcriptional start site of <i>Ccnb1ip1</i>. These newly identified cis-elements may be involved in transcriptional regulation of <i>Ccnb1ip1</i>, which is important in meiotic crossing-over and G2/M transition of the mitotic cell cycle.</p></div

Genomic, Epigenomic, and Transcriptomic Profiling towards Identifying Omics Features and Specific Biomarkers That Distinguish Uterine Leiomyosarcoma and Leiomyoma at Molecular Levels

Uterine leiomyosarcoma (LMS) is the worst malignancy among the gynecologic cancers. Uterine leiomyoma (LM), a benign tumor of myometrial origin, is the most common among women of childbearing age. Because of their similar symptoms, it is difficult to preoperatively distinguish the two conditions only by ultrasound and pelvic MRI. While histopathological diagnosis is currently the main approach used to distinguish them postoperatively, unusual histologic variants of LM tend to be misdiagnosed as LMS. Therefore, development of molecular diagnosis as an alternative or confirmatory means will help to diagnose LMS more accurately. We adopted omics-based technologies to identify genome-wide features to distinguish LMS from LM and revealed that copy number, gene expression, and DNA methylation profiles successfully distinguished these tumors. LMS was found to possess features typically observed in malignant solid tumors, such as extensive chromosomal abnormalities, overexpression of cell cycle-related genes, hypomethylation spreading through large genomic regions, and frequent hypermethylation at the polycomb group target genes and protocadherin genes. We also identified candidate expression and DNA methylation markers, which will facilitate establishing postoperative molecular diagnostic tests based on conventional quantitative assays. Our results demonstrate the feasibility of establishing such tests and the possibility of developing preoperative and noninvasive methods

Colony formation assays to evaluate the barrier activity of the <i>Ccnb1ip1</i> insulator.

<p>The <i>Ccnb1ip1</i> insulator or the cHS4 insulator was cloned at both sides of the neomycin-resistant gene (PGK-neo). Red and purple boxes represent the <i>Ccnb1ip1</i> insulator, and cHS4 insulator, respectively. Cos7 cells were transfected with these vectors in triplicates. The average numbers of the colonies formed without G418 selection are 86, 72, and 82, for PGK-neo alone, PGK-neo with <i>Ccnb1ip1</i> insulators, and PGK-neo with cHS4 insulators, respectively. The average numbers of the colonies formed with G418 selection are 9, 14, and 24, for PGK-neo alone, PGK-neo with <i>Ccnb1ip1</i> insulators, and PGK-neo with cHS4 insulators, respectively. Percentage numbers of the G418-resistant colonies relative to that for PGK-neo alone are shown for PGK-neo with <i>Ccnb1ip1</i> insulators and PGK-neo with cHS4 insulators (mean ± sd, n = 3).</p