Comparative Analyses of H3K4 and H3K27 Trimethylations Between the Mouse Cerebrum and Testis

AbstractThe global features of H3K4 and H3K27 trimethylations (H3K4me3 and H3K27me3) have been well studied in recent years, but most of these studies were performed in mammalian cell lines. In this work, we generated the genome-wide maps of H3K4me3 and H3K27me3 of mouse cerebrum and testis using ChIP-seq and their high-coverage transcriptomes using ribominus RNA-seq with SOLiD technology. We examined the global patterns of H3K4me3 and H3K27me3 in both tissues and found that modifications are closely-associated with tissue-specific expression, function and development. Moreover, we revealed that H3K4me3 and H3K27me3 rarely occur in silent genes, which contradicts the findings in previous studies. Finally, we observed that bivalent domains, with both H3K4me3 and H3K27me3, existed ubiquitously in both tissues and demonstrated an invariable preference for the regulation of developmentally-related genes. However, the bivalent domains tend towards a “winner-takes-all” approach to regulate the expression of associated genes. We also verified the above results in mouse ES cells. As expected, the results in ES cells are consistent with those in cerebrum and testis. In conclusion, we present two very important findings. One is that H3K4me3 and H3K27me3 rarely occur in silent genes. The other is that bivalent domains may adopt a “winner-takes-all” principle to regulate gene expression

TiBi-3D - a Guide through the World of Epigenetics

In the last two decades the study of changes in the genome function that are not induced by changes in DNA has consolidated a strong research field called ”epigenetics”. Chromatin state changes play an essential role in the regulation of transcription of many genes, thus controlling cell differentiation. A large part of these changes is due to histone modifications that alter the accessibility of the DNA. Current state of the art visualization methods for the analysis of epigenetic data sets are not suited to represent the relationship between the combinatorial pattern of histone modifications and their regulatory effects

Genome-wide analysis of trimethylated lysine 4 of histone H3 (H3K4me3) in Aspergillus niger

Aspergillus niger is a commercially important producer of enzymes and organic acids. In this study, I have examined the distribution of trimethylated lysine 4 of histone H3 (H3K4me3) in the A. niger genome. I performed chromatin immunoprecipitation followed by sequencing (ChIP-seq) to determine the genomic regions enriched for nucleosomes with this histone modification. I have conducted an analysis of the resulting peaksets to determine the optimal peak finding parameters for use in the detection of H3K4me3 ChIP-enriched regions in A. niger. As H3K4me3 was previously widely reported to mark actively transcribed genes, genome-wide ChIP-seq maps for maltose and xylose growth were compared with transcriptome data generated under the same growth conditions. Almost all genes that contained H3K4me3 are actively transcribed. However, nearly a third of all actively transcribed genes are not associated with H3K4me3. In addition, H3K4me3 is not associated with the majority of genes differentially expressed on maltose or xylose growth. Chromosomal maps revealed that this histone modification is non-randomly distributed in the genome. In particular, H3K4me3 is enriched at pericentromeric regions, but absent at the centromere and at regions proximal to the telomeres. Finally, H3K4me3 occasionally localized to actively transcribed regions not predicted to contain a gene model. The results of this study suggest that H3K4me3 is positively correlated with transcriptional activity, but is not a definitive marker of active gene expression. Furthermore, this modification is highly locally organized along A. niger chromosomes. Epigenetic phenomena in A. niger warrant further study to determine their significance in genome regulation

THE APPLICATION OF HISTONE ANALYSIS FOR IDENTIFYING DISTAL REGULATORY ELEMENTS - REGULATION OF FSHR

Mechanisms that regulate gene expression are fundamental to many complex biological processes and disease states. Genome-wide approaches that combine chromatin-immunoprecipitation (ChIP) and next-generation sequencing have greatly advanced our understanding of chromatin structure and the role that histone modification plays in transcriptional regulation. In particular, these advances revealed important associations between functional, non-coding DNAs and specific histone modifications, which have been used technically to identify numerous distal regulatory elements and furthered our knowledge of transcriptional regulation and cell-specific gene regulation. Fshr is a gene expressed only in testicular Sertoli cells and ovarian granulosa cells and its expression is critical for proper gonad function and fertility. Importantly, underlying its exquisite cell-specificity is a transcriptional mechanism limited to only two cell types, which previous studies revealed was dependent on distal regulatory elements. To help identify these elements, we used ChIP, combined with next-generation sequencing, to globally map Histone 3 Lysine 4 tri-methylation (H3K4me3) in Sertoli and myoid cells. H3K4me3 is a post-translational histone modification known to associate with distal regulatory elements and promoter regions of actively transcribed genes. Analysis of H3K4me3 enrichment profiles identified a distal site 3' to Fshr that was specific to Sertoli cells. Transient transfection analysis indicated the region represses Fshr promoter activity and in vitro binding analysis revealed that GATA-4 and an unknown protein bound to the region, implicating them in cell-specific regulation of Fshr. These studies provide evidence that Fshr is regulated by a distal regulatory element and have provided insight into the nature of these regulatory proteins. This work provides a database mapping H3K4me3 enrichment within Sertoli and peritubular myoid cells that can be used to identify new regulatory regions. Overall, these studies have furthered our knowledge of cell-specific gene regulation in Sertoli cells and provided new data that will lead to a better understanding of transcriptional regulation in Sertoli cells

The landscape of vascular accessible chromatin and prioritisation of functional risk alleles for cardiovascular disease

Cardiovascular disease is a leading cause of mortality and disability. A meta-analysis of genome-wide association studies (GWAS) of 181,522 coronary artery (CAD) cases and 984,058 controls identified 897 CAD risk variants at the 1% false discovery rate whereas a blood pressure (BP) GWAS meta-analysis in over 1 million individuals identified 901 BP risk alleles. To prioritise CAD and BP variants for functional studies I mapped the vascular landscape of accessible chromatin in human coronary artery endothelial cells (HCAECs) and smooth muscle cells (HCASMCs). Furthermore, I stimulated HCAECs with vascular endothelial growth factor A (VEGFA) to assess its effect on accessible chromatin. This study led to the identification of 86,811 ATAC-seq peaks in unstimulated HCAECs, 108,736 ATAC-seq peaks in VEGFA-stimulated HCAECs, and 209,743 ATAC-seq peaks in HCASMCs. Ras and cellular senescence pathways were specific to HCAEC, whereas cAMP, vascular smooth muscle contraction, and prolactin pathways were specific to HCASMC. The most significantly enriched transcription factor binding motifs in HCAEC included CTCF, BORIS, ATF3, JUNB, and Fra1, whereas in HCASMC I found BATF, AFT3, FRA1, AP-1 and JunB. A total of 17,570 peaks were found to be restricted to HCAEC accessible chromatin, when compared to ENCODE DNaseI-seq data from 125 cell types. They showed enrichment of TGF-beta and Wnt signalling pathways. There are 39,935 ATAC-seq peaks in the unique VEGFA-stimulated HCAEC, which showed enrichment for phospholipase D (PLD) signalling. Intersecting the ATAC-seq data with 21,461 CAD risk variants (all proxies of 897 index variants at r2 >0.8) as well as 14,168 BP variants (all proxies of 901 index variants at r2 >0.8) identified 499 CAD and 517 BP risk variants, respectively, located in vascular ATAC-seq peaks. These variants were further prioritised using tools such as RegulomeDB, Haploreg, and GTEx. In addition, further prioritization of functional CAD SNPs was performed based on the FGWAS approach. Examination of accessible chromatin in vascular cells can be used to prioritise functional variants, facilitating further focused studies on these loci and the eventual identification of novel therapeutics