Epigenetics & Chromatin最新文献

Background: CTCF is highly likely to be the ancestor of proteins that contain large clusters of C2H2 zinc finger domains, and its conservation is observed across most bilaterian organisms. In mammals, CTCF is the primary architectural protein involved in organizing chromosome topology and mediating enhancer-promoter interactions over long distances. In Drosophila, CTCF (dCTCF) cooperates with other architectural proteins to establish long-range interactions and chromatin boundaries. CTCFs of various organisms contain an unstructured N-terminal dimerization domain (DD) and clusters comprising eleven zinc-finger domains of the C2H2 type. The Drosophila (dCTCF) and human (hCTCF) CTCFs share sequence homology in only five C2H2 domains that specifically bind to a conserved 15 bp motif.

Results: Previously, we demonstrated that CTCFs from different organisms carry unstructured N-terminal dimerization domains (DDs) that lack sequence homology. Here we used the CTCF^attP(mCh) platform to introduce desired changes in the Drosophila CTCF gene and generated a series of transgenic lines expressing dCTCF with different variants of the N-terminal domain. Our findings revealed that the functionality of dCTCF is significantly affected by the deletion of the N-terminal DD. Additionally, we observed a strong impact on the binding of the dCTCF mutant to chromatin upon deletion of the DD. However, chromatin binding was restored in transgenic flies expressing a chimeric CTCF protein with the DD of hCTCF. Although the chimeric protein exhibited lower expression levels than those of the dCTCF variants, it efficiently bound to chromatin similarly to the wild type (wt) protein.

Conclusions: Our findings suggest that one of the evolutionarily conserved functions of the unstructured N-terminal dimerization domain is to recruit dCTCF to its genomic sites in vivo.

Chromatin state is thought to impart regulatory function to the underlying DNA sequence. This can be established through histone modifications and chromatin organisation, but exactly how these factors relate to one another to regulate gene expression is unclear. In this study, we have used super-resolution microscopy to image the Y loops of Drosophila melanogaster primary spermatocytes, which are enormous transcriptionally active chromatin fibres, each representing single transcription units that are individually resolvable in the nuclear interior. We previously found that the Y loops consist of regular clusters of nucleosomes, with an estimated median of 54 nucleosomes per cluster with wide variation.In this study, we report that the histone modifications H3K4me3, H3K27me3, and H3K36me3 are also clustered along the Y loops, with H3K4me3 more associated with diffuse chromatin compared to H3K27me3. These histone modifications form domains that can be stretches of Y loop chromatin micrometres long, or can be in short alternating domains. The different histone modifications are associated with different sizes of chromatin clusters and unique morphologies. Strikingly, a single chromatin cluster almost always only contains only one type of the histone modifications that were labelled, suggesting exclusivity, and therefore regulation at the level of individual chromatin clusters. The active mark H3K36me3 is more associated with actively elongating RNA polymerase II than H3K27me3, with polymerase often appearing on what are assumed to be looping regions on the periphery of chromatin clusters.These results provide a foundation for understanding the relationship between chromatin state, chromatin organisation, and transcription regulation - with potential implications for pause-release dynamics, splicing complex organisation and chromatin dynamics during polymerase progression along a gene.

Background: Prostate adenocarcinoma (PRAD) is the second leading cause of cancer-related deaths in men. High variability in DNA methylation and a high rate of large genomic rearrangements are often observed in PRAD.

Results: To investigate the reasons for such high variance, we integrated DNA methylation, RNA-seq, and copy number alterations datasets from The Cancer Genome Atlas (TCGA), focusing on PRAD, and employed weighted gene co-expression network analysis (WGCNA). Our results show that only single cluster of co-expressed genes is associated with genomic and epigenomic instability. Within this cluster, TP63 and TRIM29 are key transcription regulators and are downregulated in PRAD. We discovered that TP63 regulates the level of enhancer methylation in prostate basal epithelial cells. TRIM29 forms a complex with TP63 and together regulates the expression of genes specific to the prostate basal epithelium. In addition, TRIM29 binds DNA repair proteins and prevents the formation of the TMPRSS2:ERG gene fusion typically observed in PRAD.

Conclusion: Our study demonstrates that TRIM29 and TP63 are important regulators in maintaining the identity of the basal epithelium under physiological conditions. Furthermore, we uncover the role of TRIM29 in PRAD development.

Protein and nucleic acid methylation are important biochemical modifications. In addition to their well-established roles in gene regulation, they also regulate cell signaling, metabolism, and translation. Despite this high biological relevance, little is known about the general regulation of methyltransferase function. Methyltransferases are divided into superfamilies based on structural similarities and further classified into smaller families based on sequence/domain/target similarity. While members within superfamilies differ in substrate specificity, their structurally similar active sites indicate a potential for shared modes of regulation. Growing evidence from one superfamily suggests a common regulatory mode may be through heterooligomerization with other family members. Here, we describe examples of methyltransferase regulation through intrafamily heterooligomerization and discuss how this can be exploited for therapeutic use.

In the past decade, numerous epigenetic mechanisms have been discovered to be associated with cancer. The mammalian SWI/SNF complex is an ATP-dependent chromatin remodeling complex whose mutations are associated with various malignancies including breast cancer. As the SWI/SNF complex has become one of the most commonly mutated complexes in cancer, targeting epigenetic mutations acquired during breast cancer progress is a potential means of improving clinical efficacy in treatment strategies. This article reviews the composition of the SWI/SNF complex, its main roles and research progress in breast cancer, and links these findings to the latest discoveries in cancer epigenomics to discuss the potential mechanisms and therapeutic potential of SWI/SNF in breast cancer.

Background: Bivalent chromatin is an exemplar of epigenetic plasticity. This co-occurrence of active-associated H3K4me3 and inactive-associated H3K27me3 histone modifications on opposite tails of the same nucleosome occurs predominantly at promoters that are poised for future transcriptional upregulation or terminal silencing. We know little of the dynamics, resolution, and regulation of this chromatin state outside of embryonic stem cells where it was first described. This is partly due to the technical challenges distinguishing bone-fide bivalent chromatin, where both marks are on the same nucleosome, from allelic or sample heterogeneity where there is a mix of H3K4me3-only and H3K27me3-only mononucleosomes.

Results: Here, we present a robust and sensitive method to accurately map bivalent chromatin genome-wide, along with controls, from as little as 2 million cells. We optimized and refined the sequential ChIP protocol which uses two sequential overnight immunoprecipitation reactions to robustly purify nucleosomes that are truly bivalent and contain both H3K4me3 and H3K27me3 modifications. Our method generates high quality genome-wide maps with strong peak enrichment and low background, which can be analyzed using standard bioinformatic packages. Using this method, we detect 8,789 bivalent regions in mouse embryonic stem cells corresponding to 3,918 predominantly CpG rich and developmentally regulated gene promoters. Furthermore, profiling Dppa2/4 knockout mouse embryonic stem cells, which lose both H3K4me3 and H3K27me3 at approximately 10% of bivalent promoters, demonstrated the ability of our method to capture bivalent chromatin dynamics.

Conclusions: Our optimized sequential reChIP method enables high-resolution genome-wide assessment of bivalent chromatin together with all required controls in as little as 2 million cells. We share a detailed protocol and guidelines that will enable bivalent chromatin landscapes to be generated in a range of cellular contexts, greatly enhancing our understanding of bivalent chromatin and epigenetic plasticity beyond embryonic stem cells.

Background: Blood homeostasis requires the daily production of millions of terminally differentiated effector cells that all originate from hematopoietic stem cells (HSCs). HSCs are rare and exhibit unique self-renewal and multipotent properties, which depend on their ability to maintain quiescence through ill-defined processes. Defective control of cell cycle progression can eventually lead to bone marrow failure or malignancy. In particular, the molecular mechanisms tying cell cycle re-entry to cell fate commitment in HSCs remain elusive. Previous studies have identified chromatin coordination as a key regulator of differentiation in embryonic stem cells.

Results: Here, we utilized genetic inactivation of the chromatin-associated Sin3B protein to manipulate cell cycle control and found dysregulated chromatin accessibility and cell cycle progression in HSCs. Single cell transcriptional profiling of hematopoietic stem and progenitor cells (HSPCs) inactivated for Sin3B reveals aberrant progression through the G₁ phase of the cell cycle, which correlates with the engagement of specific signaling pathways, including aberrant expression of cell adhesion molecules and the interferon signaling program in LT-HSCs. In addition, we uncover the Sin3B-dependent accessibility of genomic elements controlling HSC differentiation, which points to cell cycle progression possibly dictating the priming of HSCs for differentiation.

Conclusions: Our findings provide new insights into controlled cell cycle progression as a potential regulator of HSC lineage commitment through the modulation of chromatin features.