Transcription reprogramming during cell differentiation involves targeting enhancers to genes responsible for establishment of cell fates. To understand the contribution of CTCF-mediated chromatin organization to cell lineage commitment, we analyzed 3D chromatin architecture during the differentiation of human embryonic stem cells into pancreatic islet organoids. We find that CTCF loops are formed and disassembled at different stages of the differentiation process by either recruitment of CTCF to new anchor sites or use of pre-existing sites not previously involved in loop formation. Recruitment of CTCF to new sites in the genome involves demethylation of H3K9me3 to H3K9me2, demethylation of DNA, recruitment of pioneer factors, and positioning of nucleosomes flanking the new CTCF sites. Existing CTCF sites not involved in loop formation become functional loop anchors via the establishment of new cohesin loading sites containing NIPBL and YY1 at sites between the new anchors. In both cases, formation of new CTCF loops leads to strengthening of enhancer promoter interactions and increased transcription of genes adjacent to loop anchors. These results suggest an important role for CTCF and cohesin in controlling gene expression during cell differentiation.
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Jordi Fernandez-Albert;
Michal Lipinski;
Maria T. Lopez-Cascales;
M. Jordan Rowley;
Ana M. Martin-Gonzalez;
Beatriz del Blanco;
Victor Corces;
Angel Barco
Activity-driven transcription plays an important role in many brain processes, including those underlying memory and epilepsy. Here we combine genetic tagging of nuclei and ribosomes with RNA sequencing, chromatin immunoprecipitation with sequencing, assay for transposase-accessible chromatin using sequencing and Hi-C to investigate transcriptional and chromatin changes occurring in mouse hippocampal excitatory neurons at different time points after synchronous activation during seizure and sparse activation by novel context exploration. The transcriptional burst is associated with an increase in chromatin accessibility of activity-regulated genes and enhancers, de novo binding of activity-regulated transcription factors, augmented promoter–enhancer interactions and the formation of gene loops that bring together the transcription start site and transcription termination site of induced genes and may sustain the fast reloading of RNA polymerase complexes. Some chromatin occupancy changes and interactions, particularly those driven by AP1, remain long after neuronal activation and could underlie the changes in neuronal responsiveness and circuit connectivity observed in these neuroplasticity paradigms, perhaps thereby contributing to metaplasticity in the adult brain.
Studies of 3D chromatin organization have suggested that chromosomes are hierarchically organized into large compartments composed of smaller domains called topologically associating domains (TADs). Recent evidence suggests that compartments are smaller than previously thought and that the transcriptional or chromatin state is responsible for interactions leading to the formation of small compartmental domains in all organisms. In vertebrates, CTCF forms loop domains, probably via an extrusion process involving cohesin. CTCF loops cooperate with compartmental domains to establish the 3D organization of the genome. The continuous extrusion of the chromatin fibre by cohesin may also be responsible for the establishment of enhancer–promoter interactions and stochastic aspects of the transcription process. These observations suggest that the 3D organization of the genome is an emergent property of chromatin and its components, and thus may not be only a determinant but also a consequence of its function.
Pluripotent stem cells transition between distinct naive and primed states that are controlled by overlapping sets of master regulatory transcription factors. In this issue of Cell Stem Cell, Buecker et al. (2014) and Factor et al. (2014) demonstrate that alternate enhancer usage, regulated by state-specific binding partners of master regulators, defines these pluripotent state transitions.
Cells respond to temperature stress via up- and downregulation of hundreds of genes. This process is thought to be regulated by the heat shock factor HSF1, which controls the release of RNAPII from promoter-proximal pausing. Here, we analyze the events taking place in hESCs upstream of RNAPII release. We find that temperature stress results in the activation or decommissioning of thousands of enhancers. This process involves alterations in the occupancy of transcription factors HSF1, AP-1, NANOG, KLF4, and OCT4 accompanied by nucleosome remodeling by BRG1 and changes in H3K27ac. Furthermore, redistribution of RAD21 and CTCF results in the formation and disassembly of interactions mediated by these two proteins. These alterations tether and untether enhancers to their cognate promoters or refashion insulated neighborhoods, thus transforming the landscape of enhancer-promoter interactions. Details of the 3D interactome remodeling process support loop extrusion initiating at random sites as a mechanism for the establishment of CTCF/cohesin loops. Lyu et al. show that AP-1 and pluripotency factors play a critical role in the temperature stress response in hESCs. Recruitment of these factors to regulatory sequences is accompanied by changes in chromatin 3D organization mediated by CTCF and cohesin, which regulate the establishment of new enhancer-promoter interaction via a randomly initiated loop extrusion mechanism.
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Vishnu Dileep;
Korey A. Wilson;
Claire Marchal;
Xiaowen Lyu;
Peiyao A. Zhao;
Ben Li;
Axel Poulet;
Daniel A. Bartlett;
Juan Carlos Rivera-Mulia;
Zhaohui S. Qin;
Allan J. Robins;
Thomas C. Schulz;
Michael J. Kulik;
Rachel Patton McCord;
Job Dekker;
Stephen Dalton;
Victor Corces;
David M. Gilbert
The temporal order of DNA replication is regulated during development and is highly correlated with gene expression, histone modifications and 3D genome architecture. We tracked changes in replication timing, gene expression, and chromatin conformation capture (Hi-C) A/B compartments over the first two cell cycles during differentiation of human embryonic stem cells to definitive endoderm. Remarkably, transcriptional programs were irreversibly reprogrammed within the first cell cycle and were largely but not universally coordinated with replication timing changes. Moreover, changes in A/B compartment and several histone modifications that normally correlate strongly with replication timing showed weak correlation during the early cell cycles of differentiation but showed increased alignment in later differentiation stages and in terminally differentiated cell lines. Thus, epigenetic cell fate transitions during early differentiation can occur despite dynamic and discordant changes in otherwise highly correlated genomic properties. The temporal order of DNA replication is regulated during development, and is highly correlated with gene expression, chromatin structure, and 3D-genome architecture. Gilbert and colleagues find that stable transcriptional changes of human embryonic stem cells to endoderm can occur within a single cell cycle, accompanied by discordance in replication timing and chromatin compartment that align in later differentiation stages.
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Irene Gutierrez-Perez;
M. Jordan Rowley;
Xiaowen Lyu;
Viviana Valadez-Graham;
Diana M. Vallejo;
Esther Ballesta-Illan;
Jose P. Lopez-Atalaya;
Isaac Kremsky;
Esther Caparros;
Victor Corces;
Maria Dominguez
Gutierrez-Perez et al. show that BTB domain-containing isoforms of Pipsqueak associate with architectural proteins, whereas Psq lacking BTB colocalizes with Polycomb. Induction of differentiation by the hormone 20-hydroxyecdysone results in recruitment of the ecdysone receptor and Psq lacking BTB to enhancers and establishment of interactions with promoters of activated genes.
Transgenerational inheritance requires mechanisms by which epigenetic information is transferred via gametes. Canonical thought holds that mammalian sperm chromatin would be incapable of carrying epigenetic information as post-translational modifications of histones because of their replacement with protamine proteins. Furthermore, compaction of the sperm genome would hinder DNA accessibility of proteins involved in transcriptional regulation and genome architecture. In this Minireview, we delineate the paternal chromatin remodeling events during spermatogenesis and fertilization. Sperm chromatin is epigenetically modified at various time points throughout its development. This allows for the addition of environment-specific modifications that can be passed from parents to offspring.
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Jonathan A. Beagan;
Thomas G. Gilgenast;
Jesi Kim;
Zachary Plona;
Heidi K. Norton;
Gui Hu;
Sarah C. Hsu;
Emily J. Shields;
Xiaowen Lyu;
Effie Apostolou;
Konrad Hochedlinger;
Victor Corces;
Job Dekker;
Jennifer E. Phillips-Cremins
Pluripotent genomes are folded in a topological hierarchy that reorganizes during differentiation. The extent to which chromatin architecture is reconfigured during somatic cell reprogramming is poorly understood. Here we integrate fine-resolution architecture maps with epigenetic marks and gene expression in embryonic stem cells (ESCs), neural progenitor cells (NPCs), and NPC-derived induced pluripotent stem cells (iPSCs). We find that most pluripotency genes reconnect to target enhancers during reprogramming. Unexpectedly, some NPC interactions around pluripotency genes persist in our iPSC clone. Pluripotency genes engaged in both "fully-reprogrammed" and "persistent-NPC" interactions exhibit over/undershooting of target expression levels in iPSCs. Additionally, we identify a subset of "poorly reprogrammed" interactions that do not reconnect in iPSCs and display only partially recovered, ESC-specific CTCF occupancy. 2i/LIF can abrogate persistent-NPC interactions, recover poorly reprogrammed interactions, reinstate CTCF occupancy, and restore expression levels. Our results demonstrate that iPSC genomes can exhibit imperfectly rewired 3D-folding linked to inaccurately reprogrammed gene expression.