scholarly journals Loss of Cohesin regulator PDS5A reveals repressive role of Polycomb loops

2021 ◽  
Author(s):  
Daniel Bsteh ◽  
Hagar F Moussa ◽  
Georg Michlits ◽  
Ramesh Yelagandula ◽  
Jingkui Wang ◽  
...  
Keyword(s):  
Target Genes ◽  
Embryonic Stem ◽  
Transcriptional Silencing ◽  
Chromatin Modifications ◽  
3D Genome ◽  
Long Range Interactions ◽  
Crispr Screen ◽  
Epigenetic Regulators ◽  
Polycomb Repressive Complexes

Polycomb Repressive Complexes 1 and 2 (PRC1, PRC2) are conserved epigenetic regulators that promote transcriptional silencing. PRC1 and PRC2 converge on shared targets, catalyzing repressive histone modifications. In addition, a subset of PRC1/PRC2 targets engage in long-range interactions whose functions in gene silencing are poorly understood. Using a CRISPR screen in mouse embryonic stem cells, we identified that the cohesin regulator PDS5A links transcriptional silencing by Polycomb and 3D genome organization. PDS5A deletion impairs cohesin unloading and results in derepression of subset of endogenous PRC1/PRC2 target genes. Importantly, derepression is not associated with loss of repressive Polycomb chromatin modifications. Instead, loss of PDS5A leads to aberrant cohesin activity, ectopic insulation sites and specific reduction of ultra-long Polycomb loops. We infer that these loops are important for robust silencing at a subset of Polycomb target genes and that maintenance of cohesin-dependent genome architecture is critical for Polycomb regulation.

45 Expression patterns of PRDM family genes in porcine pre-implantation embryos

2020 ◽  
Vol 32 (2) ◽  
pp. 148
Author(s):  
K. Farrell ◽  
K. Uh ◽  
K. Lee
Keyword(s):  
Embryo Development ◽  
Internal Control ◽  
Target Genes ◽  
Expression Patterns ◽  
Embryonic Stem ◽  
Germinal Vesicle ◽  
Early Embryo ◽  
Cell Stage ◽  
Early Embryo Development

Establishing proper levels of pluripotency is essential for normal development. The genome of gametes is remodelled upon fertilisation and pluripotency-related genes are expressed in blastocysts. Multiple pluripotency-related genes are involved in the well-orchestrated process; however, detailed mechanistic actions remain elusive. The PRDM family genes are reported to be closely related to the pluripotency. A previous report noted that PRDM14 plays an important role in the maintenance of pluripotency in human embryonic stem cells (ESCs) and potentially murine ESCs; loss of PRDM14 was found to cause abnormalities in genome-wide epigenetic status. Similarly, PRDM15 was found to be a key regulator of pluripotency in mouse ESCs. Structural similarities among the PRDM family suggest that other PRDM family genes may help to establish and maintain pluripotency in embryos. Unfortunately, little is known about the expression profile of PRDM family in porcine embryos. To expand our understanding of the role of PRDM family in porcine embryos, expression patterns of PRDM gene family were investigated using reverse transcription quantitative (RTq)-PCR. Candidate PRDM family genes were selected based on previous RNA-Seq data in porcine oocytes/embryos. To conduct this study, germinal vesicle (GV), MII, zygote, 4-cell, and blastocyst samples were collected. Complementary DNA synthesised from the samples was used for RT-qPCR to analyse the expression pattern of selected PRDM family genes: PRDM2, PRDM4, PRDM6, PRDM14, and PRDM15. The expression of target genes was normalized to the YWHAG level, an internal control. Then, GV stage was used as a control for ΔΔCT analysis. Two technical replications and three biological replications were performed. Analysis of variance was used for statistical analysis and P-values<0.05 were considered significant. There was a significant decrease in PRDM2 expression in 4-cell and blastocyst, PRDM4 expression in 4-cell, and PRDM6 in all stages (MII, zygote, 4-cell, and blastocyst), compared with the GV stage. Because zygotic genome activation occurs at the 4-cell stage in the pig, the significant decrease in gene expression (PRDM2, PRDM4, and PRDM6) indicates they may be maternally originated and involved in the reprogramming process following fertilisation. On the other hand, there was a significant increase in PRDM15 expression in blastocysts and the PRDM14 transcript was only detected in blastocysts in all three biological replicates, suggesting that the genes are most likely involved in pluripotency maintenance, as was found in previous human studies. These results indicate that PRDM family genes are differentially expressed during early embryo development in pigs and may play a role in maintenance of pluripotency. For further study, we intend to evaluate the role of PRDM family genes during early embryo development in pigs.

scholarly journals Functional correlation of H3K9me2 and nuclear compartment formation

2020 ◽  
Author(s):  
Kei Fukuda ◽  
Chikako Shimura ◽  
Hisashi Miura ◽  
Akie Tanigawa ◽  
Takehiro Suzuki ◽  
...  
Keyword(s):  
Transcriptional Activation ◽  
Genome Organization ◽  
Mammalian Cells ◽  
Embryonic Stem ◽  
Epigenetic Mark ◽  
3 Dimensional ◽  
3D Genome ◽  
Functional Correlation ◽  
Genome Wide

AbstractBackgroundHistone H3 lysine 9 dimethylation (H3K9me2) is a highly conserved silencing epigenetic mark. Chromatin marked with H3K9me2 forms large domains in mammalian cells and correlates well with lamina-associated domains and the B compartment. However, the role of H3K9me2 in 3-dimensional (3D) genome organization remains unclear.ResultsWe investigated the genome-wide H3K9me2 distribution, the transcriptome and 3D genome organization in mouse embryonic stem cells (mESCs) upon the inhibition or depletion of H3K9 methyltransferases (MTases) G9a/GLP, SETDB1, and SUV39H1/2. We found that H3K9me2 is regulated by these five MTases; however, H3K9me2 and transcription in the A and B compartments were largely regulated by different sets of the MTases: H3K9me2 in the A compartments were mainly regulated by G9a/GLP and SETDB1, while H3K9me2 in the B compartments were regulated by all five H3K9 MTases. Furthermore, decreased H3K9me2 correlated with the changes to the more active compartmental state that accompanied transcriptional activation.ConclusionOur data showed that H3K9me2 domain formation is functionally linked to 3D genome organization.

scholarly journals EOMES is responsible for WNT memory and can substitute for WNT in mesendoderm specification

2021 ◽  
Author(s):  
Anna Yoney ◽  
Lu Bai ◽  
Ali H. Brivanlou ◽  
Eric D Siggia
Keyword(s):  
Target Genes ◽  
Embryonic Stem ◽  
Model Systems ◽  
Mechanistic Basis ◽  
Vitro Model ◽  
Transcriptional Output ◽  
Accessible Chromatin ◽  
Pluripotency Maintenance

Embryogenesis is guided by a limited set of signaling pathways that are reused at different times and places throughout development. How a context dependent signaling response is generated has been a central question of developmental biology, which can now be addressed with in vitro model systems. Our previous work in human embryonic stem cells (hESCs) established that pre-exposure of cells to WNT/β-catenin signaling is sufficient to switch the output of ACTIVIN/SMAD2 signaling from pluripotency maintenance to mesendoderm (ME) differentiation. A body of previous literature has established the role of both pathways in ME differentiation. However, our work demonstrated that the two signals do not need to be present simultaneously and that hESCs have a means to record WNT signals. Here we demonstrate that hESCs have accessible chromatin at SMAD2 binding sites near pluripotency and ME-associated target genes and that WNT priming does not alter SMAD2 binding. Rather our results indicate that stable transcriptional output at ME genes results from WNT-dependent production of an additional SMAD2 co-factor, EOMES. We show that expression of EOMES can replace WNT signaling in ME differentiation, providing a mechanistic basis for WNT-priming and memory in early development.

scholarly journals DNA methylation directs polycomb-dependent 3D genome re- organisation in naive pluripotency

2019 ◽  
Author(s):  
Katy A McLaughlin ◽  
Ilya M Flyamer ◽  
John P Thomson ◽  
Heidi K Mjoseng ◽  
Ruchi Shukla ◽  
...  
Keyword(s):  
Dna Methylation ◽  
Stem Cells ◽  
Gene Regulation ◽  
Ground State ◽  
Embryonic Stem ◽  
Genome Organisation ◽  
Dna Hypomethylation ◽  
3D Genome ◽  
And Function

The DNA hypomethylation that occurs when embryonic stem cells (ESCs) are directed to the ground state of naive pluripotency by culturing in 2i conditions results in redistribution of polycomb (H3K27me3) away from its target loci. Here we demonstrate that 3D genome organisation is also altered in 2i. We found chromatin decompaction at polycomb target loci as well as loss of long-range polycomb interactions. By preventing DNA hypomethylation during the transition to the ground-state, we are able to restore the H3K27me3 distribution, and polycomb-mediated 3D genome organisation that is characteristic of primed ESCs grown in serum, to ESCs in 2i. However, these cells retain the functional characteristics of 2i ground state ESCs. Our findings demonstrate the central role of DNA methylation in shaping major aspects of 3D genome organisation but caution against assuming causal roles for the epigenome and 3D genome in gene regulation and function in ESCs.

scholarly journals Relaxed 3D genome conformation facilitates the pluripotent to totipotent state transition in embryonic stem cells

2020 ◽  
Author(s):  
Jiali Yu ◽  
Yezhang Zhu ◽  
Jiahui Gu ◽  
Chaoran Xue ◽  
Long Zhang ◽  
...  
Keyword(s):  
Stem Cells ◽  
Embryonic Stem Cells ◽  
Genome Organization ◽  
Critical Role ◽  
Embryonic Stem ◽  
List Type ◽  
Down Regulation ◽  
Chromatin Loops ◽  
3D Genome

SUMMARYThe 3D genome organization is crucial for gene regulation. Although recent studies have revealed a uniquely relaxed genome conformation in totipotent early blastmeres of both fertilized and cloned embryos, how weakened higher-order chromatin structure is functionally linked to totipotency acquisition remains elusive. Using low-input Hi-C, ATAC-seq, and ChIP-seq, we systematically examined the dynamics of 3D genome and epigenome during pluripotency-to-totipotency transition in mouse embryonic stem cells (ESCs). The totipotent 2-cell-embro-like cells (2CLCs) exhibit more relaxed chromatin architecture compared to ESCs, including global weakening of both enhancer-promoter interactions and TAD insulation. While the former leads to inactivation of ESC enhancers and down-regulation of pluripotent genes, the latter may facilitate contacts between the new enhancers arising in 2CLCs and neighboring 2C genes. Importantly, disruption of chromatin loops by depleting CTCF or cohesin promotes ESC to 2CLC transition. Our results thus establish a critical role of 3D genome organization in totipotency acquisition.HIGHLIGHTSGlobal weakening of the 3D genome conformation during ESC to 2CLC transitionLoss of enhancer-promoter loops and down-regulation of pluripotent genes in 2CLCsInactivation of ESC enhancers and formation of new enhancers in 2CLCsDisruption of chromatin loops by depleting CTCF or cohesin promotes 2CLC emergence

Abstract 139: Role of Histone Lysine Demethylase PHF8 in Epigenetic Regulation of Embryonic Stem Cell Cardiac Differentiation

2016 ◽  
Vol 119 (suppl_1) ◽  
Author(s):  
Yan Tang ◽  
Yujia Yue ◽  
Suresh K Verma ◽  
Mohsin Khan ◽  
Raj Kishore
Keyword(s):  
Progenitor Cells ◽  
Expression Profiles ◽  
Embryonic Stem ◽  
Cardiac Differentiation ◽  
Lineage Commitment ◽  
Epigenetic Control ◽  
Lineage Decision ◽  
Cardiac Lineage ◽  
Epigenetic Regulators

Epigenetic control mechanisms play a key role in the regulation of lineage commitment of stem/progenitor cells, while the epigenetic regulators involved in the determination of cardiomyogenic lineage are incompletely defined. Using in vitro cardiac differentiation system of mESCs with Brachyury and Nkx2.5 selection, we analyzed expression profiles of epigenetic regulators at critical stages of cardiomyogenesis by RT 2 profiler PCR arrays. To identify the potential epigenetic regulators in the cardiac lineage decision, we compared their expression levels in Brachyury + mesodermal cells with Brachyury - cells, and in Nkx2.5 + cardiac progenitor cells with the Nkx2.5 - cells, respectively. To further understand the role of these epigenetic regulators in cardiac lineage commitment, we knockout these genes and investigate their function. For example, deletion of the H3K9me2 demethylase PHF8 in mESCs did not affect self-renewal, proliferation or early ectodermal/endodermal differentiation, but it did promote the mesodermal lineage commitment with the enhanced cardiomyocyte differentiation. The effects were accompanied by a reduction in apoptosis, without significant differences between differentiating wide-type (ph8 +/Y ) and ph8 -/Y ESCs in cell cycle progression or proliferation. Functionally, PHF8 promoted the loss of a repressive mark H3K9me2 from the transcription start site of a proapoptotic gene pmaip1 and activated its transcription. These results reveal the new epigenetic control mechanism in mesodemal and cardiac differentiation and establish a link between the apoptosis and cell lineage decision as well as cardiogenesis.

scholarly journals Regulation of mammalian 3D genome organization and histone H3K9 dimethylation by H3K9 methyltransferases

2021 ◽  
Vol 4 (1) ◽  
Author(s):  
Kei Fukuda ◽  
Chikako Shimura ◽  
Hisashi Miura ◽  
Akie Tanigawa ◽  
Takehiro Suzuki ◽  
...  
Keyword(s):  
Transcriptional Activation ◽  
Genome Organization ◽  
Mammalian Cells ◽  
Embryonic Stem ◽  
Epigenetic Mark ◽  
3 Dimensional ◽  
3D Genome ◽  
Genome Wide ◽  
H3k9 Dimethylation

AbstractHistone H3 lysine 9 dimethylation (H3K9me2) is a highly conserved silencing epigenetic mark. Chromatin marked with H3K9me2 forms large domains in mammalian cells and overlaps well with lamina-associated domains and the B compartment defined by Hi-C. However, the role of H3K9me2 in 3-dimensional (3D) genome organization remains unclear. Here, we investigated genome-wide H3K9me2 distribution, transcriptome, and 3D genome organization in mouse embryonic stem cells following the inhibition or depletion of H3K9 methyltransferases (MTases): G9a, GLP, SETDB1, SUV39H1, and SUV39H2. We show that H3K9me2 is regulated by all five MTases; however, H3K9me2 and transcription in the A and B compartments are regulated by different MTases. H3K9me2 in the A compartments is primarily regulated by G9a/GLP and SETDB1, while H3K9me2 in the B compartments is regulated by all five MTases. Furthermore, decreased H3K9me2 correlates with changes to more active compartmental state that accompanied transcriptional activation. Thus, H3K9me2 contributes to inactive compartment setting.

scholarly journals Variant PCGF1-PRC1 links PRC2 recruitment with differentiation-associated transcriptional inactivation at target genes

2021 ◽  
Vol 12 (1) ◽  
Author(s):  
Hiroki Sugishita ◽  
Takashi Kondo ◽  
Shinsuke Ito ◽  
Manabu Nakayama ◽  
Nayuta Yakushiji-Kaminatsui ◽  
...  
Keyword(s):  
Stem Cells ◽  
Crucial Role ◽  
Target Genes ◽  
Embryonic Stem ◽  
Embryoid Bodies ◽  
Polycomb Group ◽  
Gene Repression ◽  
Down Regulation ◽  
Aberrant Expression ◽  
Polycomb Repressive Complexes

AbstractPolycomb repressive complexes-1 and -2 (PRC1 and 2) silence developmental genes in a spatiotemporal manner during embryogenesis. How Polycomb group (PcG) proteins orchestrate down-regulation of target genes upon differentiation, however, remains elusive. Here, by differentiating embryonic stem cells into embryoid bodies, we reveal a crucial role for the PCGF1-containing variant PRC1 complex (PCGF1-PRC1) to mediate differentiation-associated down-regulation of a group of genes. Upon differentiation cues, transcription is down-regulated at these genes, in association with PCGF1-PRC1-mediated deposition of histone H2AK119 mono-ubiquitination (H2AK119ub1) and PRC2 recruitment. In the absence of PCGF1-PRC1, both H2AK119ub1 deposition and PRC2 recruitment are disrupted, leading to aberrant expression of target genes. PCGF1-PRC1 is, therefore, required for initiation and consolidation of PcG-mediated gene repression during differentiation.

scholarly journals PHF19 mediated regulation of proliferation and invasiveness in prostate cancer cells

eLife ◽  
2020 ◽  
Vol 9 ◽  
Author(s):  
Payal Jain ◽  
Cecilia Ballare ◽  
Enrique Blanco ◽  
Pedro Vizan ◽  
Luciano Di Croce
Keyword(s):  
Prostate Cancer ◽  
Cell Proliferation ◽  
Cancer Cells ◽  
Target Genes ◽  
Embryonic Stem ◽  
Prostate Cancer Cells ◽  
Genome Wide ◽  
A Genome ◽  
Matrix Organization

The Polycomb-like protein PHF19/PCL3 associates with PRC2 and mediates its recruitment to chromatin in embryonic stem cells. PHF19 is also overexpressed in many cancers. However, neither PHF19 targets nor misregulated pathways involving PHF19 are known. Here, we investigate the role of PHF19 in prostate cancer cells. We find that PHF19 interacts with PRC2 and binds to PRC2 targets on chromatin. PHF19 target genes are involved in proliferation, differentiation, angiogenesis, and extracellular matrix organization. Depletion of PHF19 triggers an increase in MTF2/PCL2 chromatin recruitment, with a genome-wide gain in PRC2 occupancy and H3K27me3 deposition. Transcriptome analysis shows that PHF19 loss promotes deregulation of key genes involved in growth, metastasis, invasion, and of factors that stimulate blood vessels formation. Consistent with this, PHF19 silencing reduces cell proliferation, while promotes invasive growth and angiogenesis. Our findings reveal a role for PHF19 in controlling the balance between cell proliferation and invasiveness in prostate cancer.

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