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Testicular germ cell tumors (TGCTs) can be treated with cisplatin-based therapy. However, a clinically significant number of cisplatin-resistant patients die from progressive disease as no effective alternatives exist. Curative cisplatin therapy results in acute and life-long toxicities in the young TGCT patient population providing a rationale to decrease cisplatin exposure. In contrast to genetic alterations, recent evidence suggests that epigenetics is a major driving factor for TGCT formation, progression, and response to chemotherapy. Hence, targeting epigenetic pathways with "epidrugs" is one potential relatively unexplored strategy to advance TGCT treatment beyond cisplatin. In this report, we demonstrate for the first time that targeting polycomb demethylases KDM6A and KDM6B with epidrug GSK-J4 can treat both cisplatin-sensitive and -resistant TGCTs. While GSK-J4 had minimal effects alone on TGCT tumor growth in vivo, it dramatically sensitized cisplatin-sensitive and -resistant TGCTs to cisplatin. We validated KDM6A/KDM6B as the target of GSK-J4 since KDM6A/KDM6B genetic depletion had a similar effect to GSK-J4 on cisplatin-mediated anti-tumor activity and transcriptome alterations. Pharmacologic and genetic targeting of KDM6A/KDM6B potentiated or primed the p53-dominant transcriptional response to cisplatin, with also evidence for basal activation of p53. Further, several chromatin modifier genes, including BRD4, lysine demethylases, chromodomain helicase DNA binding proteins, and lysine methyltransferases, were repressed with cisplatin only in KDM6A/KDM6B-targeted cells, implying that KDM6A/KDM6B inhibition sets the stage for extensive chromatin remodeling of TGCT cells upon cisplatin treatment. Our findings demonstrate that targeting polycomb demethylases is a new potent pharmacologic strategy for treating cisplatin resistant TGCTs that warrants clinical development. [ABSTRACT FROM AUTHOR]

Polycomb proteins are a fundamental repressive system that plays crucial developmental roles by orchestrating cell-type-specific transcription programs that govern cell identity. Direct alterations of Polycomb activity are indeed implicated in human pathologies, including developmental disorders and cancer. General Polycomb repression is coordinated by three distinct activities that regulate the deposition of two histone post-translational modifications: tri-methylation of histone H3 lysine 27 (H3K27me3) and histone H2A at lysine 119 (H2AK119ub1). These activities exist in large and heterogeneous multiprotein ensembles consisting of common enzymatic cores regulated by heterogeneous non-catalytic modules composed of a large number of accessory proteins with diverse biochemical properties. Here, we have analyzed the current molecular knowledge, focusing on the functional interaction between the core enzymatic activities and their regulation mediated by distinct accessory modules. This provides a comprehensive analysis of the molecular details that control the establishment and maintenance of Polycomb repression, examining their underlying coordination and highlighting missing information and emerging new features of Polycomb-mediated transcriptional control. The Polycomb machinery is a major repressive system essential for development, stem cell, and tissue homeostasis directly involved in several human pathologies. Tamburri et al. explored the molecular complexity by which different core enzymatic activities and regulatory modules coordinate to establish and maintain repressive domains to control cell transcriptional identities. [ABSTRACT FROM AUTHOR]

Abstract Testicular germ cell tumors (TGCTs) can be treated with cisplatin-based therapy. However, a clinically significant number of cisplatin-resistant patients die from progressive disease as no effective alternatives exist. Curative cisplatin therapy results in acute and life-long toxicities in the young TGCT patient population providing a rationale to decrease cisplatin exposure. In contrast to genetic alterations, recent evidence suggests that epigenetics is a major driving factor for TGCT formation, progression, and response to chemotherapy. Hence, targeting epigenetic pathways with “epidrugs” is one potential relatively unexplored strategy to advance TGCT treatment beyond cisplatin. In this report, we demonstrate for the first time that targeting polycomb demethylases KDM6A and KDM6B with epidrug GSK-J4 can treat both cisplatin-sensitive and -resistant TGCTs. While GSK-J4 had minimal effects alone on TGCT tumor growth in vivo, it dramatically sensitized cisplatin-sensitive and -resistant TGCTs to cisplatin. We validated KDM6A/KDM6B as the target of GSK-J4 since KDM6A/KDM6B genetic depletion had a similar effect to GSK-J4 on cisplatin-mediated anti-tumor activity and transcriptome alterations. Pharmacologic and genetic targeting of KDM6A/KDM6B potentiated or primed the p53-dominant transcriptional response to cisplatin, with also evidence for basal activation of p53. Further, several chromatin modifier genes, including BRD4, lysine demethylases, chromodomain helicase DNA binding proteins, and lysine methyltransferases, were repressed with cisplatin only in KDM6A/KDM6B-targeted cells, implying that KDM6A/KDM6B inhibition sets the stage for extensive chromatin remodeling of TGCT cells upon cisplatin treatment. Our findings demonstrate that targeting polycomb demethylases is a new potent pharmacologic strategy for treating cisplatin resistant TGCTs that warrants clinical development.

Heterochromatin is a gene-repressive protein-nucleic acid ultrastructure that is initially nucleated by DNA sequences. However, following nucleation, heterochromatin can then propagate along the chromatin template in a sequence-independent manner in a reaction termed spreading. At the heart of this process are enzymes that deposit chemical information on chromatin, which attracts the factors that execute chromatin compaction and transcriptional or co/post-transcriptional gene silencing. Given that these enzymes deposit guiding chemical information on chromatin they are commonly termed writers. While the processes of nucleation and central actions of writers have been extensively studied and reviewed, less is understood about how the spreading process is regulated. We discuss how the chromatin substrate is prepared for heterochromatic spreading, and how trans-acting factors beyond writer enzymes regulate it. We examine mechanisms by which trans-acting factors in Suv39, PRC2, SETDB1 and SIR writer systems regulate spreading of the respective heterochromatic marks across chromatin. While these systems are in some cases evolutionarily and mechanistically quite distant, common mechanisms emerge which these trans-acting factors exploit to tune the spreading reaction.

The ovarian reserve defines female reproductive lifespan, which in humans spans decades. The ovarian reserve consists of oocytes residing in primordial follicles arrested in meiotic prophase I and is maintained independent of DNA replication and cell proliferation, thereby lacking stem cell-based maintenance. Largely unknown is how cellular states of the ovarian reserve are established and maintained for decades. Our recent study revealed that a distinct chromatin state is established during ovarian reserve formation in mice, uncovering a novel window of epigenetic programming in female germline development. We showed that an epigenetic regulator, Polycomb Repressive Complex 1 (PRC1), establishes a repressive chromatin state in perinatal mouse oocytes that is essential for prophase I-arrested oocytes to form the ovarian reserve. Here we discuss the biological roles and mechanisms underlying epigenetic programming in ovarian reserve formation, highlighting current knowledge gaps and emerging research areas in female reproductive biology.

Proneural gliomas are brain tumors characterized by enrichment of oligodendrocyte progenitor cell (OPC) transcripts and genetic alterations. In this study we sought to identify transcriptional and epigenetic differences between OPCs with Trp53 deletion and PDGF-BB overexpression (BB-p53n) and those carrying only p53 deletion (p53n). In culture, the BB-p53n OPCs display growth characteristics more similar to glioma cells than p53n OPCs. When injected in mouse brains, BB-p53n OPC form tumors, while the p53n OPCs do not. Unbiased histone proteomics and transcriptomic analysis on these OPC populations identified higher levels of the histone H3K27me3 mark and lower levels of the histone H4K20me3. The transcriptome of the BB-p53n OPCs was characterized by higher levels of transcripts related to proliferation and cell adhesion compared to p53n OPCs. Pharmacological inhibition of the enzyme responsible for histone H3K27 trimethylation (EZH2i) in BB-p53n OPCs, reduced cell cycle transcripts and increased the expression of differentiation markers, but was not sufficient to restore their growth characteristics. This suggests that PDGF-BB overexpression in p53n OPCs favors the early stages of transformation, by promoting proliferation and halting differentiation in a H3K27me3-dependent pathway, and favoring growth characteristics in a H3K27me3 independent manner

During early development, sea lamprey embryos undergo programmatic elimination of DNA from somatic progenitor cells in a process termed programmed genome rearrangement (PGR). Eliminated DNA eventually becomes condensed into micronuclei, which are then physically degraded and permanently lost from the cell. Previous studies indicated that many of the genes eliminated during PGR have mammalian homologs that are bound by polycomb repressive complex (PRC) in embryonic stem cells. To test whether PRC components play a role in the faithful elimination of germline‐specific sequences, we used a combination of CRISPR/Cas9 and lightsheet microscopy to investigate the impact of gene knockouts on early development and the progression through stages of DNA elimination. Analysis of knockout embryos for the core PRC2 subunits EZH, SUZ12, and EED show that disruption of all three genes results in an increase in micronucleus number, altered distribution of micronuclei within embryos, and an increase in micronucleus volume in mutant embryos. While the upstream events of DNA elimination are not strongly impacted by loss of PRC2 components, this study suggests that PRC2 plays a role in the later stages of elimination related to micronucleus condensation and degradation. These findings also suggest that other genes/epigenetic pathways may work in parallel during DNA elimination to mediate chromatin structure, accessibility, and the ultimate loss of germline‐specific DNA. Research Highlights: Cas9 was used efficiently mutate core PRC2 genes EZH, SUZ12, and EED.Whole embryo imaging revealed impacts on micronucleus distribution and compaction during later stages of DNA loss.Initial targeting of DNA for elimination is likely upstream of PRC2. [ABSTRACT FROM AUTHOR]

When exposed to low temperatures, plants undergo a drastic reprogramming of their transcriptome in order to adapt to their new environmental conditions, which primes them for potential freezing temperatures. While the involvement of transcription factors in this process, termed cold acclimation, has been deeply investigated, the potential contribution of chromatin regulation remains largely unclear. A large proportion of cold-inducible genes carries the repressive mark histone 3 lysine 27 trimethylation (H3K27me3), which has been hypothesized as maintaining them in a silenced state in the absence of stress, but which would need to be removed or counteracted upon stress perception. However, the fate of H3K27me3 during cold exposure has not been studied genome-wide. In this study, we offer an epigenome profiling of H3K27me3 and its antagonistic active mark H3K4me3 during short-term cold exposure. Both chromatin marks undergo rapid redistribution upon cold exposure, however, the gene sets undergoing H3K4me3 or H3K27me3 differential methylation are distinct, refuting the simplistic idea that gene activation relies on a switch from an H3K27me3 repressed chromatin to an active form enriched in H3K4me3. Coupling the ChIP-seq experiments with transcriptome profiling reveals that differential histone methylation only weakly correlates with changes in expression. Interestingly, only a subset of cold-regulated genes lose H3K27me3 during their induction, indicating that H3K27me3 is not an obstacle to transcriptional activation. In the H3K27me3 methyltransferase curly leaf (clf) mutant, many cold regulated genes display reduced H3K27me3 levels but their transcriptional activity is not altered prior or during a cold exposure, suggesting that H3K27me3 may serve a more intricate role in the cold response than simply repressing the cold-inducible genes in naïve conditions. [ABSTRACT FROM AUTHOR]

Polycomb group (PcG) proteins are a subset of epigenetic factors that are highly conserved throughout evolution. In mammals, PcG proteins can be classified into two muti-proteins complexes: Polycomb repressive complex 1 (PRC1) and PRC2. Increasing evidence has demonstrated that PcG complexes play critical roles in the regulation of gene expression, genomic imprinting, chromosome X-inactivation, and chromatin structure. Accordingly, the dysfunction of PcG proteins is tightly orchestrated with abnormal developmental processes. Here, we summarized and discussed the current knowledge of the biochemical and molecular functions of PcG complexes, especially the PRC1 and PRC2 in mammalian development including embryonic development and tissue development, which will shed further light on the deep understanding of the basic knowledge of PcGs and their functions for reproductive health and developmental disorders. [ABSTRACT FROM AUTHOR]

Epigenetic processes involving long non-coding RNAs regulate endothelial gene expression. However, the underlying regulatory mechanisms causing endothelial dysfunction remain to be elucidated. Enhancer of zeste homolog 2 (EZH2) is an important rheostat of histone H3K27 trimethylation (H3K27me3) that represses endothelial targets, but EZH2 RNA binding capacity and EZH2:RNA functional interactions have not been explored in post-ischemic angiogenesis. We used formaldehyde/UV-assisted crosslinking ligation and sequencing of hybrids and identified a new role for maternally expressed gene 3 (MEG3). MEG3 formed the predominant RNA:RNA hybrid structures in endothelial cells. Moreover, MEG3:EZH2 assists recruitment onto chromatin. By EZH2-chromatin immunoprecipitation, following MEG3 depletion, we demonstrated that MEG3 controls recruitment of EZH2/H3K27me3 onto integrin subunit alpha4 (ITGA4) promoter. Both MEG3 knockdown or EZH2 inhibition (A-395) promoted ITGA4 expression and improved endothelial cell migration and adhesion to fibronectin in vitro. The A-395 inhibitor re-directed MEG3-assisted chromatin remodeling, offering a direct therapeutic benefit by increasing endothelial function and resilience. This approach subsequently increased the expression of ITGA4 in arterioles following ischemic injury in mice, thus promoting arteriogenesis. Our findings show a context-specific role for MEG3 in guiding EZH2 to repress ITGA4. Novel therapeutic strategies could antagonize MEG3:EZH2 interaction for pre-clinical studies.

Light is a crucial environmental factor that impacts various aspects of plant development. Phytochromes, as light sensors, regulate myriads of downstream genes to mediate developmental reprogramming in response to changes in environmental conditions. CONSTITUTIVELY PHOTOMORPHOGENIC 1 (COP1) is an E3 ligase for a number of substrates in light signaling, acting as a central repressor of photomorphogenesis. The interplay between phytochrome B (phyB) and COP1 forms an antagonistic regulatory module that triggers extensive gene expression reprogramming when exposed to light. Here, we uncover a role of COP1 in light-dependent chromatin remodeling through the regulation of VIL1 (VIN3-LIKE 1)/VERNALIZATION 5, a Polycomb protein. VIL1 directly interacts with phyB and regulates photomorphogenesis through the formation of repressive chromatin loops at downstream growth-promoting genes in response to light. Furthermore, we reveal that COP1 governs light-dependent formation of chromatin loop and limiting a repressive histone modification to fine-tune expressions of growth-promoting genes during photomorphogenesis through VIL1. [ABSTRACT FROM AUTHOR]

Summary: We are becoming aware of a growing number of organisms that do not express genetic information equally from both parents as a result of an epigenetic phenomenon called genomic imprinting. Recently, it was shown that the entire paternal genome is repressed during the diploid phase of the life cycle of the liverwort Marchantia polymorpha. The deposition of the repressive epigenetic mark H3K27me3 on the male pronucleus is responsible for the imprinted state, which is reset by the end of meiosis. Here, we put these recent reports in perspective of other forms of imprinting and discuss the potential mechanisms of imprinting in bryophytes and the causes of its evolution. [ABSTRACT FROM AUTHOR]

The Drosophila PRC1 complex regulates gene expression by modifying histone proteins and chromatin architecture. Two PRC1 subunits, PSC and Ph, are most implicated in chromatin architecture. In vitro, PRC1 compacts chromatin and inhibits transcription and nucleosome remodeling. The long disordered C-terminal region of PSC (PSC-CTR) is important for these activities, while Ph has little effect. In cells, Ph is important for condensate formation, long-range chromatin interactions, and gene regulation, and its polymerizing sterile alpha motif (SAM) is implicated in these activities. In vitro, truncated Ph containing the SAM and two other conserved domains (mini-Ph) undergoes phase separation with chromatin, suggesting a mechanism for SAM-dependent condensate formation in vivo. How the distinct activities of PSC and Ph on chromatin function together in PRC1 is not known. To address this question, we analyzed structures formed with large chromatin templates and PRC1 in vitro. PRC1 bridges chromatin into extensive fibrillar networks. Ph, its SAM, and SAM polymerization activity have little effect on these structures. Instead, the PSC-CTR controls their growth, and is sufficient for their formation. To understand how phase separation driven by Ph SAM intersects with the chromatin bridging activity of the PSC-CTR, we used mini-Ph to form condensates with chromatin and then challenged them with PRC1 lacking Ph (PRC1ΔPh). PRC1ΔPh converts mini-Ph chromatin condensates into clusters of small non-fusing condensates and bridged fibers. These condensates retain a high level of chromatin compaction and do not intermix. Thus, phase separation of chromatin by mini-Ph, followed by the action of the PSC-CTR, creates a unique chromatin organization with regions of high nucleosome density and extraordinary stability. We discuss how this coordinated sequential activity of two proteins found in the same complex may occur and the possible implications of stable chromatin architectures in maintaining transcription states. [ABSTRACT FROM AUTHOR]

Since their discovery, the Hox genes, with their incredible power to reprogram the identity of complete body regions, a phenomenon called homeosis, have captured the fascination of many biologists. Recent research has provided new insights into the function of Hox proteins in different germ layers and the mechanisms they employ to control tissue morphogenesis. We focus in this review on the ectoderm and mesoderm to highlight new findings and discuss them with regards to established concepts of Hox target gene regulation. Furthermore, we highlight the molecular mechanisms involved the transcriptional repression of specific groups of Hox target genes, and summarize the role of Hox mediated gene silencing in tissue development. Finally, we reflect on recent findings identifying a large number of tissue-specific Hox interactor partners, which open up new avenues and directions towards a better understanding of Hox function and specificity in different tissues. [ABSTRACT FROM AUTHOR]

Germline epigenetic programming, including genomic imprinting, substantially influences offspring development. Polycomb Repressive Complex 2 (PRC2) plays an important role in Histone 3 Lysine 27 trimethylation (H3K27me3)-dependent imprinting, loss of which leads to growth and developmental changes in mouse offspring. In this study, we show that offspring from mouse oocytes lacking the PRC2 protein Embryonic Ectoderm Development (EED) were initially developmentally delayed, characterised by low blastocyst cell counts and substantial growth delay in mid-gestation embryos. This initial developmental delay was resolved as offspring underwent accelerated fetal development and growth in late gestation resulting in offspring that were similar stage and weight to controls at birth. The accelerated development and growth in offspring from Eed-null oocytes was associated with remodelling of the placenta, which involved an increase in fetal and maternal tissue size, conspicuous expansion of the glycogen-enriched cell population, and delayed parturition. Despite placental remodelling and accelerated offspring fetal growth and development, placental efficiency, and fetal blood glucose levels were low, and the fetal blood metabolome was unchanged. Moreover, while expression of the H3K27me3-imprinted gene and amino acid transporter Slc38a4 was increased, fetal blood levels of individual amino acids were similar to controls, indicating that placental amino acid transport was not enhanced. Genome-wide analyses identified extensive transcriptional dysregulation and DNA methylation changes in affected placentas, including a range of imprinted and non-imprinted genes. Together, while deletion of Eed in growing oocytes resulted in fetal growth and developmental delay and placental hyperplasia, our data indicate a remarkable capacity for offspring fetal growth to be normalised despite inefficient placental function and the loss of H3K27me3-dependent genomic imprinting.

When exposed to low temperatures, plants undergo a drastic reprogramming of their transcriptome in order to adapt to their new environmental conditions, which primes them for potential freezing temperatures. While the involvement of transcription factors in this process, termed cold acclimation, has been deeply investigated, the potential contribution of chromatin regulation remains largely unclear. A large proportion of cold-inducible genes carries the repressive mark histone 3 lysine 27 trimethylation (H3K27me3), which has been hypothesized as maintaining them in a silenced state in the absence of stress, but which would need to be removed or counteracted upon stress perception. However, the fate of H3K27me3 during cold exposure has not been studied genome-wide. In this study, we offer an epigenome profiling of H3K27me3 and its antagonistic active mark H3K4me3 during short-term cold exposure. Both chromatin marks undergo rapid redistribution upon cold exposure, however, the gene sets undergoing H3K4me3 or H3K27me3 differential methylation are distinct, refuting the simplistic idea that gene activation relies on a switch from an H3K27me3 repressed chromatin to an active form enriched in H3K4me3. Coupling the ChIP-seq experiments with transcriptome profiling reveals that differential histone methylation only weakly correlates with changes in expression. Interestingly, only a subset of cold-regulated genes lose H3K27me3 during their induction, indicating that H3K27me3 is not an obstacle to transcriptional activation. In the H3K27me3 methyltransferase curly leaf (clf) mutant, many cold regulated genes display reduced H3K27me3 levels but their transcriptional activity is not altered prior or during a cold exposure, suggesting that H3K27me3 may serve a more intricate role in the cold response than simply repressing the cold-inducible genes in naïve conditions.

A fundamental goal of developmental biology is to understand how cell and tissue fates are specified. The imaginal discs of Drosophila are excellent model systems for addressing this paradigm as their fate can be redirected when discs regenerate after injury or when key selector genes are misregulated. Here, we show that when Polycomb expression is reduced, the wing selector gene vestigial is ectopically activated. This leads to the inappropriate formation of the Vestigial-Scalloped complex, which forces the eye to transform into a wing. We further demonstrate that disrupting this complex does not simply block wing formation or restore eye development. Instead, immunohistochemistry and high-throughput genomic analysis show that the eye-antennal disc unexpectedly undergoes hyperplastic growth with multiple domains being organized into other imaginal discs and tissues. These findings provide insight into the complex developmental landscape that tissues must navigate before adopting their final fate. [ABSTRACT FROM AUTHOR]

Heterochromatin is a gene-repressive protein–nucleic acid ultrastructure that is initially nucleated by DNA sequences. However, following nucleation, heterochromatin can then propagate along the chromatin template in a sequence-independent manner in a reaction termed spreading. At the heart of this process are enzymes that deposit chemical information on chromatin, which attracts the factors that execute chromatin compaction and transcriptional or co/post-transcriptional gene silencing. Given that these enzymes deposit guiding chemical information on chromatin they are commonly termed ‘writers’. While the processes of nucleation and central actions of writers have been extensively studied and reviewed, less is understood about how the spreading process is regulated. We discuss how the chromatin substrate is prepared for heterochromatic spreading, and how trans-acting factors beyond writer enzymes regulate it. We examine mechanisms by which trans-acting factors in Suv39, PRC2, SETDB1 and SIR writer systems regulate spreading of the respective heterochromatic marks across chromatin. While these systems are in some cases evolutionarily and mechanistically quite distant, common mechanisms emerge which these trans-acting factors exploit to tune the spreading reaction.

The chromatin of animal cells contains two types of histones: canonical histones that are expressed during S phase of the cell cycle to package the newly replicated genome, and variant histones with specialized functions that are expressed throughout the cell cycle and in nonproliferating cells. Determining whether and how canonical and variant histones cooperate to regulate genome function is integral to understanding how chromatin-based processes affect normal and pathological development. Here, we demonstrate that variant histone H3.3 is essential for Drosophila development only when canonical histone gene copy number is reduced, suggesting that coordination between canonical H3.2 and variant H3.3 expression is necessary to provide sufficient H3 protein for normal genome function. To identify genes that depend upon, or are involved in, this coordinate regulation we screened for heterozygous chromosome 3 deficiencies that impair development of flies bearing reduced H3.2 and H3.3 gene copy number. We identified two regions of chromosome 3 that conferred this phenotype, one of which contains the Polycomb gene, which is necessary for establishing domains of facultative chromatin that repress master regulator genes during development. We further found that reduction in Polycomb dosage decreases viability of animals with no H3.3 gene copies. Moreover, heterozygous Polycomb mutations result in de-repression of the Polycomb target gene Ubx and cause ectopic sex combs when either canonical or variant H3 gene copy number is reduced. We conclude that Polycomb-mediated facultative heterochromatin function is compromised when canonical and variant H3 gene copy number falls below a critical threshold. [ABSTRACT FROM AUTHOR]

Polycomb group (PcG) proteins maintain the silenced state of key developmental genes, but how these proteins are recruited to specific regions of the genome is still not completely understood. In Drosophila, PcG proteins are recruited to Polycomb response elements (PREs) comprised of a flexible array of sites for sequence-specific DNA binding proteins, "PcG recruiters," including Pho, Spps, Cg, and GAF. Pho is thought to play a central role in PcG recruitment. Early data showed that mutation of Pho binding sites in PREs in transgenes abrogated the ability of those PREs to repress gene expression. In contrast, genome-wide experiments in pho mutants or by Pho knockdown showed that PcG proteins can bind to PREs in the absence of Pho. Here, we directly addressed the importance of Pho binding sites in 2 engrailed (en) PREs at the endogenous locus and in transgenes. Our results show that Pho binding sites are required for PRE activity in transgenes with a single PRE. In a transgene, 2 PREs together lead to stronger, more stable repression and confer some resistance to the loss of Pho binding sites. Making the same mutation in Pho binding sites has little effect on PcG-protein binding at the endogenous en gene. Overall, our data support the model that Pho is important for PcG binding but emphasize how multiple PREs and chromatin environment increase the ability of PREs to function in the absence of Pho. This supports the view that multiple mechanisms contribute to PcG recruitment in Drosophila. [ABSTRACT FROM AUTHOR]

Valosin-containing protein (VCP) binds and extracts ubiquitylated cargo to regulate protein homeostasis. VCP has been studied primarily in aging and disease contexts, but it also affects germline development. However, the precise molecular functions of VCP in the germline, particularly in males, are poorly understood. Using the Drosophila male germline as a model system, we find that VCP translocates from the cytosol to the nucleus as germ cells transition into the meiotic spermatocyte stage. Importantly, nuclear translocation of VCP appears to be one crucial event stimulated by testis-specific TBP-associated factors (tTAFs) to drive spermatocyte differentiation. VCP promotes the expression of several tTAF-target genes, and VCP knockdown, like tTAF loss of function, causes cells to arrest in early meiotic stages. At a molecular level, VCP activity supports spermatocyte gene expression by downregulating a repressive histone modification, mono-ubiquitylated H2A (H2Aub), during meiosis. Remarkably, experimentally blocking H2Aub in VCPRNAi testes is sufficient to overcome the meiotic-arrest phenotype and to promote development through the spermatocyte stage. Collectively, our data highlight VCP as a downstream effector of tTAFs that downregulates H2Aub to facilitate meiotic progression. [ABSTRACT FROM AUTHOR]

The establishment and stable inheritance of individual patterns of gene expression in different cell types are required for the development of multicellular organisms. The important epigenetic regulators are the Polycomb group (PcG) and Trithorax group (TrxG) proteins, which control the silenced and active states of genes, respectively. In Drosophila, the PcG/TrxG group proteins are recruited to the DNA regulatory sequences termed the Polycomb response elements (PREs). The PREs are composed of the binding sites for different DNA-binding proteins, the so-called PcG recruiters. Currently, the role of the PcG recruiters in the targeting of the PcG proteins to PREs is well documented. However, there are examples where the PcG recruiters are also implicated in the active transcription and in the TrxG function. In addition, there is increasing evidence that the genome-wide PcG recruiters interact with the chromatin outside of the PREs and overlap with the proteins of differing regulatory classes. Recent studies of the interactomes of the PcG recruiters significantly expanded our understanding that they have numerous interactors besides the PcG proteins and that their functions extend beyond the regulation of the PRE repressive activity. Here, we summarize current data about the functions of the PcG recruiters. [ABSTRACT FROM AUTHOR]

The cohesin complex has a range of crucial functions in the cell. Cohesin is essential for mediating chromatid cohesion during mitosis, for repair of double‐strand DNA breaks, and for control of gene transcription. This last function has been the subject of intense research ever since the discovery of cohesin's role in the long‐range regulation of the cut gene in Drosophila. Subsequent research showed that the expression of some genes is exquisitely sensitive to cohesin depletion, while others remain relatively unperturbed. Sensitivity to cohesin depletion is also remarkably cell type‐ and/or condition‐specific. The relatively recent discovery that cohesin is integral to forming chromatin loops via loop extrusion should explain much of cohesin's gene regulatory properties, but surprisingly, loop extrusion has failed to identify a 'one size fits all' mechanism for how cohesin controls gene expression. This review will illustrate how early examples of cohesin‐dependent gene expression integrate with later work on cohesin's role in genome organization to explain mechanisms by which cohesin regulates gene expression. [ABSTRACT FROM AUTHOR]

Chromatin is an essential contributor to the regulation of transcription. The two histone post-translational modifications H3K4me3 and H3K27me3 act as an activator and repressor of gene expression, respectively, and are usually described as being mutually exclusive. However, recent work revealed that both marks might co-exist at several loci, forming a distinctive chromatin state called bivalency. While this state has been detected on a handful of genes involved in plant development and stress responses, its role in the regulation of transcription remains unclear. In an effort to shed more light on the putative function(s) of bivalency in plants, this review details the potential players involved in its setting and reading, and explores how this chromatin state might contribute to the control of gene expression. We propose that bivalency maintains transcriptional plasticity by facilitating transitions between a repressed and an active state and/or by preventing irreversible silencing of its targets. We also highlight recently developed techniques that could be used for further investigating bivalency. [ABSTRACT FROM AUTHOR]

Autophagy is a conserved cytoprotective process, aberrations in which lead to numerous degenerative disorders. While the cytoplasmic components of autophagy have been extensively studied, the epigenetic regulation of autophagy genes, especially in stem cells, is less understood. Deciphering the epigenetic regulation of autophagy genes becomes increasingly relevant given the therapeutic benefits of small-molecule epigenetic inhibitors in novel treatment modalities. We observe that, during retinoic acid-mediated differentiation of mouse embryonic stem cells (mESCs), autophagy is induced, and identify the Polycomb group histone methyl transferase EZH2 as a regulator of this process. In mESCs, EZH2 represses several autophagy genes, including the autophagy regulator DNA damageregulated autophagy modulator protein 1 (Dram1). EZH2 facilitates the formation of a bivalent chromatin domain at the Dram1 promoter, allowing gene expression and autophagy induction during differentiation while retaining the repressive H3K27me3 mark. EZH2 inhibition leads to loss of the bivalent domain, with consequent 'hyper-expression' of Dram1, accompanied by extensive cell death. This study shows that Polycomb group proteins help maintain a balance between autophagy and cell death during stem cell differentiation, in part, by regulating the expression of the Dram1 gene. [ABSTRACT FROM AUTHOR]

Simple Summary: Epigenetic factors control how genes are expressed in different cell types. Members of the Polycomb Repressive Complexes (PRCs) are critical for epigenetic control of gene transcription in blood cells, and mutations and deletions in these factors are common in the blood cancer acute myeloid leukaemia (AML). This review article provides an overview of how the structure and function of PRCs are affected by genetic alterations in AML, with a primary focus on PRC2 core factors. We document how mutations and deletions in PRC2 factors are linked to other AML-associated genetic alterations and discuss how these observations might inform potential treatment avenues in future. Epigenetic dysregulation is a hallmark of many haematological malignancies and is very frequent in acute myeloid leukaemia (AML). A cardinal example is the altered activity of the Polycomb Repressive Complex 2 (PRC2) due to somatic mutations and deletions in genes encoding PRC2 core factors that are necessary for correct complex assembly. These genetic alterations typically lead to reduced histone methyltransferase activity that, in turn, has been strongly linked to poor prognosis and chemoresistance. In this review, we provide an overview of genetic alterations of PRC components in AML, with particular reference to structural and functional features of PRC2 factors. We further review genetic interactions between these alterations and other AML-associated mutations in both adult and paediatric leukaemias. Finally, we discuss reported prognostic links between PRC2 mutations and deletions and disease outcomes and potential implications for therapy. [ABSTRACT FROM AUTHOR]

Characterization of gene regulatory networks is fundamental to understanding homeostatic development. This process can be simplified by analyzing relatively simple genomes such as the genome of Drosophila melanogaster. In this work we have developed a computational framework in Drosophila to explore for the presence of gene regulatory circuits between two large groups of transcriptional regulators: the epigenetic group of the Polycomb/trithorax (PcG/trxG) proteins and the microRNAs (miRNAs). We have searched genome-wide for miRNA targets in PcG / trxG transcripts as well as for Polycomb Response Elements (PREs) in miRNA genes. Our results show that 10% of the analyzed miRNAs could be controlling PcG/trxG gene expression, while 40% of those miRNAs are putatively controlled by the selected set of PcG/trxG proteins. The integration of these analyses has resulted in the predicted existence of 3 classes of miRNA-PcG/trxG crosstalk interactions that define potential regulatory circuits. In the first class, miRNA-PcG circuits are defined by miRNAs that reciprocally crosstalk with PcG. In the second, miRNA-trxG circuits are defined by miRNAs that reciprocally crosstalk with trxG. In the third class, miRNA-PcG/trxG shared circuits are defined by miRNAs that crosstalk with both PcG and trxG regulators. These putative regulatory circuits may uncover a novel mechanism in Drosophila for the control of PcG/trxG and miRNAs levels of expression. The computational framework developed here for Drosophila melanogaster can serve as a model case for similar analyses in other species. Moreover, our work provides, for the first time, a new and useful resource for the Drosophila community to consult prior to experimental studies investigating the epigenetic regulatory networks of miRNA-PcG/trxG mediated gene expression. [Display omitted] • A computational framework to define epigenetic regulatory circuits in Drosophila. • Regulatory interactions between microRNAs and Polycomb (PcG)/ trithorax (trxG) genes. • 35 and 15 microRNAs potentially regulate PcG and trxG gene expression respectively. • 106 pre-miRNAs are potentially regulated by PcG/trxG proteins. • miRNA-PcG/trxG crosstalk interactions define 21 gene regulatory circuits. [ABSTRACT FROM AUTHOR]

Polycomb and Trithorax group proteins (PcG and TrxG) are epigenetic factors responsible for the repression and activation of transcription, respectively. In Drosophila, PcG/TrxG proteins are recruited to specialized DNA elements called Polycomb response elements (PREs). Depending on the context, these elements may repress, activate, or be neutral with respect to the promoter of the target gene. Previously, in transgenic studies using PhiC31 site-specific integration system, we have demonstrated that sites for architectural proteins inserted next to PRE can induce the repressive activity of bxdPRE by stimulating the binding of PcG/TrxG factors to this element. However, this effect may depend on additional DNA elements present at the integration site after PhiC31-dependent transgene insertion. In the present study, using an alternative system of integration based on CRISPR/Cas9-catalyzed homology-directed repair, we have proved that the binding sites of the architectural protein Su(Hw) are indeed able to induce the repressive activity of bxdPRE and recruitment of PcG/TrxG proteins, regardless of the heterogeneous DNA-sequences present at the site of integration after PhiC31-dependent insertion of the transgenes. [ABSTRACT FROM AUTHOR]

The Drosophila PRC1 complex regulates gene expression by modifying histone proteins and chromatin architecture. Two PRC1 subunits, PSC and Ph, are most implicated in chromatin architecture. In vitro, PRC1 compacts chromatin and inhibits transcription and nucleosome remodeling. The long disordered C-terminal region of PSC (PSC-CTR) is important for these activities, while Ph has little effect. In cells, Ph is important for condensate formation, long-range chromatin interactions, and gene regulation, and its polymerizing sterile alpha motif (SAM) is implicated in these activities. In vitro, truncated Ph containing the SAM and two other conserved domains (mini-Ph) undergoes phase separation with chromatin, suggesting a mechanism for SAM-dependent condensate formation in vivo. How the distinct activities of PSC and Ph on chromatin function together in PRC1 is not known. To address this question, we analyzed structures formed with large chromatin templates and PRC1 in vitro. PRC1 bridges chromatin into extensive fibrillar networks. Ph, its SAM, and SAM polymerization activity have little effect on these structures. Instead, the PSC-CTR controls their growth, and is sufficient for their formation. To understand how phase separation driven by Ph SAM intersects with the chromatin bridging activity of the PSC-CTR, we used mini-Ph to form condensates with chromatin and then challenged them with PRC1 lacking Ph (PRC1ΔPh). PRC1ΔPh converts mini-Ph chromatin condensates into clusters of small non-fusing condensates and bridged fibers. These condensates retain a high level of chromatin compaction and do not intermix. Thus, phase separation of chromatin by mini-Ph, followed by the action of the PSC-CTR, creates a unique chromatin organization with regions of high nucleosome density and extraordinary stability. We discuss how this coordinated sequential activity of two proteins found in the same complex may occur and the possible implications of stable chromatin architectures in maintaining transcription states.

In many animal species with a bilateral symmetry, Hox genes are clustered either at one or at several genomic loci. This organization has a functional relevance, as the transcriptional control applied to each gene depends upon its relative position within the gene cluster. It was previously noted that vertebrate Hox clusters display a much higher level of genomic organization than their invertebrate counterparts. The former are always more compact than the latter, they are generally devoid of repeats and of interspersed genes, and all genes are transcribed by the same DNA strand, suggesting that particular factors constrained these clusters toward a tighter structure during the evolution of the vertebrate lineage. Here, we investigate the importance of uniform transcriptional orientation by engineering several alleles within the HoxD cluster, such as to invert one or several transcription units, with or without a neighboring CTCF site. We observe that the association between the tight structure of mammalian Hox clusters and their regulation makes inversions likely detrimental to the proper implementation of this complex genetic system. We propose that the consolidation of Hox clusters in vertebrates, including transcriptional polarity, evolved in conjunction with the emergence of global gene regulation via the flanking regulatory landscapes, to optimize a coordinated response of selected subsets of target genes in cis.

Genes involved in sexual reproduction diverge rapidly as a result of reproductive fitness. Here, we identify a novel protein domain in the germline-specific Polycomb protein SCML2 that is required for the establishment of unique gene expression programs after the mitosis-to-meiosis transition in spermatogenesis. We term this novel domain, which is comprised of rapidly evolved, DNA-binding repeat units of 28 amino acids, the SCML2 DNA-binding (SDB) repeats. These repeats are acquired in a specific subgroup of the rodent lineage, having been subjected to positive selection in the course of evolution. Mouse SCML2 has two DNA-binding domains: one is the SDB repeats and the other is an RNA-binding region, which is conserved in human SCML2. For the recruitment of SCML2 to target loci, the SDB repeats cooperate with the other functional domains of SCML2 to bind chromatin. The cooperative action of these domains enables SCML2 to sense DNA hypomethylation in an in vivo chromatin environment, thereby enabling SCML2 to bind to hypomethylated chromatin. We propose that the rapid evolution of SCML2 is due to reproductive adaptation, which has promoted species-specific gene expression programs in spermatogenesis.

Abstract Background RNA-DNA hybrid (R-loop)-associated long noncoding RNAs (lncRNAs), including the Arabidopsis lncRNA AUXIN-REGULATED PROMOTER LOOP (APOLO), are emerging as important regulators of three-dimensional chromatin conformation and gene transcriptional activity. Results Here, we show that in addition to the PRC1-component LIKE HETEROCHROMATIN PROTEIN 1 (LHP1), APOLO interacts with the methylcytosine-binding protein VARIANT IN METHYLATION 1 (VIM1), a conserved homolog of the mammalian DNA methylation regulator UBIQUITIN-LIKE CONTAINING PHD AND RING FINGER DOMAINS 1 (UHRF1). The APOLO-VIM1-LHP1 complex directly regulates the transcription of the auxin biosynthesis gene YUCCA2 by dynamically determining DNA methylation and H3K27me3 deposition over its promoter during the plant thermomorphogenic response. Strikingly, we demonstrate that the lncRNA UHRF1 Protein Associated Transcript (UPAT), a direct interactor of UHRF1 in humans, can be recognized by VIM1 and LHP1 in plant cells, despite the lack of sequence homology between UPAT and APOLO. In addition, we show that increased levels of APOLO or UPAT hamper VIM1 and LHP1 binding to YUCCA2 promoter and globally alter the Arabidopsis transcriptome in a similar manner. Conclusions Collectively, our results uncover a new mechanism in which a plant lncRNA coordinates Polycomb action and DNA methylation through the interaction with VIM1, and indicates that evolutionary unrelated lncRNAs with potentially conserved structures may exert similar functions by interacting with homolog partners.

The functional maturation of neurons is a prolonged process that extends past the mitotic exit and is mediated by the chromatin-dependent orchestration of gene transcription programs. We find that expression of this maturation gene program in mouse cerebellar granule neurons (CGNs) requires dynamic changes in the genomic distribution of histone H3 lysine 27 trimethylation (H3K27me3), demonstrating a function for this chromatin modification beyond its role in cell fate specification. The developmental loss of H3K27me3 at promoters of genes activated as CGNs mature is facilitated by the lysine demethylase and ASD-risk gene, Kdm6b. Interestingly, inhibition of the H3K27 methyltransferase EZH2 in newborn CGNs not only blocks the repression of progenitor genes but also impairs the induction of mature CGN genes, showing the importance of bidirectional H3K27me3 regulation across the genome. These data demonstrate that H3K27me3 turnover in developing postmitotic neurons regulates the temporal coordination of gene expression programs that underlie functional neuronal maturation.

Polycomb group proteins (PcG) play a crucial role in developmental programs in eukaryotic organisms, including plants. PcG-mediated gene repression is achieved by epigenetic histone modification on target chromatins. Loss of PcG components leads to severe developmental defects. CURLY LEAF (CLF), a PcG component in Arabidopsis, catalyzes the trimethylation of histone H3 on lysine 27 (H3K27me3), a repressive histone mark in numerous genes in Arabidopsis. In this study, we isolated a single homolog of Arabidopsis CLF, namely, BrCLF, in Brassica rapa ssp. trilocularis. Transcriptomic analysis revealed that BrCLF participated in B. rapa developmental processes, such as seed dormancy, leaf and flower organ development, and floral transition. BrCLF was also involved in stress signaling and stress-responsive metabolism, such as aliphatic and indolic glucosinolate metabolism in B. rapa. Epigenome analysis showed that H3K27me3 was substantially enriched in genes related to these developmental and stress-responsive processes. Thus, this study provided a basis for elucidating the molecular mechanism of the PcG-mediated regulation of development and stress responses in B. rapa. [ABSTRACT FROM AUTHOR]

During mammalian neurodevelopment, signaling pathways converge upon transcription factors (TFs) to establish appropriate gene expression programmes leading to the production of distinct neural and glial cell types. This process is partially regulated by the dynamic modulation of chromatin states by epigenetic systems, including the polycomb group (PcG) family of co-repressors. PcG proteins form multi-subunit assemblies that sub-divide into distinct, yet functionally related families. Polycomb repressive complexes 1 and 2 (PRC1 and 2) modify the chemical properties of chromatin by covalently modifying histone tails via H2A ubiquitination (H2AK119ub1) and H3 methylation, respectively. In contrast to the PRCs, the Polycomb repressive deubiquitinase (PR-DUB) complex removes H2AK119ub1 from chromatin through the action of the C-terminal hydrolase BAP1. Genetic screening has identified several PcG mutations that are causally associated with a range of congenital neuropathologies associated with both localised and/or systemic growth abnormalities. As PRC1 and PR-DUB hold opposing functions to control H2AK119ub1 levels across the genome, it is plausible that such neurodevelopmental disorders arise through a common mechanism. In this review, we will focus on advancements regarding the composition and opposing molecular functions of mammalian PRC1 and PR-DUB, and explore how their dysfunction contributes to the emergence of neurodevelopmental disorders. [ABSTRACT FROM AUTHOR]

The Polycomb group (PcG) complex PRC1 localizes in the nucleus in condensed structures called Polycomb bodies. The PRC1 subunit Polyhomeotic (Ph) contains an oligomerizing sterile alpha motif (SAM) that is implicated in both PcG body formation and chromatin organization in Drosophila and mammalian cells. A truncated version of Ph containing the SAM (mini-Ph) forms phase-separated condensates with DNA or chromatin in vitro, suggesting that PcG bodies may form through SAM-driven phase separation. In cells, Ph forms multiple small condensates, while mini-Ph typically forms a single large nuclear condensate. We therefore hypothesized that sequences outside of mini-Ph, which are predicted to be intrinsically disordered, are required for proper condensate formation. We identified three distinct low-complexity regions in Ph based on sequence composition. We systematically tested the role of each of these sequences in Ph condensates using live imaging of transfected Drosophila S2 cells. Each sequence uniquely affected Ph SAM-dependent condensate size, number, and morphology, but the most dramatic effects occurred when the central, glutamine-rich intrinsically disordered region (IDR) was removed, which resulted in large Ph condensates. Like mini-Ph condensates, condensates lacking the glutamine-rich IDR excluded chromatin. Chromatin fractionation experiments indicated that the removal of the glutamine-rich IDR reduced chromatin binding and that the removal of either of the other IDRs increased chromatin binding. Our data suggest that all three IDRs, and functional interactions among them, regulate Ph condensate size and number. Our results can be explained by a model in which tight chromatin binding by Ph IDRs antagonizes Ph SAM-driven phase separation. Our observations highlight the complexity of regulation of biological condensates housed in single proteins. [ABSTRACT FROM AUTHOR]

• OPCs lacking p53 form tumors when injected in mice only if they also express PDGF-BB. • PDGF-BB overexpression in p53 null OPC results in more proliferation, less differentiation and altered growth characteristics. • PDGF-BB overexpressing p53 null OPC display high levels of the repressive H3K27me3 histone mark and low levels of H4K20me3. • Pharmacological inhibition of H3K27 methylation alone is not sufficient to revert the growth phenotype of the PDGF-BB overexpressing p53 null OPC. Proneural gliomas are brain tumors characterized by enrichment of oligodendrocyte progenitor cell (OPC) transcripts and genetic alterations. In this study we sought to identify transcriptional and epigenetic differences between OPCs with Trp53 deletion and PDGF-BB overexpression (BB-p53n) and those carrying only p53 deletion (p53n). In culture, the BB-p53n OPCs display growth characteristics more similar to glioma cells than p53n OPCs. When injected in mouse brains, BB-p53n OPC form tumors, while the p53n OPCs do not. Unbiased histone proteomics and transcriptomic analysis on these OPC populations identified higher levels of the histone H3K27me3 mark and lower levels of the histone H4K20me3. The transcriptome of the BB-p53n OPCs was characterized by higher levels of transcripts related to proliferation and cell adhesion compared to p53n OPCs. Pharmacological inhibition of the enzyme responsible for histone H3K27 trimethylation (EZH2i) in BB-p53n OPCs, reduced cell cycle transcripts and increased the expression of differentiation markers, but was not sufficient to restore their growth characteristics. This suggests that PDGF-BB overexpression in p53n OPCs favors the early stages of transformation, by promoting proliferation and halting differentiation in a H3K27me3-dependent pathway, and favoring growth characteristics in a H3K27me3 independent manner [ABSTRACT FROM AUTHOR]

Abstract Background Neural progenitors produce diverse cells in a stereotyped birth order, but can specify each cell type for only a limited duration. In the Drosophila embryo, neuroblasts (neural progenitors) specify multiple, distinct neurons by sequentially expressing a series of temporal identity transcription factors with each division. Hunchback (Hb), the first of the series, specifies early-born neuronal identity. Neuroblast competence to generate early-born neurons is terminated when the hb gene relocates to the neuroblast nuclear lamina, rendering it refractory to activation in descendent neurons. Mechanisms and trans-acting factors underlying this process are poorly understood. Here we identify Corto, an enhancer of Trithorax/Polycomb (ETP) protein, as a new regulator of neuroblast competence. Methods We used the GAL4/UAS system to drive persistent misexpression of Hb in neuroblast 7–1 (NB7-1), a model lineage for which the early competence window has been well characterized, to examine the role of Corto in neuroblast competence. We used immuno-DNA Fluorescence in situ hybridization (DNA FISH) in whole embryos to track the position of the hb gene locus specifically in neuroblasts across developmental time, comparing corto mutants to control embryos. Finally, we used immunostaining in whole embryos to examine Corto’s role in repression of Hb and a known target gene, Abdominal B (Abd-B). Results We found that in corto mutants, the hb gene relocation to the neuroblast nuclear lamina is delayed and the early competence window is extended. The delay in gene relocation occurs after hb transcription is already terminated in the neuroblast and is not due to prolonged transcriptional activity. Further, we find that Corto genetically interacts with Posterior Sex Combs (Psc), a core subunit of polycomb group complex 1 (PRC1), to terminate early competence. Loss of Corto does not result in derepression of Hb or its Hox target, Abd-B, specifically in neuroblasts. Conclusions These results show that in neuroblasts, Corto genetically interacts with PRC1 to regulate timing of nuclear architecture reorganization and support the model that distinct mechanisms of silencing are implemented in a step-wise fashion during development to regulate cell fate gene expression in neuronal progeny.

Abstract The Polycomb repressive complex 2 (PRC2) is an essential chromatin regulatory complex involved in repressing the transcription of diverse developmental genes. PRC2 consists of a core complex; possessing H3K27 methyltransferase activity and various associated factors that are important to modulate its function. During evolution, the composition of PRC2 and the functionality of PRC2 components have changed considerably. Here, we compare the PRC2 complex members of Drosophila and mammals and describe their adaptation to altered biological needs. We also highlight how the PRC2.1 subcomplex has gained multiple novel functions and discuss the implications of these changes for the function of PRC2 in chromatin regulation.

Polycomb group proteins (PcG) play a crucial role in developmental programs in eukaryotic organisms, including plants. PcG-mediated gene repression is achieved by epigenetic histone modification on target chromatins. Loss of PcG components leads to severe developmental defects. CURLY LEAF (CLF), a PcG component in Arabidopsis, catalyzes the trimethylation of histone H3 on lysine 27 (H3K27me3), a repressive histone mark in numerous genes in Arabidopsis. In this study, we isolated a single homolog of Arabidopsis CLF, namely, BrCLF, in Brassica rapa ssp. trilocularis. Transcriptomic analysis revealed that BrCLF participated in B. rapa developmental processes, such as seed dormancy, leaf and flower organ development, and floral transition. BrCLF was also involved in stress signaling and stress-responsive metabolism, such as aliphatic and indolic glucosinolate metabolism in B. rapa. Epigenome analysis showed that H3K27me3 was substantially enriched in genes related to these developmental and stress-responsive processes. Thus, this study provided a basis for elucidating the molecular mechanism of the PcG-mediated regulation of development and stress responses in B. rapa.

Repressive H3K27me3 and active H3K4me2/3 together form bivalent chromatin domains, molecular hallmarks of developmental potential. In the male germline, these domains are thought to persist into sperm to establish totipotency in the next generation. However, it remains unknown how H3K27me3 is established on specific targets in the male germline. Here, we demonstrate that a germline-specific Polycomb protein, SCML2, binds to H3K4me2/3-rich hypomethylated promoters in undifferentiated spermatogonia to facilitate H3K27me3. Thus, SCML2 establishes bivalent domains in the male germline of mice. SCML2 regulates two major classes of bivalent domains: Class I domains are established on developmental regulator genes that are silent throughout spermatogenesis, while class II domains are established on somatic genes silenced during late spermatogenesis. We propose that SCML2-dependent H3K27me3 in the male germline prepares the expression of developmental regulator and somatic genes in embryonic development.

Epigenetic factors (EFs) regulate multiple aspects of cerebral cortex development, including proliferation, differentiation, laminar fate, and regional identity. The same neurodevelopmental processes are also regulated by transcription factors (TFs), notably the Pax6→ Tbr2→ Tbr1 cascade expressed sequentially in radial glial progenitors (RGPs), intermediate progenitors, and postmitotic projection neurons, respectively. Here, we studied the EF landscape and its regulation in embryonic mouse neocortex. Microarray and in situ hybridization assays revealed that many EF genes are expressed in specific cortical cell types, such as intermediate progenitors, or in rostrocaudal gradients. Furthermore, many EF genes are directly bound and transcriptionally regulated by Pax6, Tbr2, or Tbr1, as determined by chromatin immunoprecipitation-sequencing and gene expression analysis of TF mutant cortices. Our analysis demonstrated that Pax6, Tbr2, and Tbr1 form a direct feedforward genetic cascade, with direct feedback repression. Results also revealed that each TF regulates multiple EF genes that control DNA methylation, histone marks, chromatin remodeling, and non-coding RNA. For example, Tbr1 activates Rybp and Auts2 to promote the formation of non-canonical Polycomb repressive complex 1 (PRC1). Also, Pax6, Tbr2, and Tbr1 collectively drive massive changes in the subunit isoform composition of BAF chromatin remodeling complexes during differentiation: for example, a novel switch from Bcl7c (Baf40c) to Bcl7a (Baf40a), the latter directly activated by Tbr2. Of 11 subunits predominantly in neuronal BAF, 7 were transcriptionally activated by Pax6, Tbr2, or Tbr1. Using EFs, Pax6→ Tbr2→ Tbr1 effect persistent changes of gene expression in cell lineages, to propagate features such as regional and laminar identity from progenitors to neurons.

Xenopus tropicalis tadpoles have the capacity for scarless regeneration of appendages including the limb and tail. Following injury, transcriptional programs must be activated and inactivated with high spatial and temporal resolution to result in a properly patterned appendage. Functional studies have established that histone‐modifying enzymes that act to close chromatin are required for regeneration, but the genomic regions sensitive to these activities are not fully established. Here we show that early inhibition of HDAC or EZH2 activity results in incomplete tail regeneration. To identify how each of these perturbations impacts chromatin accessibility, we applied an assay for transposase‐accessible chromatin (ATAC‐seq) to HDAC or EZH2‐inhibited regenerating tadpoles. We find that neither perturbation results in a global increase in chromatin accessibility, but that both inhibitors have targeted effects on chromatin accessibility and gene expression. Upon HDAC inhibition, regulatory regions neighbouring genes associated with neuronal regeneration are preferentially accessible, whereas regions associated with immune response and apoptosis are preferentially accessible following EZH2 inhibition. Together, these results suggest distinct roles for these two chromatin‐closing activities in appendage regeneration. [ABSTRACT FROM AUTHOR]

Genetic and epigenetic changes might facilitate the acquisition of stem cell-like phenotypes of tumors, resulting in worse patients outcome. Although the role of chromobox (CBX) domain proteins, a family of epigenetic factors that recognize specific histone marks, in the pathogenesis of several tumor types is well documented, little is known about their association with cancer stemness. Here, we have characterized the relationship between the CBX family members' expression and cancer stemness in liver, lung, pancreatic, and uterine tumors using publicly available TCGA and GEO databases and harnessing several bioinformatic tools (i.e., Oncomine, GEPIA2, TISIDB, GSCA, UALCAN, R2 platform, Enrichr, GSEA). We demonstrated that significant upregulation of CBX3 and downregulation of CBX7 are consistently associated with enriched cancer stem-cell-like phenotype across distinct tumor types. High CBX3 expression is observed in higher-grade tumors that exhibit stem cell-like traits, and CBX3-associated gene expression profiles are robustly enriched with stemness markers and targets for c-Myc transcription factor regardless of the tumor type. Similar to high-stemness tumors, CBX3-overexpressing cancers manifest a higher mutation load. On the other hand, higher-grade tumors are characterized by the significant downregulation of CBX7, and CBX7-associated gene expression profiles are significantly depleted with stem cell markers. In contrast to high-stemness tumors, cancer with CBX7 upregulation exhibit a lower mutation burden. Our results clearly demonstrate yet unrecognized association of high CBX3 and low CBX7 expression with cancer stem cell-like phenotype of solid tumors. [ABSTRACT FROM AUTHOR]

Although all cells in the human body are made of the same DNA, these cells undergo differentiation and behave differently during development, through integration of external and internal stimuli via 'specific mechanisms.' Epigenetics is one such mechanism that comprises DNA/RNA, histone modifications, and non-coding RNAs that regulate transcription without changing the genetic code. The discovery of the first Polycomb mutant phenotype in Drosophila started the study of epigenetics more than 80 years ago. Since then, a considerable number of Polycomb Group (PcG) genes in Drosophila have been discovered to be preserved in mammals, including humans. PcG proteins exert their influence through gene repression by acting in complexes, modifying histones, and compacting the chromatin within the nucleus. In this article, we discuss how our knowledge of the PcG repression mechanism in Drosophila translates to human communicable disease research. [ABSTRACT FROM AUTHOR]