PCGF homologs, CBX proteins, and RYBP define functionally distinct PRC1 family complexes

The heterogeneous nature of mammalian PRC1 complexes has hindered our understanding of their biological functions. Here, we present a comprehensive proteomic and genomic analysis that uncovered six major groups of PRC1 complexes, each containing a distinct PCGF subunit, a RING1A/B ubiquitin ligase, and a unique set of associated polypeptides. These PRC1 complexes differ in their genomic localization, and only a small subset colocalize with H3K27me3. Further biochemical dissection revealed that the six PCGF-RING1A/B combinations form multiple complexes through association with RYBP or its homolog YAF2, which prevents the incorporation of other canonical PRC1 subunits, such as CBX, PHC, and SCM. Although both RYBP/YAF2- and CBX/PHC/SCM-containing complexes compact chromatin, only RYBP stimulates the activity of RING1B toward H2AK119ub1, suggesting a central role in PRC1 function. Knockdown of RYBP in embryonic stem cells compromised their ability to form embryoid bodies, likely because of defects in cell proliferation and maintenance of H2AK119ub1 levels.     

YTHDF1 facilitates PRC1-mediated H2AK119ub in human ES cells

Polycomb repressive complexes (PRCs) play critical roles in cell fate decisions during normal development as well as disease progression through mediating histone modifications such as H3K27me3 and H2AK119ub. How exactly PRCs recruited to chromatin remains to be fully illuminated. Here, we report that YTHDF1, the N6-methyladenine (m⁶A) RNA reader that was previously known to be mainly cytoplasmic, associates with RNF2, a PRC1 protein that mediates H2AK119ub in human embryonic stem cells (hESCs). A portion of YTHDF1 localizes in the nuclei and associates with RNF2/H2AK119ub on a subset of gene loci related to neural development functions. Knock-down YTHDF1 attenuates H2AK119ub modification on these genes and promotes neural differentiation in hESCs. Our findings provide a noncanonical mechanism that YTHDF1 participates in PRC1 functions in hESCs.     

Interdependence of PRC1 and PRC2 for recruitment to Polycomb Response Elements

Polycomb Group (PcG) proteins are epigenetic repressors essential for control of development and cell differentiation. They form multiple complexes of which PRC1 and PRC2 are evolutionary conserved and obligatory for repression. The targeting of PRC1 and PRC2 is poorly understood and was proposed to be hierarchical and involve tri-methylation of histone H3 (H3K27me3) and/or monoubiquitylation of histone H2A (H2AK118ub). Here, we present a strict test of this hypothesis using the Drosophila model. We discover that neither H3K27me3 nor H2AK118ub is required for targeting PRC complexes to Polycomb Response Elements (PREs). We find that PRC1 can bind PREs in the absence of PRC2 but at many PREs PRC2 requires PRC1 to be targeted. We show that one role of H3K27me3 is to allow PcG complexes anchored at PREs to interact with surrounding chromatin. In contrast, the bulk of H2AK118ub is unrelated to PcG repression. These findings radically change our view of how PcG repression is targeted and suggest that PRC1 and PRC2 can communicate independently of histone modifications.     

Histone H2A Mono-Ubiquitination Is a Crucial Step to Mediate PRC1-Dependent Repression of Developmental Genes to Maintain ES Cell Identity

Two distinct Polycomb complexes, PRC1 and PRC2, collaborate to maintain epigenetic repression of key developmental loci in embryonic stem cells (ESCs). PRC1 and PRC2 have histone modifying activities, catalyzing mono-ubiquitination of histone H2A (H2AK119u1) and trimethylation of H3 lysine 27 (H3K27me3), respectively. Compared to H3K27me3, localization and the role of H2AK119u1 are not fully understood in ESCs. Here we present genome-wide H2AK119u1 maps in ESCs and identify a group of genes at which H2AK119u1 is deposited in a Ring1-dependent manner. These genes are a distinctive subset of genes with H3K27me3 enrichment and are the central targets of Polycomb silencing that are required to maintain ESC identity. We further show that the H2A ubiquitination activity of PRC1 is dispensable for its target binding and its activity to compact chromatin at Hox loci, but is indispensable for efficient repression of target genes and thereby ESC maintenance. These data demonstrate that multiple effector mechanisms including H2A ubiquitination and chromatin compaction combine to mediate PRC1-dependent repression of genes that are crucial for the maintenance of ESC identity. Utilization of these diverse effector mechanisms might provide a means to maintain a repressive state that is robust yet highly responsive to developmental cues during ES cell self-renewal and differentiation.     

PcG蛋白转录抑制复合物组份的功能及构成的时空特征

PcG蛋白主要以PRC1和PRC2两组复合物的形式存在,通过参与核小体组蛋白翻译后修饰等机制,发挥调控靶基因转录的功能. PRC1复合体中的RING1A/B具有使组蛋白H2AK119泛素化的活性;PRC2中的EZH2具有使组蛋白H3K27三甲基化的活性,形成PRC1锚定到核小体上的位点. PcG蛋白的表达特征具有发育阶段和细胞类型时空特异性. 长链非编码RNA等反式作用因子能募集PcG蛋白结合于靶基因,发挥靶向作用. 本文就PcG蛋白功能、构成的时空特异性、募集机制及其与疾病发生的关系研究进展做一综述.     

Histone monoubiquitylation position determines specificity and direction of enzymatic cross-talk with histone methyltransferases Dot1L and PRC2

It is well established that chromatin is a destination for signal transduction, affecting many DNA-templated processes. Histone proteins in particular are extensively post-translationally modified. We are interested in how the complex repertoire of histone modifications is coordinately regulated to generate meaningful combinations of "marks" at physiologically relevant genomic locations. One important mechanism is "cross-talk" between pre-existing histone post-translational modifications and enzymes that subsequently add or remove modifications on chromatin. Here, we use chemically defined "designer" nucleosomes to investigate novel enzymatic cross-talk relationships between the most abundant histone ubiquitylation sites, H2AK119ub and H2BK120ub, and two important histone methyltransferases, Dot1L and PRC2. Although the presence of H2Bub in nucleosomes greatly stimulated Dot1L methylation of H3K79, we found that H2Aub did not influence Dot1L activity. In contrast, we show that H2Aub inhibited PRC2 methylation of H3K27, but H2Bub did not influence PRC2 activity. Taken together, these results highlight how the position of nucleosome monoubiquitylation affects the specificity and direction of cross-talk with enzymatic activities on chromatin.     

RYBP represses endogenous retroviruses and preimplantation- and germ line-specific genes in mouse embryonic stem cells

Polycomb repressive complexes (PRCs) are important chromatin regulators of embryonic stem (ES) cell function. RYBP binds Polycomb H2A monoubiquitin ligases Ring1A and Ring1B and has been suggested to assist PRC localization to their targets. Moreover, constitutive inactivation of RYBP precludes ES cell formation. Using ES cells conditionally deficient in RYBP, we found that RYBP is not required for maintenance of the ES cell state, although mutant cells differentiate abnormally. Genome-wide chromatin association studies showed RYBP binding to promoters of Polycomb targets, although its presence is dispensable for gene repression. We discovered, using Eed-knockout (KO) ES cells, that RYBP binding to promoters was independent of H3K27me3. However, recruiting of PRC1 subunits Ring1B and Mel18 to their targets was not altered in the absence of RYBP. In contrast, we have found that RYBP efficiently represses endogenous retroviruses (murine endogenous retrovirus [MuERV] class) and preimplantation (including zygotic genome activation stage)- and germ line-specific genes. These observations support a selective repressor activity for RYBP that is dispensable for Polycomb function in the ES cell state. Also, they suggest a role for RYBP in epigenetic resetting during preimplantation development through repression of germ line genes and PcG targets before formation of pluripotent epiblast cells.     

Distinct dynamics of HISTONE3 variants between the two fertilization products in plants

Sexual reproduction involves epigenetic reprogramming comprising DNA methylation and histone modifications. In addition, dynamics of HISTONE3 (H3) variant H3.3 upon fertilization are conserved in animals, suggesting an essential role. In contrast to H3, H3.3 marks actively transcribed regions of the genome and can be deposited in a replication-independent manner. Although H3 variants are conserved in plants, their dynamics during fertilization have remained unexplored. We overcame technical limitations to live imaging of the fertilization process in Arabidopsis thaliana and studied dynamics of the male-gamete-specific H3.3 and the centromeric Histone Three Related 12 (HTR12). The double-fertilization process in plants produces the zygote and the embryo-nourishing endosperm. We show that the zygote is characterized by replication-independent removal of paternal H3.3 and homogeneous incorporation of parental chromatin complements. In the endosperm, the paternal H3.3 is passively diluted by replication while the paternal chromatin remains segregated apart from the maternal chromatin (gonomery). Hence epigenetic regulations distinguish the two products of fertilization in plants. H3.3-replication-independent dynamics and gonomery also mark the first zygotic divisions in animal species. We thus propose the convergent selection of parental epigenetic imbalance involving H3 variants in sexually reproducing organisms.     

Polycomb repressive complex 1: Regulators of neurogenesis from embryonic to adult stage

Development of vertebrate nervous system is a complex process which involves differential gene expression and disruptions in this process or in the mature brain, may lead to neurological disorders and diseases. Extensive work that spanned several decades using rodent models and recent work on stem cells have helped uncover the intricate process of neuronal differentiation and maturation. There are various morphological changes, genetic and epigenetic modifications which occur during normal mammalian neural development, one of the chromatin modifications that controls vital gene expression are the posttranslational modifications on histone proteins, that controls accessibility of translational machinery. Among the histone modifiers, polycomb group proteins (PcGs), such as Ezh2, Eed and Suz12 form large protein complexes—polycomb repressive complex 2 (PRC2); while Ring1b and Bmi1 proteins form core of PRC1 along with accessory proteins such as Cbx, Hph, Rybp and Pcgfs catalyse histone modifications such as H3K27me3 and H2AK119ub1. PRC1 proteins are known to play critical role in X chromosome inactivation in females but they also repress the expression of key developmental genes and tightly regulate the mammalian neuronal development. In this review we have discussed the signalling pathways, morphogens and nuclear factors that initiate, regulate and maintain cells of the nervous system. Further, we have extensively reviewed the recent literature on the role of Ring1b and Bmi1 in mammalian neuronal development and differentiation; as well as highlighted questions that are still unanswered.     

Dynamics of histone H3 acetylation in the nucleosome core during mouse pre-implantation development

In mammals, the time period that follows fertilization is characterized by extensive chromatin remodeling, which enables epigenetic reprogramming of the gametes. Major changes in chromatin structure persist until the time of implantation, when the embryo develops into a blastocyst, which comprises the inner cell mass and the trophectoderm. Changes in DNA methylation, histone variant incorporation, and covalent modifications of the histones tails have been intensively studied during pre-implantation development. However, modifications within the core of the nucleosomes have not been systematically analyzed. Here, we report the first characterization and temporal analysis of 3 key acetylated residues in the core of the histone H3: H3K64ac, H3K122ac, and H3K56ac, all located at structurally important positions close to the DNA. We found that all 3 acetylations occur during pre-implantation development, but with different temporal kinetics. Globally, H3K64ac and H3K56ac were detected throughout cleavage stages, while H3K122ac was only weakly detectable during this time. Our work contributes to the understanding of the contribution of histone modifications in the core of the nucleosome to the “marking” of the newly established embryonic chromatin and unveils new modification pathways potentially involved in epigenetic reprogramming.     

Mechanisms of histone H3 lysine 27 trimethylation remodeling during early mammalian development

During fertilization, two of the most differentiated cells in the mammalian organism, a sperm and oocyte, are combined to form a pluripotent embryo. Dynamic changes in chromatin structure allow the transition of the chromatin on these specialized cells into an embryonic configuration capable of generating every cell type. Initially, this reprogramming activity is supported by oocyte-derived factors accumulated during oogenesis as proteins and mRNAs; however, the underlying molecular mechanisms that govern it remain poorly characterized. Trimethylation of histone H3 at lysine 27 (H3K27me3) is a repressive epigenetic mark that changes dynamically during pre-implantation development in mice, bovine and pig embryos. Here we present data and hypotheses related to the potential mechanisms behind H3K27me3 remodeling during early development. We postulate that the repressive H3K27me3 mark is globally erased from the parental genomes in order to remove the gametic epigenetic program and to establish a pluripotent embryonic epigenome. We discuss information gathered in mice, pigs, and bovine, with the intent of providing a comparative analysis of the reprogramming of this epigenetic mark during early mammalian development.     

Mechanisms of histone H3 lysine 27 trimethylation remodeling during early mammalian development

During fertilization, two of the most differentiated cells in the mammalian organism, a sperm and oocyte, are combined to form a pluripotent embryo. Dynamic changes in chromatin structure allow the transition of the chromatin on these specialized cells into an embryonic configuration capable of generating every cell type. Initially, this reprogramming activity is supported by oocyte-derived factors accumulated during oogenesis as proteins and mRNAs; however, the underlying molecular mechanisms that govern it remain poorly characterized. Trimethylation of histone H3 at lysine 27 (H3K27me3) is a repressive epigenetic mark that changes dynamically during pre-implantation development in mice, bovine and pig embryos. Here we present data and hypotheses related to the potential mechanisms behind H3K27me3 remodeling during early development. We postulate that the repressive H3K27me3 mark is globally erased from the parental genomes in order to remove the gametic epigenetic program and to establish a pluripotent embryonic epigenome. We discuss information gathered in mice, pigs, and bovine, with the intent of providing a comparative analysis of the reprogramming of this epigenetic mark during early mammalian development.     

Parental epigenetic control of embryogenesis: a balance between inheritance and reprogramming?

At fertilization, fusion of two differentiated gametes forms the zygote that is capable of forming all of the varied cell lineages of an organism. It is widely thought that the acquisition of totipotency involves extensive epigenetic reprogramming of the germline state into an embryonic state. However, recent data argue that this reprogramming is incomplete and that substantial epigenetic information passes from one generation to the next. In this review we summarize the changes in chromatin states that take place during mammalian gametogenesis and examine the evidence that early mammalian embryogenesis may be affected by inheritance of epigenetic information from the parental generation.