RESUMO
The ChIP-seq signal of histone modifications at promoters is a good predictor of gene expression in different cellular contexts, but whether this is also true at enhancers is not clear. To address this issue, we develop quantitative models to characterize the relationship of gene expression with histone modifications at enhancers or promoters. We use embryonic stem cells (ESCs), which contain a full spectrum of active and repressed (poised) enhancers, to train predictive models. As many poised enhancers in ESCs switch towards an active state during differentiation, predictive models can also be trained on poised enhancers throughout differentiation and in development. Remarkably, we determine that histone modifications at enhancers, as well as promoters, are predictive of gene expression in ESCs and throughout differentiation and development. Importantly, we demonstrate that their contribution to the predictive models varies depending on their location in enhancers or promoters. Moreover, we use a local regression (LOESS) to normalize sequencing data from different sources, which allows us to apply predictive models trained in a specific cellular context to a different one. We conclude that the relationship between gene expression and histone modifications at enhancers is universal and different from promoters. Our study provides new insight into how histone modifications relate to gene expression based on their location in enhancers or promoters.
Assuntos
Elementos Facilitadores Genéticos , Expressão Gênica , Código das Histonas/genética , Modelos Genéticos , Regiões Promotoras Genéticas , Animais , Diferenciação Celular/genética , Células Cultivadas , Sequenciamento de Cromatina por Imunoprecipitação/estatística & dados numéricos , Biologia Computacional , Humanos , Camundongos , Células-Tronco Embrionárias Murinas/metabolismo , Análise de RegressãoRESUMO
Polycomb group proteins (PcG) are transcriptional repressors that control cell identity and development. In mammals, five different CBX proteins associate with the core Polycomb repressive complex 1 (PRC1). In mouse embryonic stem cells (ESCs), CBX6 and CBX7 are the most highly expressed CBX family members. CBX7 has been recently characterized, but little is known regarding the function of CBX6. Here, we show that CBX6 is essential for ESC identity. Its depletion destabilizes the pluripotency network and triggers differentiation. Mechanistically, we find that CBX6 is physically and functionally associated to both canonical PRC1 (cPRC1) and non-canonical PRC1 (ncPRC1) complexes. Notably, in contrast to CBX7, CBX6 is recruited to chromatin independently of H3K27me3. Taken together, our findings reveal that CBX6 is an essential component of ESC biology that contributes to the structural and functional complexity of the PRC1 complex.
Assuntos
Diferenciação Celular/genética , Perfilação da Expressão Gênica , Células-Tronco Embrionárias Murinas/metabolismo , Proteínas do Grupo Polycomb/genética , Animais , Células HEK293 , Humanos , Camundongos , Células-Tronco Embrionárias Murinas/citologia , Complexo Repressor Polycomb 1/genética , Complexo Repressor Polycomb 1/metabolismo , Proteínas do Grupo Polycomb/metabolismo , Interferência de RNARESUMO
Although the core subunits of Polycomb group (PcG) complexes are well characterized, little is known about the dynamics of these protein complexes during cellular differentiation. We used quantitative interaction proteomics and genome-wide profiling to study PcG proteins in mouse embryonic stem cells (ESCs) and neural progenitor cells (NPCs). We found that the stoichiometry and genome-wide binding of PRC1 and PRC2 were highly dynamic during neural differentiation. Intriguingly, we observed a downregulation and loss of PRC2 from chromatin marked with trimethylated histone H3 K27 (H3K27me3) during differentiation, whereas PRC1 was retained at these sites. Additionally, we found PRC1 at enhancer and promoter regions independently of PRC2 binding and H3K27me3. Finally, overexpression of NPC-specific PRC1 interactors in ESCs led to increased Ring1b binding to, and decreased expression of, NPC-enriched Ring1b-target genes. In summary, our integrative analyses uncovered dynamic PcG subcomplexes and their widespread colocalization with active chromatin marks during differentiation.
Assuntos
Diferenciação Celular/genética , Cromatina/metabolismo , Histonas/genética , Células-Tronco Embrionárias Murinas/metabolismo , Células-Tronco Neurais/metabolismo , Proteínas do Grupo Polycomb/genética , Animais , Linhagem Celular , Cromatina/química , Cromatografia Líquida , Cromossomos Artificiais Bacterianos , Regulação da Expressão Gênica , Estudo de Associação Genômica Ampla , Histonas/metabolismo , Camundongos , Células-Tronco Embrionárias Murinas/citologia , Células-Tronco Neurais/citologia , Proteínas do Grupo Polycomb/classificação , Proteínas do Grupo Polycomb/metabolismo , Mapeamento de Interação de Proteínas , Proteômica/métodos , Transdução de Sinais , Espectrometria de Massas em TandemRESUMO
Polycomb complexes (PRC1 and PRC2) are essential regulators of epigenetic gene silencing in embryonic and adult stem cells. Emerging evidence suggests that the core subunit composition regulates distinct biological processes, yet little is known about the mechanistic underpinnings of how differently composed Polycomb complexes instruct and maintain cell fate. Here we find that Mel18, also known as Pcgf2 and one of six Pcgf paralogs, uniquely regulates PRC1 to specify mesoderm cell fate in embryonic stem cells. Mechanistically, Mel18 functions as a classical Polycomb protein during early cardiac mesoderm differentiation by repressing pluripotency, lineage specification, late cardiac differentiation, and negative regulators of the BMP pathway. However, Mel18 also positively regulates expression of key mesoderm transcription factors, revealing an unexpected function of Mel18 in gene activation during cardiac differentiation. Taken together, our findings reveal that Mel18 is required to specify PRC1 function in both a context- and stage-specific manner.
Assuntos
Linhagem da Célula , Regulação da Expressão Gênica , Mesoderma/citologia , Células-Tronco Embrionárias Murinas/citologia , Proteínas do Grupo Polycomb/metabolismo , Animais , Proteínas Morfogenéticas Ósseas/metabolismo , Cartilagem/citologia , Diferenciação Celular , Corpos Embrioides/citologia , Genoma , Camundongos , Células-Tronco Embrionárias Murinas/metabolismo , Miocárdio/citologia , Miócitos Cardíacos/citologia , Complexo Repressor Polycomb 1/deficiência , Complexo Repressor Polycomb 1/metabolismo , Ligação Proteica , Estabilidade Proteica , Transdução de Sinais , Transcrição GênicaRESUMO
Embryonic stem cells (ESCs) are characterized by their ability to self-renew and to differentiate into all cell types of a given organism. Understanding the molecular mechanisms that govern the ESC state is of great interest not only for basic research-for instance, ESCs represent a perfect system to study cellular differentiation in vitro-but also for their potential implications in human health, as these mechanisms are likewise involved in cancer progression and could be exploited in regenerative medicine. In this minireview, we focus on the latest insights into the molecular mechanisms mediated by the pluripotency factors as well as their roles during differentiation. We also discuss recent advances in understanding the function of the epigenetic regulators, Polycomb and MLL complexes, in ESC biology.
Assuntos
Células-Tronco Embrionárias/citologia , Epigênese Genética , Regulação da Expressão Gênica no Desenvolvimento , Camundongos/genética , Células-Tronco Pluripotentes/citologia , Animais , Diferenciação Celular , Células-Tronco Embrionárias/metabolismo , Histona-Lisina N-Metiltransferase/genética , Histona-Lisina N-Metiltransferase/metabolismo , Proteínas de Homeodomínio/genética , Proteínas de Homeodomínio/metabolismo , Camundongos/embriologia , Proteína de Leucina Linfoide-Mieloide/genética , Proteína de Leucina Linfoide-Mieloide/metabolismo , Proteína Homeobox Nanog , Fator 3 de Transcrição de Octâmero/genética , Fator 3 de Transcrição de Octâmero/metabolismo , Células-Tronco Pluripotentes/metabolismo , Proteínas do Grupo Polycomb/genética , Proteínas do Grupo Polycomb/metabolismo , Fatores de Transcrição SOXB1/genética , Fatores de Transcrição SOXB1/metabolismo , Transdução de SinaisRESUMO
The molecular mechanisms underlying specification from embryonic stem cells (ESCs) and maintenance of neural progenitor cells (NPCs) are largely unknown. Recently, we reported that the Zuotin-related factor 1 (Zrf1) is necessary for chromatin displacement of the Polycomb-repressive complex 1 (PRC1). We found that Zrf1 is required for NPC specification from ESCs and that it promotes the expression of NPC markers, including the key regulator Pax6. Moreover, Zrf1 is essential to establish and maintain Wnt ligand expression levels, which are necessary for NPC self-renewal. Reactivation of proper Wnt signaling in Zrf1-depleted NPCs restores Pax6 expression and the self-renewal capacity. ESC-derived NPCs in vitro resemble most of the characteristics of the self-renewing NPCs located in the developing embryonic cortex, which are termed radial glial cells (RGCs). Depletion of Zrf1 in vivo impairs the expression of key self-renewal regulators and Wnt ligand genes in RGCs. Thus, we demonstrate that Zrf1 plays an essential role in NPC generation and maintenance.