RESUMO
Epigenetic mechanisms have been proposed to play crucial roles in mammalian development, but their precise functions are only partially understood. To investigate epigenetic regulation of embryonic development, we differentiated human embryonic stem cells into mesendoderm, neural progenitor cells, trophoblast-like cells, and mesenchymal stem cells and systematically characterized DNA methylation, chromatin modifications, and the transcriptome in each lineage. We found that promoters that are active in early developmental stages tend to be CG rich and mainly engage H3K27me3 upon silencing in nonexpressing lineages. By contrast, promoters for genes expressed preferentially at later stages are often CG poor and primarily employ DNA methylation upon repression. Interestingly, the early developmental regulatory genes are often located in large genomic domains that are generally devoid of DNA methylation in most lineages, which we termed DNA methylation valleys (DMVs). Our results suggest that distinct epigenetic mechanisms regulate early and late stages of ES cell differentiation.
Assuntos
Metilação de DNA , Células-Tronco Embrionárias/metabolismo , Epigenômica , Regulação da Expressão Gênica no Desenvolvimento , Animais , Diferenciação Celular , Cromatina/metabolismo , Ilhas de CpG , Células-Tronco Embrionárias/citologia , Histonas/metabolismo , Humanos , Metilação , Neoplasias/genética , Regiões Promotoras Genéticas , Peixe-Zebra/embriologiaRESUMO
Aggregation of the RNA-binding protein TAR DNA-binding protein 43 (TDP-43) is the key neuropathological feature of neurodegenerative diseases, including amyotrophic lateral sclerosis (ALS) and frontotemporal lobar degeneration (FTLD). In physiological conditions, TDP-43 is predominantly nuclear, forms oligomers, and is contained in biomolecular condensates assembled by liquid-liquid phase separation (LLPS). In disease, TDP-43 forms cytoplasmic or intranuclear inclusions. How TDP-43 transitions from physiological to pathological states remains poorly understood. Using a variety of cellular systems to express structure-based TDP-43 variants, including human neurons and cell lines with near-physiological expression levels, we show that oligomerization and RNA binding govern TDP-43 stability, splicing functionality, LLPS, and subcellular localization. Importantly, our data reveal that TDP-43 oligomerization is modulated by RNA binding. By mimicking the impaired proteasomal activity observed in ALS/FTLD patients, we found that monomeric TDP-43 forms inclusions in the cytoplasm, whereas its RNA binding-deficient counterpart aggregated in the nucleus. These differentially localized aggregates emerged via distinct pathways: LLPS-driven aggregation in the nucleus and aggresome-dependent inclusion formation in the cytoplasm. Therefore, our work unravels the origins of heterogeneous pathological species reminiscent of those occurring in TDP-43 proteinopathy patients.
Assuntos
Esclerose Lateral Amiotrófica , Degeneração Lobar Frontotemporal , Humanos , Esclerose Lateral Amiotrófica/metabolismo , Degeneração Lobar Frontotemporal/metabolismo , Proteínas de Ligação a DNA/metabolismo , Neurônios/metabolismo , RNA/genéticaRESUMO
The differentiation of follicular dendritic cells (FDC) is essential to the remarkable microanatomic plasticity of lymphoid follicles. Here we show that FDC arise from ubiquitous perivascular precursors (preFDC) expressing platelet-derived growth factor receptor ß (PDGFRß). PDGFRß-Cre-driven reporter gene recombination resulted in FDC labeling, whereas conditional ablation of PDGFRß(+)-derived cells abolished FDC, indicating that FDC originate from PDGFRß(+) cells. Lymphotoxin-α-overexpressing prion protein (PrP)(+) kidneys developed PrP(+) FDC after transplantation into PrP(-) mice, confirming that preFDC exist outside lymphoid organs. Adipose tissue-derived PDGFRß(+) stromal-vascular cells responded to FDC maturation factors and, when transplanted into lymphotoxin ß receptor (LTßR)(-) kidney capsules, differentiated into Mfge8(+)CD21/35(+)FcγRIIß(+)PrP(+) FDC capable of trapping immune complexes and recruiting B cells. Spleens of lymphocyte-deficient mice contained perivascular PDGFRß(+) FDC precursors whose expansion required both lymphoid tissue inducer (LTi) cells and lymphotoxin. The ubiquity of preFDC and their strategic location at blood vessels may explain the de novo generation of organized lymphoid tissue at sites of lymphocytic inflammation.
Assuntos
Vasos Sanguíneos/citologia , Células Dendríticas Foliculares/citologia , Baço/citologia , Células-Tronco/citologia , Animais , Linfócitos B/imunologia , Células Dendríticas Foliculares/imunologia , Células Dendríticas Foliculares/metabolismo , Centro Germinativo/citologia , Centro Germinativo/imunologia , Centro Germinativo/metabolismo , Inflamação/patologia , Células Matadoras Naturais/imunologia , Camundongos , Receptor beta de Fator de Crescimento Derivado de Plaquetas/metabolismo , Organismos Livres de Patógenos Específicos , Baço/metabolismoRESUMO
Nuclear pore complexes have recently been shown to play roles in gene activation; however their potential involvement in metazoan transcription remains unclear. Here we show that the nucleoporins Sec13, Nup98, and Nup88, as well as a group of FG-repeat nucleoporins, bind to the Drosophila genome at functionally distinct loci that often do not represent nuclear envelope contact sites. Whereas Nup88 localizes to silent loci, Sec13, Nup98, and a subset of FG-repeat nucleoporins bind to developmentally regulated genes undergoing transcription induction. Strikingly, RNAi-mediated knockdown of intranuclear Sec13 and Nup98 specifically inhibits transcription of their target genes and prevents efficient reactivation of transcription after heat shock, suggesting an essential role of NPC components in regulating complex gene expression programs of multicellular organisms.
Assuntos
Cromatina/metabolismo , Proteínas de Drosophila/metabolismo , Drosophila melanogaster/metabolismo , Regulação da Expressão Gênica , Complexo de Proteínas Formadoras de Poros Nucleares/metabolismo , Animais , Drosophila melanogaster/genética , Resposta ao Choque Térmico , RNA Polimerase II/metabolismo , Transcrição GênicaRESUMO
Naive CD4⺠T cells can differentiate into specific helper and regulatory T cell lineages in order to combat infection and disease. The correct response to cytokines and a controlled balance of these populations is critical for the immune system and the avoidance of autoimmune disorders. To investigate how early cell-fate commitment is regulated, we generated the first human genome-wide maps of histone modifications that reveal enhancer elements after 72 hr of in vitro polarization toward T helper 1 (Th1) and T helper 2 (Th2) cell lineages. Our analysis indicated that even at this very early time point, cell-specific gene regulation and enhancers were at work directing lineage commitment. Further examination of lineage-specific enhancers identified transcription factors (TFs) with known and unknown T cell roles as putative drivers of lineage-specific gene expression. Lastly, an integrative analysis of immunopathogenic-associated SNPs suggests a role for distal regulatory elements in disease etiology.
Assuntos
Cromatina/metabolismo , Histonas/metabolismo , Doenças do Sistema Imunitário/imunologia , Células Th1/imunologia , Células Th2/imunologia , Diferenciação Celular/genética , Linhagem da Célula/genética , Predisposição Genética para Doença , Estudo de Associação Genômica Ampla , Histonas/genética , Humanos , Doenças do Sistema Imunitário/genética , Polimorfismo de Nucleotídeo Único , Regiões Promotoras Genéticas , Equilíbrio Th1-Th2RESUMO
A correction in the paper by Seiboth et al. [(2018). J. Synchrotron Rad. 25, 108-115] is made.
RESUMO
The human cell nucleus serves as an important organelle holding the genetic blueprint for life. In this work, X-ray ptychography was applied to assess the masses of human cell nuclei using its unique phase shift information. Measurements were carried out at the I13-1 beamline at the Diamond Light Source that has extremely large transverse coherence properties. The ptychographic diffractive imaging approach allowed imaging of large structures that gave quantitative measurements of the phase shift in 2D projections. In this paper a modified ptychography algorithm that improves the quality of the reconstruction for weak scattering samples is presented. The application of this approach to calculate the mass of several human nuclei is also demonstrated.
Assuntos
Núcleo Celular/ultraestrutura , Microscopia de Contraste de Fase/métodos , Algoritmos , Humanos , Processamento de Imagem Assistida por Computador/métodos , Síncrotrons , Difração de Raios X , Raios XRESUMO
PARP inhibitors are used for treatment of tumors lacking function of the double-strand DNA break repair proteins BRCA1 or BRCA2 and are already approved for several cancer types. Thus, it is clinically crucial to determine germline as well as somatic BRCA1/2 mutations in those patients. The amplicon-based Oncomine BRCA1 and BRCA2 Assay is a test routinely used in diagnostics with FFPE specimens. The assay is validated for the detection of mutations, however, data on its performance in detecting large genomic rearrangements in FFPE tissue, is scarce. We cross-validated Oncomine BRCA1 and BRCA2 Assay in blood samples and/or FFPE tissue with multiplex ligation-dependent probe amplification (MLPA) for exon deletions and with OncoScan and an in-house hybridization-based target capture assay (MelArray) with a customized pipeline for the detection of loss of heterozygosity (LOH) and heterozygous versus complete gene loss. The Oncomine BRCA1 and BRCA2 Assay could detect both exon deletion and mono- and bi-allelic losses of the BRCA1/2 genes. We show that the therapeutically relevant large genomic rearrangements are reliably detected with the amplicon-based Oncomine BRCA1 and BRCA2 Assay in FFPE tumor tissue. Based on our data, we suggest tumor BRCA testing as standard diagnostic prescreening prior to germline BRCA testing.
Assuntos
Proteína BRCA1/genética , Proteína BRCA2/genética , Neoplasias/genética , Inibidores de Poli(ADP-Ribose) Polimerases/uso terapêutico , Quebras de DNA de Cadeia Dupla/efeitos dos fármacos , Reparo do DNA/efeitos dos fármacos , Feminino , Rearranjo Gênico/genética , Genoma Humano/genética , Humanos , Perda de Heterozigosidade/genética , Masculino , Mutação , Neoplasias/tratamento farmacológico , Neoplasias/patologiaRESUMO
Overall, neonatal cancer is uncommon. Because of its rarity and heterogeneity, diagnosis can be challenging. We report a unique case of a myoepithelial carcinoma in a 7 week old girl. Molecular diagnostic workup revealed a EWSR1-KLF15 gene fusion which was previously described in only six cases of myoepithelial tumors so far. All cases occurred in children and adolescents. To our knowledge, this is the first report of a congenital EWSR1-KLF15 fusion positive myoepithelial tumor in an infant.
Assuntos
Biomarcadores Tumorais/genética , Fusão Gênica , Fatores de Transcrição Kruppel-Like/genética , Mioepitelioma/genética , Proteína EWS de Ligação a RNA/genética , Neoplasias de Tecidos Moles/genética , Feminino , Humanos , Lactente , Mioepitelioma/diagnóstico , Mioepitelioma/patologia , Neoplasias de Tecidos Moles/diagnóstico , Neoplasias de Tecidos Moles/patologiaRESUMO
Pol II(G) is a distinct form of RNA polymerase II that contains the tightly associated Gdown1 polypeptide (encoded by POLR2M). Unlike Pol II, Pol II(G) is highly dependent upon Mediator for robust activator-dependent transcription in a biochemically defined in vitro system. Here, in vitro studies show that Gdown1 competes with TFIIF for binding to the RPB1 and RPB5 subunits of Pol II, thereby inhibiting an essential function of TFIIF in preinitiation complex assembly, but also that Mediator can actually facilitate Pol II(G) binding to the promoter prior to subsequent Mediator functions. Complementary ChIP and RNAi analyses reveal that Pol II(G) is recruited to promoter regions of subsets of actively transcribed genes, where it appears to restrict transcription. These and other results suggest that Pol II(G) may act to modulate some genes while simultaneously, as a poised (noninitiated) polymerase, setting the stage for Mediator-dependent enhancement of their activity.
Assuntos
Regulação da Expressão Gênica , RNA Polimerase II/fisiologia , Fatores de Transcrição TFII/fisiologia , Transcrição Gênica , Ligação Competitiva , RNA Polimerases Dirigidas por DNA/metabolismo , Células HeLa , Humanos , Complexo Mediador/metabolismo , Complexo Mediador/fisiologia , RNA Polimerase II/genética , RNA Polimerase II/metabolismo , Fatores de Transcrição TFII/genética , Fatores de Transcrição TFII/metabolismoRESUMO
BACKGROUND: Screening programmes for tuberculosis (TB) among immigrants rarely consider the heterogeneity of risk related to migrants' country of origin. We assess the performance of a large screening programme in asylum seekers by analysing (i) the difference in yield and numbers needed to screen (NNS) by country and WHO-reported TB burden, (ii) the possible impact of screening thresholds on sensitivity, and (iii) the value of WHO-estimated TB burden to improve the prediction accuracy of screening yield. METHODS: We combined individual data of 119,037 asylum seekers screened for TB in Germany (2002-2015) with TB estimates of the World Health Organization (WHO) (1990-2014) for their 81 countries of origin. Adjusted rate ratios (aRR) and 95% credible intervals (CrI) of the observed yield of screening were calculated in Bayesian Poisson regression models by categories of WHO-estimated TB incidence. We assessed changes in sensitivity depending on screening thresholds, used WHO TB estimates as prior information to predict TB in asylum seekers, and modelled country-specific probabilities of numbers needed to screen (NNS) conditional on different screening thresholds. RESULTS: The overall yield was 82 per 100,000 and the annual yield ranged from 44.1 to 279.7 per 100,000. Country-specific yields ranged from 10 (95%- CrI: 1-47) to 683 (95%-CrI: 306-1336) per 100,000 in Iraqi and Somali asylum seekers, respectively. The observed yield was higher in asylum seekers from countries with a WHO-estimated TB incidence > 50 relative to those from countries ≤50 per 100,000 (aRR: 4.17, 95%-CrI: 2.86-6.59). Introducing a threshold in the range of a WHO-estimated TB incidence of 50 and 100 per 100,000 resulted in the lowest "loss" in sensitivity. WHO's TB prevalence estimates improved prediction accuracy for eight of the 11 countries, and allowed modelling country-specific probabilities of NNS. CONCLUSIONS: WHO's TB data can inform the estimation of screening yield and thus be used to improve screening efficiency in asylum seekers. This may help to develop more targeted screening strategies by reducing uncertainty in estimates of expected country-specific yield, and identify thresholds with lowest loss in sensitivity. Further modelling studies are needed which combine clinical, diagnostic and country-specific parameters.
Assuntos
Emigrantes e Imigrantes/estatística & dados numéricos , Tuberculose/diagnóstico , Adolescente , Adulto , Idoso , Teorema de Bayes , Criança , Controle de Doenças Transmissíveis , Feminino , Alemanha , Humanos , Masculino , Programas de Rastreamento , Pessoa de Meia-Idade , Modelos Estatísticos , Prevalência , Saúde Pública , Refugiados , Tuberculose/epidemiologia , Organização Mundial da Saúde , Adulto JovemRESUMO
BACKGROUND: A comprehensive in-hospital patient management with reasonable and economic resource allocation is arguably the major challenge of health-care systems worldwide, especially in elderly, frail, and polymorbid patients. The need for patient management tools to improve the transition process and allocation of health care resources in routine clinical care particularly for the inpatient setting is obvious. To address these issues, a large prospective trial is warranted. METHODS: The "Integrative Hospital Treatment in Older patients to benchmark and improve Outcome and Length of stay" (In-HospiTOOL) study is an investigator-initiated, multicenter effectiveness trial to compare the effects of a novel in-hospital management tool on length of hospital stay, readmission rate, quality of care, and other clinical outcomes using a time-series model. The study aims to include approximately 35`000 polymorbid medical patients over an 18-month period, divided in an observation, implementation, and intervention phase. Detailed data on treatment and outcome of polymorbid medical patients during the in-hospital stay and after 30 days will be gathered to investigate differences in resource use, inter-professional collaborations and to establish representative benchmarking data to promote measurement and display of quality of care data across seven Swiss hospitals. The trial will inform whether the "In-HospiTOOL" optimizes inter-professional collaboration and thereby reduces length of hospital stay without harming subjective and objective patient-oriented outcome markers. DISCUSSION: Many of the current quality-mirroring tools do not reflect the real need and use of resources, especially in polymorbid and elderly patients. In addition, a validated tool for optimization of patient transition and discharge processes is still missing. The proposed multicenter effectiveness trial has potential to improve interprofessional collaboration and optimizes resource allocation from hospital admission to discharge. The results will enable inter-hospital comparison of transition processes and accomplish a benchmarking for inpatient care quality.
Assuntos
Benchmarking/normas , Múltiplas Afecções Crônicas/terapia , Adolescente , Adulto , Idoso , Ensaios Clínicos como Assunto , Pesquisa Comparativa da Efetividade , Atenção à Saúde/estatística & dados numéricos , Prestação Integrada de Cuidados de Saúde/normas , Hospitalização/estatística & dados numéricos , Humanos , Relações Interprofissionais , Tempo de Internação/estatística & dados numéricos , Pessoa de Meia-Idade , Estudos Multicêntricos como Assunto , Alta do Paciente/normas , Readmissão do Paciente/normas , Transferência de Pacientes/normas , Ensaios Clínicos Pragmáticos como Assunto , Estudos Prospectivos , Qualidade da Assistência à Saúde , Alocação de Recursos , Adulto JovemRESUMO
Wavefront errors of rotationally parabolic refractive X-ray lenses made of beryllium (Be CRLs) have been recovered for various lens sets and X-ray beam configurations. Due to manufacturing via an embossing process, aberrations of individual lenses within the investigated ensemble are very similar. By deriving a mean single-lens deformation for the ensemble, aberrations of any arbitrary lens stack can be predicted from the ensemble with \bar{\sigma} = 0.034λ. Using these findings the expected focusing performance of current Be CRLs are modeled for relevant X-ray energies and bandwidths and it is shown that a correction of aberrations can be realised without prior lens characterization but simply based on the derived lens deformation. The performance of aberration-corrected Be CRLs is discussed and the applicability of aberration-correction demonstrated over wide X-ray energy ranges.
RESUMO
Favorable-risk human acute myeloid leukemia (AML) engrafts poorly in currently used immunodeficient mice, possibly because of insufficient environmental support of these leukemic entities. To address this limitation, we here transplanted primary human AML with isolated nucleophosmin (NPM1) mutation and AML with inv(16) in mice in which human versions of genes encoding cytokines important for myelopoiesis (macrophage colony-stimulating factor [M-CSF], interleukin-3, granulocyte-macrophage colony-stimulating factor, and thrombopoietin) were knocked into their respective mouse loci. NPM1mut AML engrafted with higher efficacy in cytokine knock-in (KI) mice and showed a trend toward higher bone marrow engraftment levels in comparison with NSG mice. inv(16) AML engrafted with high efficacy and was serially transplantable in cytokine KI mice but, in contrast, exhibited virtually no engraftment in NSG mice. Selected use of cytokine KI mice revealed that human M-CSF was required for inv(16) AML engraftment. Subsequent transcriptome profiling in an independent AML patient study cohort demonstrated high expression of M-CSF receptor and enrichment of M-CSF inducible genes in inv(16) AML cases. This study thus provides a first xenotransplantation mouse model for and informs on the disease biology of inv(16) AML.
Assuntos
Modelos Animais de Doenças , Leucemia Mieloide Aguda , Transplante de Neoplasias/métodos , Transplante Heterólogo/métodos , Animais , Aberrações Cromossômicas , Cromossomos Humanos Par 16/genética , Citocinas , Técnicas de Introdução de Genes , Xenoenxertos , Humanos , Leucemia Mieloide Aguda/genética , Leucemia Mieloide Aguda/patologia , Camundongos , Mutação , Proteínas Nucleares/genética , NucleofosminaRESUMO
The laboratory mouse is the most widely used mammalian model organism in biomedical research. The 2.6 × 10(9) bases of the mouse genome possess a high degree of conservation with the human genome, so a thorough annotation of the mouse genome will be of significant value to understanding the function of the human genome. So far, most of the functional sequences in the mouse genome have yet to be found, and the cis-regulatory sequences in particular are still poorly annotated. Comparative genomics has been a powerful tool for the discovery of these sequences, but on its own it cannot resolve their temporal and spatial functions. Recently, ChIP-Seq has been developed to identify cis-regulatory elements in the genomes of several organisms including humans, Drosophila melanogaster and Caenorhabditis elegans. Here we apply the same experimental approach to a diverse set of 19 tissues and cell types in the mouse to produce a map of nearly 300,000 murine cis-regulatory sequences. The annotated sequences add up to 11% of the mouse genome, and include more than 70% of conserved non-coding sequences. We define tissue-specific enhancers and identify potential transcription factors regulating gene expression in each tissue or cell type. Finally, we show that much of the mouse genome is organized into domains of coordinately regulated enhancers and promoters. Our results provide a resource for the annotation of functional elements in the mammalian genome and for the study of mechanisms regulating tissue-specific gene expression.
Assuntos
Regulação da Expressão Gênica/genética , Genoma/genética , Camundongos/genética , Mapeamento Físico do Cromossomo , Sequências Reguladoras de Ácido Nucleico/genética , Acetilação , Animais , Cromatina/metabolismo , Imunoprecipitação da Cromatina , Sequência Conservada , Elementos Facilitadores Genéticos/genética , Evolução Molecular , Masculino , Metilação , Camundongos Endogâmicos C57BL , Anotação de Sequência Molecular , Motivos de Nucleotídeos , Especificidade de Órgãos , Regiões Promotoras Genéticas/genética , Análise de Sequência de DNA , Fatores de Transcrição/metabolismoRESUMO
Nuclear-architecture defects have been shown to correlate with the manifestation of a number of human diseases as well as ageing. It is therefore plausible that diseases whose manifestations correlate with ageing might be connected to the appearance of nuclear aberrations over time. We decided to evaluate nuclear organization in the context of ageing-associated disorders by focusing on a leucine-rich repeat kinase 2 (LRRK2) dominant mutation (G2019S; glycine-to-serine substitution at amino acid 2019), which is associated with familial and sporadic Parkinson's disease as well as impairment of adult neurogenesis in mice. Here we report on the generation of induced pluripotent stem cells (iPSCs) derived from Parkinson's disease patients and the implications of LRRK2(G2019S) mutation in human neural-stem-cell (NSC) populations. Mutant NSCs showed increased susceptibility to proteasomal stress as well as passage-dependent deficiencies in nuclear-envelope organization, clonal expansion and neuronal differentiation. Disease phenotypes were rescued by targeted correction of the LRRK2(G2019S) mutation with its wild-type counterpart in Parkinson's disease iPSCs and were recapitulated after targeted knock-in of the LRRK2(G2019S) mutation in human embryonic stem cells. Analysis of human brain tissue showed nuclear-envelope impairment in clinically diagnosed Parkinson's disease patients. Together, our results identify the nucleus as a previously unknown cellular organelle in Parkinson's disease pathology and may help to open new avenues for Parkinson's disease diagnoses as well as for the potential development of therapeutics targeting this fundamental cell structure.
Assuntos
Proteínas Mutantes/metabolismo , Células-Tronco Neurais/patologia , Doença de Parkinson/patologia , Proteínas Serina-Treonina Quinases/genética , Proteínas Serina-Treonina Quinases/metabolismo , Apoptose , Diferenciação Celular , Divisão Celular , Linhagem Celular , Células Clonais/metabolismo , Células Clonais/patologia , Células-Tronco Embrionárias/metabolismo , Células-Tronco Embrionárias/patologia , Técnicas de Introdução de Genes , Humanos , Células-Tronco Pluripotentes Induzidas/metabolismo , Células-Tronco Pluripotentes Induzidas/patologia , Serina-Treonina Proteína Quinase-2 com Repetições Ricas em Leucina , Proteínas Mutantes/genética , Mutação , Células-Tronco Neurais/metabolismo , Membrana Nuclear/genética , Membrana Nuclear/patologia , Complexo de Endopeptidases do Proteassoma/metabolismo , Estresse FisiológicoRESUMO
Both diffusible factors acting in trans and chromatin components acting in cis are implicated in gene regulation, but the extent to which either process causally determines a cell's transcriptional identity is unclear. We recently used cell fusion to define a class of silent genes termed "cis-silenced" (or "occluded") genes, which remain silent even in the presence of trans-acting transcriptional activators. We further showed that occlusion of lineage-inappropriate genes plays a critical role in maintaining the transcriptional identities of somatic cells. Here, we present, for the first time, a comprehensive map of occluded genes in somatic cells. Specifically, we mapped occluded genes in mouse fibroblasts via fusion to a dozen different rat cell types followed by whole-transcriptome profiling. We found that occluded genes are highly prevalent and stable in somatic cells, representing a sizeable fraction of silent genes. Occluded genes are also highly enriched for important developmental regulators of alternative lineages, consistent with the role of occlusion in safeguarding cell identities. Alongside this map, we also present whole-genome maps of DNA methylation and eight other chromatin marks. These maps uncover a complex relationship between chromatin state and occlusion. Furthermore, we found that DNA methylation functions as the memory of occlusion in a subset of occluded genes, while histone deacetylation contributes to the implementation but not memory of occlusion. Our data suggest that the identities of individual cell types are defined largely by the occlusion status of their genomes. The comprehensive reference maps reported here provide the foundation for future studies aimed at understanding the role of occlusion in development and disease.
Assuntos
Regulação da Expressão Gênica , Inativação Gênica , Sequências Reguladoras de Ácido Nucleico , Transativadores/genética , Transcrição Gênica , Animais , Fusão Celular , Linhagem Celular , Cromatina/genética , Metilação de DNA/genética , Genoma , Histonas/genética , Histonas/metabolismo , Camundongos , RatosRESUMO
Systematic annotation of gene regulatory elements is a major challenge in genome science. Direct mapping of chromatin modification marks and transcriptional factor binding sites genome-wide has successfully identified specific subtypes of regulatory elements. In Drosophila several pioneering studies have provided genome-wide identification of Polycomb response elements, chromatin states, transcription factor binding sites, RNA polymerase II regulation and insulator elements; however, comprehensive annotation of the regulatory genome remains a significant challenge. Here we describe results from the modENCODE cis-regulatory annotation project. We produced a map of the Drosophila melanogaster regulatory genome on the basis of more than 300 chromatin immunoprecipitation data sets for eight chromatin features, five histone deacetylases and thirty-eight site-specific transcription factors at different stages of development. Using these data we inferred more than 20,000 candidate regulatory elements and validated a subset of predictions for promoters, enhancers and insulators in vivo. We identified also nearly 2,000 genomic regions of dense transcription factor binding associated with chromatin activity and accessibility. We discovered hundreds of new transcription factor co-binding relationships and defined a transcription factor network with over 800 potential regulatory relationships.
Assuntos
Drosophila melanogaster/genética , Genoma de Inseto/genética , Anotação de Sequência Molecular , Sequências Reguladoras de Ácido Nucleico/genética , Animais , Cromatina/metabolismo , Montagem e Desmontagem da Cromatina , Imunoprecipitação da Cromatina , Elementos Facilitadores Genéticos/genética , Histona Desacetilases/metabolismo , Elementos Isolantes/genética , Regiões Promotoras Genéticas/genética , Reprodutibilidade dos Testes , Elementos Silenciadores Transcricionais/genética , Fatores de Transcrição/metabolismoRESUMO
During mammalian development, DNA methylation patterns need to be reset in primordial germ cells (PGCs) and preimplantation embryos. However, many LTR retrotransposons and imprinted genes are impervious to such global epigenetic reprogramming via hitherto undefined mechanisms. Here, we report that a subset of such genomic regions are resistant to widespread erasure of DNA methylation in mouse embryonic stem cells (mESCs) lacking the de novo DNA methyltransferases (Dnmts) Dnmt3a and Dnmt3b. Intriguingly, these loci are enriched for H3K9me3 in mESCs, implicating this mark in DNA methylation homeostasis. Indeed, deletion of the H3K9 methyltransferase SET domain bifurcated 1 (Setdb1) results in reduced H3K9me3 and DNA methylation levels at specific loci, concomitant with increased 5-hydroxymethylation (5hmC) and ten-eleven translocation 1 binding. Taken together, these data reveal that Setdb1 promotes the persistence of DNA methylation in mESCs, likely reflecting one mechanism by which DNA methylation is maintained at LTR retrotransposons and imprinted genes during developmental stages when DNA methylation is reprogrammed.
Assuntos
Metilação de DNA , Impressão Genômica , Histona-Lisina N-Metiltransferase/metabolismo , Retroelementos/genética , Animais , Células Cultivadas , DNA (Citosina-5-)-Metiltransferases/deficiência , DNA (Citosina-5-)-Metiltransferases/genética , DNA (Citosina-5-)-Metiltransferases/metabolismo , Metilação de DNA/genética , DNA Metiltransferase 3A , Células-Tronco Embrionárias/metabolismo , Retrovirus Endógenos/genética , Retrovirus Endógenos/metabolismo , Epigênese Genética , Feminino , Histona-Lisina N-Metiltransferase/deficiência , Histona-Lisina N-Metiltransferase/genética , Histonas/química , Histonas/genética , Histonas/metabolismo , Masculino , Camundongos , Camundongos Knockout , DNA Metiltransferase 3BRESUMO
We assessed evidence for a contextual effect of positive intergroup contact, whereby the effect of intergroup contact between social contexts (the between-level effect) on outgroup prejudice is greater than the effect of individual-level contact within contexts (the within-level effect). Across seven large-scale surveys (five cross-sectional and two longitudinal), using multilevel analyses, we found a reliable contextual effect. This effect was found in multiple countries, operationalizing context at multiple levels (regions, districts, and neighborhoods), and with and without controlling for a range of demographic and context variables. In four studies (three cross-sectional and one longitudinal) we showed that the association between context-level contact and prejudice was largely mediated by more tolerant norms. In social contexts where positive contact with outgroups was more commonplace, norms supported such positive interactions between members of different groups. Thus, positive contact reduces prejudice on a macrolevel, whereby people are influenced by the behavior of others in their social context, not merely on a microscale, via individuals' direct experience of positive contact with outgroup members. These findings reinforce the view that contact has a significant role to play in prejudice reduction, and has great policy potential as a means to improve intergroup relations, because it can simultaneously impact large numbers of people.