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1.
Mol Cell ; 84(3): 411-412, 2024 Feb 01.
Artigo em Inglês | MEDLINE | ID: mdl-38307001

RESUMO

In this issue of Molecular Cell, Matsui et al.1 examine lineage determination by pioneer transcription factors, finding that they control cell fate in cooperation with PRDM family members by repressing alternative-lineage and precocious gene expression through establishment of bivalent enhancers.


Assuntos
Fatores de Transcrição , Fatores de Transcrição/genética , Fatores de Transcrição/metabolismo , Linhagem da Célula/genética , Diferenciação Celular/genética
2.
Nucleic Acids Res ; 52(7): 3607-3622, 2024 Apr 24.
Artigo em Inglês | MEDLINE | ID: mdl-38281186

RESUMO

Biologically precise enhancer licensing by lineage-determining transcription factors enables activation of transcripts appropriate to biological demand and prevents deleterious gene activation. This essential process is challenged by the millions of matches to most transcription factor binding motifs present in many eukaryotic genomes, leading to questions about how transcription factors achieve the exquisite specificity required. The importance of chromatin remodeling factors to enhancer activation is highlighted by their frequent mutation in developmental disorders and in cancer. Here, we determine the roles of CHD4 in enhancer licensing and maintenance in breast cancer cells and during cellular reprogramming. In unchallenged basal breast cancer cells, CHD4 modulates chromatin accessibility. Its depletion leads to redistribution of transcription factors to previously unoccupied sites. During cellular reprogramming induced by the pioneer factor GATA3, CHD4 activity is necessary to prevent inappropriate chromatin opening. Mechanistically, CHD4 promotes nucleosome positioning over GATA3 binding motifs to compete with transcription factor-DNA interaction. We propose that CHD4 acts as a chromatin proof-reading enzyme that prevents unnecessary gene expression by editing chromatin binding activities of transcription factors.


Assuntos
Cromatina , Complexo Mi-2 de Remodelação de Nucleossomo e Desacetilase , Feminino , Humanos , Sítios de Ligação , Neoplasias da Mama/genética , Neoplasias da Mama/metabolismo , Linhagem Celular Tumoral , Reprogramação Celular/genética , Cromatina/metabolismo , Montagem e Desmontagem da Cromatina , Elementos Facilitadores Genéticos , Fator de Transcrição GATA3/metabolismo , Fator de Transcrição GATA3/genética , Complexo Mi-2 de Remodelação de Nucleossomo e Desacetilase/metabolismo , Complexo Mi-2 de Remodelação de Nucleossomo e Desacetilase/genética , Nucleossomos/metabolismo , Nucleossomos/genética , Ligação Proteica , Fatores de Transcrição/metabolismo
3.
Nature ; 551(7678): 105-109, 2017 11 02.
Artigo em Inglês | MEDLINE | ID: mdl-29072299

RESUMO

T helper 17 (TH17) cells are critically involved in host defence, inflammation, and autoimmunity. Transforming growth factor ß (TGFß) is instrumental in TH17 cell differentiation by cooperating with interleukin-6 (refs 6, 7). Yet, the mechanism by which TGFß enables TH17 cell differentiation remains elusive. Here we reveal that TGFß enables TH17 cell differentiation by reversing SKI-SMAD4-mediated suppression of the expression of the retinoic acid receptor (RAR)-related orphan receptor γt (RORγt). We found that, unlike wild-type T cells, SMAD4-deficient T cells differentiate into TH17 cells in the absence of TGFß signalling in a RORγt-dependent manner. Ectopic SMAD4 expression suppresses RORγt expression and TH17 cell differentiation of SMAD4-deficient T cells. However, TGFß neutralizes SMAD4-mediated suppression without affecting SMAD4 binding to the Rorc locus. Proteomic analysis revealed that SMAD4 interacts with SKI, a transcriptional repressor that is degraded upon TGFß stimulation. SKI controls histone acetylation and deacetylation of the Rorc locus and TH17 cell differentiation via SMAD4: ectopic SKI expression inhibits H3K9 acetylation of the Rorc locus, Rorc expression, and TH17 cell differentiation in a SMAD4-dependent manner. Therefore, TGFß-induced disruption of SKI reverses SKI-SMAD4-mediated suppression of RORγt to enable TH17 cell differentiation. This study reveals a critical mechanism by which TGFß controls TH17 cell differentiation and uncovers the SKI-SMAD4 axis as a potential therapeutic target for treating TH17-related diseases.


Assuntos
Diferenciação Celular , Proteínas de Ligação a DNA/metabolismo , Proteínas Proto-Oncogênicas/metabolismo , Proteínas Repressoras/metabolismo , Proteína Smad4/metabolismo , Células Th17/citologia , Células Th17/metabolismo , Fator de Crescimento Transformador beta/metabolismo , Animais , Diferenciação Celular/genética , Feminino , Deleção de Genes , Humanos , Interleucina-6/metabolismo , Masculino , Camundongos , Complexos Multiproteicos/química , Complexos Multiproteicos/metabolismo , Membro 3 do Grupo F da Subfamília 1 de Receptores Nucleares/deficiência , Membro 3 do Grupo F da Subfamília 1 de Receptores Nucleares/genética , Membro 3 do Grupo F da Subfamília 1 de Receptores Nucleares/metabolismo , Receptores de Fatores de Crescimento Transformadores beta/metabolismo , Transdução de Sinais , Proteína Smad4/deficiência , Proteína Smad4/genética
4.
Proc Natl Acad Sci U S A ; 117(31): 18439-18447, 2020 08 04.
Artigo em Inglês | MEDLINE | ID: mdl-32675241

RESUMO

In mammals, repressive histone modifications such as trimethylation of histone H3 Lys9 (H3K9me3), frequently coexist with DNA methylation, producing a more stable and silenced chromatin state. However, it remains elusive how these epigenetic modifications crosstalk. Here, through structural and biochemical characterizations, we identified the replication foci targeting sequence (RFTS) domain of maintenance DNA methyltransferase DNMT1, a module known to bind the ubiquitylated H3 (H3Ub), as a specific reader for H3K9me3/H3Ub, with the recognition mode distinct from the typical trimethyl-lysine reader. Disruption of the interaction between RFTS and the H3K9me3Ub affects the localization of DNMT1 in stem cells and profoundly impairs the global DNA methylation and genomic stability. Together, this study reveals a previously unappreciated pathway through which H3K9me3 directly reinforces DNMT1-mediated maintenance DNA methylation.


Assuntos
DNA (Citosina-5-)-Metiltransferase 1/metabolismo , Metilação de DNA , Heterocromatina/metabolismo , Histonas/metabolismo , DNA (Citosina-5-)-Metiltransferase 1/genética , Heterocromatina/genética , Histonas/química , Histonas/genética , Humanos , Lisina/genética , Lisina/metabolismo , Metilação , Processamento de Proteína Pós-Traducional
5.
Nucleic Acids Res ; 48(9): 4756-4768, 2020 05 21.
Artigo em Inglês | MEDLINE | ID: mdl-32232341

RESUMO

Estrogen receptors (ER) are activated by the steroid hormone 17ß-estradiol. Estrogen receptor alpha (ER-α) forms a regulatory network in mammary epithelial cells and in breast cancer with the transcription factors FOXA1 and GATA3. GATA3 is one of the most frequently mutated genes in breast cancer and is capable of specifying chromatin localization of FOXA1 and ER-α. How GATA3 mutations found in breast cancer impact genomic localization of ER-α and the transcriptional network downstream of ER-α and FOXA1 remains unclear. Here, we investigate the function of a recurrent patient-derived GATA3 mutation (R330fs) on this regulatory network. Genomic analysis indicates that the R330fs mutant can disrupt localization of ER-α and FOXA1. Loci co-bound by all three factors are enriched for genes integral to mammary gland development as well as epithelial cell biology. This gene set is differentially regulated in GATA3 mutant cells in culture and in tumors bearing similar mutations in vivo. The altered distribution of ER-α and FOXA1 in GATA3-mutant cells is associated with altered chromatin architecture, which leads to differential gene expression. These results suggest an active role for GATA3 zinc finger 2 mutants in ER-α positive breast tumors.


Assuntos
Neoplasias da Mama/genética , Receptor alfa de Estrogênio/metabolismo , Fator de Transcrição GATA3/genética , Fator de Transcrição GATA3/metabolismo , Regulação Neoplásica da Expressão Gênica , Redes Reguladoras de Genes , Fator 3-alfa Nuclear de Hepatócito/metabolismo , Neoplasias da Mama/metabolismo , Linhagem Celular Tumoral , Cromatina/metabolismo , Feminino , Humanos , Mutação , Transcrição Gênica
6.
Adv Exp Med Biol ; 1389: 269-293, 2022.
Artigo em Inglês | MEDLINE | ID: mdl-36350514

RESUMO

Covalent modification of DNA via deposition of a methyl group at the 5' position on cytosine residues alters the chemical groups available for interaction in the major groove of DNA. This modification, thereby, alters the affinity and specificity of DNA-binding proteins; some of them favor interaction with methylated DNA, and others disfavor it. Molecular recognition of cytosine methylation by proteins often initiates sequential regulatory events that impact gene expression and chromatin structure. The known methyl-DNA-binding proteins have unique domains responsible for DNA methylation recognition: (1) the methyl-CpG-binding domain (MBD), (2) the SET- and RING finger-associated domain (SRA), and (3) some of TF families, such as the C2H2 zinc finger domain, basic helix-loop-helix (bHLH), basic leucine-zipper (bZIP), and homeodomain proteins. Structural analyses have revealed that each domain has a characteristic methylated DNA-binding pattern, and the difference in the recognition mechanisms renders the DNA methylation mark able to transmit complicated biological information. Recent genetic and genomic studies have revealed novel functions of methyl-DNA-binding proteins. These emerging data have also provided glimpses into how methyl-DNA-binding proteins possess unique features and, presumably, functions. In this chapter, we summarize structural and biochemical analyses elucidating the mechanisms for recognition of DNA methylation and correlate this information with emerging genomic and functional data.


Assuntos
Citosina , Metilação de DNA , Humanos , Citosina/química , Proteínas de Ligação a DNA/metabolismo , DNA/metabolismo , Domínios Proteicos , Ilhas de CpG/genética
7.
Proc Biol Sci ; 288(1949): 20202718, 2021 04 28.
Artigo em Inglês | MEDLINE | ID: mdl-33878919

RESUMO

A key goal of conservation is to protect biodiversity by supporting the long-term persistence of viable, natural populations of wild species. Conservation practice has long been guided by genetic, ecological and demographic indicators of risk. Emerging evidence of animal culture across diverse taxa and its role as a driver of evolutionary diversification, population structure and demographic processes may be essential for augmenting these conventional conservation approaches and decision-making. Animal culture was the focus of a ground-breaking resolution under the Convention on the Conservation of Migratory Species of Wild Animals (CMS), an international treaty operating under the UN Environment Programme. Here, we synthesize existing evidence to demonstrate how social learning and animal culture interact with processes important to conservation management. Specifically, we explore how social learning might influence population viability and be an important resource in response to anthropogenic change, and provide examples of how it can result in phenotypically distinct units with different, socially learnt behavioural strategies. While identifying culture and social learning can be challenging, indirect identification and parsimonious inferences may be informative. Finally, we identify relevant methodologies and provide a framework for viewing behavioural data through a cultural lens which might provide new insights for conservation management.


Assuntos
Biodiversidade , Conservação dos Recursos Naturais , Animais , Animais Selvagens , Evolução Biológica , Aprendizagem
8.
Mol Ecol ; 30(23): 6162-6177, 2021 12.
Artigo em Inglês | MEDLINE | ID: mdl-34416064

RESUMO

Runs of homozygosity (ROH) occur when offspring inherit haplotypes that are identical by descent from each parent. Length distributions of ROH are informative about population history; specifically, the probability of inbreeding mediated by mating system and/or population demography. Here, we investigated whether variation in killer whale (Orcinus orca) demographic history is reflected in genome-wide heterozygosity and ROH length distributions, using a global data set of 26 genomes representative of geographic and ecotypic variation in this species, and two F1 admixed individuals with Pacific-Atlantic parentage. We first reconstructed demographic history for each population as changes in effective population size through time using the pairwise sequential Markovian coalescent (PSMC) method. We found a subset of populations declined in effective population size during the Late Pleistocene, while others had more stable demography. Genomes inferred to have undergone ancestral declines in effective population size, were autozygous at hundreds of short ROH (<1 Mb), reflecting high background relatedness due to coalescence of haplotypes deep within the pedigree. In contrast, longer and therefore younger ROH (>1.5 Mb) were found in low latitude populations, and populations of known conservation concern. These include a Scottish killer whale, for which 37.8% of the autosomes were comprised of ROH >1.5 Mb in length. The fate of this population, in which only two adult males have been sighted in the past five years, and zero fecundity over the last two decades, may be inextricably linked to its demographic history and consequential inbreeding depression.


Assuntos
Orca , Animais , Genoma , Homozigoto , Endogamia , Masculino , Polimorfismo de Nucleotídeo Único , Densidade Demográfica , Orca/genética
10.
Proc Natl Acad Sci U S A ; 115(26): 6727-6732, 2018 06 26.
Artigo em Inglês | MEDLINE | ID: mdl-29891665

RESUMO

Cardiac development relies on proper cardiomyocyte differentiation, including expression and assembly of cell-type-specific actomyosin subunits into a functional cardiac sarcomere. Control of this process involves not only promoting expression of cardiac sarcomere subunits but also repressing expression of noncardiac myofibril paralogs. This level of transcriptional control requires broadly expressed multiprotein machines that modify and remodel the chromatin landscape to restrict transcription machinery access. Prominent among these is the nucleosome remodeling and deacetylase (NuRD) complex, which includes the catalytic core subunit CHD4. Here, we demonstrate that direct CHD4-mediated repression of skeletal and smooth muscle myofibril isoforms is required for normal cardiac sarcomere formation, function, and embryonic survival early in gestation. Through transcriptomic and genome-wide analyses of CHD4 localization, we identified unique CHD4 binding sites in smooth muscle myosin heavy chain, fast skeletal α-actin, and the fast skeletal troponin complex genes. We further demonstrate that in the absence of CHD4, cardiomyocytes in the developing heart form a hybrid muscle cell that contains cardiac, skeletal, and smooth muscle myofibril components. These misexpressed paralogs intercalate into the nascent cardiac sarcomere to disrupt sarcomere formation and cause impaired cardiac function in utero. These results demonstrate the genomic and physiological requirements for CHD4 in mammalian cardiac development.


Assuntos
DNA Helicases/fisiologia , Regulação da Expressão Gênica no Desenvolvimento , Cardiopatias Congênitas/genética , Complexo Mi-2 de Remodelação de Nucleossomo e Desacetilase/fisiologia , Miócitos Cardíacos/fisiologia , Sarcômeros/fisiologia , Animais , DNA Helicases/química , DNA Helicases/deficiência , Feminino , Técnicas de Silenciamento de Genes , Genes Letais , Coração/diagnóstico por imagem , Coração/embriologia , Cardiopatias Congênitas/diagnóstico por imagem , Cardiopatias Congênitas/embriologia , Cardiopatias Congênitas/patologia , Masculino , Camundongos , Proteínas Musculares/biossíntese , Proteínas Musculares/genética , Miofibrilas/metabolismo , Miofibrilas/patologia , Nucleossomos/metabolismo , Nucleossomos/ultraestrutura , Sarcômeros/ultraestrutura , Transcrição Gênica , Ultrassonografia Pré-Natal
11.
Proc Natl Acad Sci U S A ; 115(18): E4189-E4198, 2018 05 01.
Artigo em Inglês | MEDLINE | ID: mdl-29666266

RESUMO

Early transient developmental exposure to an endocrine active compound, diethylstilbestrol (DES), a synthetic estrogen, causes late-stage effects in the reproductive tract of adult mice. Estrogen receptor alpha (ERα) plays a role in mediating these developmental effects. However, the developmental mechanism is not well known in male tissues. Here, we present genome-wide transcriptome and DNA methylation profiling of the seminal vesicles (SVs) during normal development and after DES exposure. ERα mediates aberrations of the mRNA transcriptome in SVs of adult mice following neonatal DES exposure. This developmental exposure impacts differential diseases between male (SVs) and female (uterus) tissues when mice reach adulthood due to most DES-altered genes that appear to be tissue specific during mouse development. Certain estrogen-responsive gene changes in SVs are cell-type specific. DNA methylation dynamically changes during development in the SVs of wild-type (WT) and ERα-knockout (αERKO) mice, which increases both the loss and gain of differentially methylated regions (DMRs). There are more gains of DMRs in αERKO compared with WT. Interestingly, the methylation changes between the two genotypes are in different genomic loci. Additionally, the expression levels of a subset of DES-altered genes are associated with their DNA methylation status following developmental DES exposure. Taken together, these findings provide an important basis for understanding the molecular and cellular mechanism of endocrine-disrupting chemicals (EDCs), such as DES, during development in the male mouse tissues. This unique evidence contributes to our understanding of developmental actions of EDCs in human health.


Assuntos
Metilação de DNA/efeitos dos fármacos , Dietilestilbestrol/efeitos adversos , Receptor alfa de Estrogênio/metabolismo , Estrogênios não Esteroides/efeitos adversos , Regulação da Expressão Gênica/efeitos dos fármacos , Glândulas Seminais/metabolismo , Transcriptoma/efeitos dos fármacos , Animais , Metilação de DNA/genética , Dietilestilbestrol/farmacologia , Receptor alfa de Estrogênio/agonistas , Receptor alfa de Estrogênio/genética , Estrogênios não Esteroides/farmacologia , Loci Gênicos , Masculino , Camundongos , Camundongos Knockout
13.
Genet Med ; 22(2): 389-397, 2020 02.
Artigo em Inglês | MEDLINE | ID: mdl-31388190

RESUMO

PURPOSE: Sifrim-Hitz-Weiss syndrome (SIHIWES) is a recently described multisystemic neurodevelopmental disorder caused by de novo variants inCHD4. In this study, we investigated the clinical spectrum of the disorder, genotype-phenotype correlations, and the effect of different missense variants on CHD4 function. METHODS: We collected clinical and molecular data from 32 individuals with mostly de novo variants in CHD4, identified through next-generation sequencing. We performed adenosine triphosphate (ATP) hydrolysis and nucleosome remodeling assays on variants from five different CHD4 domains. RESULTS: The majority of participants had global developmental delay, mild to moderate intellectual disability, brain anomalies, congenital heart defects, and dysmorphic features. Macrocephaly was a frequent but not universal finding. Additional common abnormalities included hypogonadism in males, skeletal and limb anomalies, hearing impairment, and ophthalmic abnormalities. The majority of variants were nontruncating and affected the SNF2-like region of the protein. We did not identify genotype-phenotype correlations based on the type or location of variants. Alterations in ATP hydrolysis and chromatin remodeling activities were observed in variants from different domains. CONCLUSION: The CHD4-related syndrome is a multisystemic neurodevelopmental disorder. Missense substitutions in different protein domains alter CHD4 function in a variant-specific manner, but result in a similar phenotype in humans.


Assuntos
Complexo Mi-2 de Remodelação de Nucleossomo e Desacetilase/genética , Transtornos do Neurodesenvolvimento/genética , Anormalidades Múltiplas/genética , Adolescente , Adulto , Criança , Pré-Escolar , Montagem e Desmontagem da Cromatina/genética , Deficiências do Desenvolvimento/genética , Feminino , Estudos de Associação Genética , Genótipo , Perda Auditiva/genética , Cardiopatias Congênitas/genética , Humanos , Lactente , Recém-Nascido , Deficiência Intelectual/genética , Masculino , Megalencefalia/genética , Complexo Mi-2 de Remodelação de Nucleossomo e Desacetilase/metabolismo , Anormalidades Musculoesqueléticas/genética , Mutação de Sentido Incorreto/genética , Fenótipo , Síndrome , Fatores de Transcrição/genética
14.
Mol Cell ; 45(6): 708-9, 2012 Mar 30.
Artigo em Inglês | MEDLINE | ID: mdl-22464440

RESUMO

In this issue of Molecular Cell, Ginno et al. (2012) describe unusual sequence features at promoter CpG islands that can lead to formation of persistent RNA-DNA hybrids (R loops), which are proposed to prevent genomic DNA methylation.

15.
Mol Ecol ; 28(2): 484-502, 2019 01.
Artigo em Inglês | MEDLINE | ID: mdl-30187987

RESUMO

Recent exploration into the interactions and relationship between hosts and their microbiota has revealed a connection between many aspects of the host's biology, health and associated micro-organisms. Whereas amplicon sequencing has traditionally been used to characterize the microbiome, the increasing number of published population genomics data sets offers an underexploited opportunity to study microbial profiles from the host shotgun sequencing data. Here, we use sequence data originally generated from killer whale Orcinus orca skin biopsies for population genomics, to characterize the skin microbiome and investigate how host social and geographical factors influence the microbial community composition. Having identified 845 microbial taxa from 2.4 million reads that did not map to the killer whale reference genome, we found that both ecotypic and geographical factors influence community composition of killer whale skin microbiomes. Furthermore, we uncovered key taxa that drive the microbiome community composition and showed that they are embedded in unique networks, one of which is tentatively linked to diatom presence and poor skin condition. Community composition differed between Antarctic killer whales with and without diatom coverage, suggesting that the previously reported episodic migrations of Antarctic killer whales to warmer waters associated with skin turnover may control the effects of potentially pathogenic bacteria such as Tenacibaculum dicentrarchi. Our work demonstrates the feasibility of microbiome studies from host shotgun sequencing data and highlights the importance of metagenomics in understanding the relationship between host and microbial ecology.


Assuntos
Metagenômica , Microbiota/genética , Pele/microbiologia , Orca/microbiologia , Animais , Regiões Antárticas , Diatomáceas/genética , Geografia , Orca/parasitologia
16.
Mol Ecol ; 28(14): 3427-3444, 2019 07.
Artigo em Inglês | MEDLINE | ID: mdl-31131963

RESUMO

Reconstruction of the demographic and evolutionary history of populations assuming a consensus tree-like relationship can mask more complex scenarios, which are prevalent in nature. An emerging genomic toolset, which has been most comprehensively harnessed in the reconstruction of human evolutionary history, enables molecular ecologists to elucidate complex population histories. Killer whales have limited extrinsic barriers to dispersal and have radiated globally, and are therefore a good candidate model for the application of such tools. Here, we analyse a global data set of killer whale genomes in a rare attempt to elucidate global population structure in a nonhuman species. We identify a pattern of genetic homogenisation at lower latitudes and the greatest differentiation at high latitudes, even between currently sympatric lineages. The processes underlying the major axis of structure include high drift at the edge of species' range, likely associated with founder effects and allelic surfing during postglacial range expansion. Divergence between Antarctic and non-Antarctic lineages is further driven by ancestry segments with up to four-fold older coalescence time than the genome-wide average; relicts of a previous vicariance during an earlier glacial cycle. Our study further underpins that episodic gene flow is ubiquitous in natural populations, and can occur across great distances and after substantial periods of isolation between populations. Thus, understanding the evolutionary history of a species requires comprehensive geographic sampling and genome-wide data to sample the variation in ancestry within individuals.


Assuntos
Fluxo Gênico , Genoma , Orca/genética , Alelos , Animais , Regiões Antárticas , Sequência de Bases , Núcleo Celular/genética , DNA Mitocondrial/genética , Deriva Genética , Variação Genética , Geografia , Cadeias de Markov , Modelos Genéticos , Filogenia , Análise de Componente Principal
17.
Nucleic Acids Res ; 45(22): 12723-12738, 2017 Dec 15.
Artigo em Inglês | MEDLINE | ID: mdl-29040764

RESUMO

Transposable elements, including endogenous retroviruses (ERVs), constitute a large fraction of the mammalian genome. They are transcriptionally silenced during early development to protect genome integrity and aberrant transcription. However, the mechanisms that control their repression are not fully understood. To systematically study ERV repression, we carried out an RNAi screen in mouse embryonic stem cells (ESCs) and identified a list of novel regulators. Among them, Rif1 displays the strongest effect. Rif1 depletion by RNAi or gene deletion led to increased transcription and increased chromatin accessibility at ERV regions and their neighboring genes. This transcriptional de-repression becomes more severe when DNA methylation is lost. On the mechanistic level, Rif1 directly occupies ERVs and is required for repressive histone mark H3K9me3 and H3K27me3 assembly and DNA methylation. It interacts with histone methyltransferases and facilitates their recruitment to ERV regions. Importantly, Rif1 represses ERVs in human ESCs as well, and the evolutionally-conserved HEAT-like domain is essential for its function. Finally, Rif1 acts as a barrier during somatic cell reprogramming, and its depletion significantly enhances reprogramming efficiency. Together, our study uncovered many previously uncharacterized repressors of ERVs, and defined an essential role of Rif1 in the epigenetic defense against ERV activation.


Assuntos
Cromatina/genética , Retrovirus Endógenos/genética , Proteínas de Ligação a Telômeros/genética , Ativação Viral , Animais , Linhagem Celular , Células Cultivadas , Cromatina/metabolismo , Metilação de DNA , Células-Tronco Embrionárias/metabolismo , Retrovirus Endógenos/fisiologia , Células HEK293 , Histona Metiltransferases , Histona-Lisina N-Metiltransferase/metabolismo , Histonas/metabolismo , Humanos , Metilação , Camundongos , Interferência de RNA , Proteínas de Ligação a Telômeros/metabolismo
18.
J Pharmacol Exp Ther ; 364(2): 156-169, 2018 02.
Artigo em Inglês | MEDLINE | ID: mdl-29180358

RESUMO

Patients with chronic constipation benefit from treatment with 5-hydroxytryptamine 4 (5-HT4) receptor agonists. However, the first-generation 5-HT4 receptor agonists cisapride and tegaserod were withdrawn from the market owing to rare cardiovascular adverse events that were not 5-HT4-receptor-related but due to the lack of selectivity of these drugs. Here we report the nonclinical cardiovascular profile of the selective 5-HT4 receptor agonist prucalopride. To assess its non-5-HT4 receptor-mediated effects on cardiovascular electrophysiological parameters, in vitro studies were performed in human ether-à-go-go-related gene-transfected cells, guinea pig ventricular myocytes and papillary muscle preparations, rabbit and dog Purkinje fibers, and the Langendorff rabbit heart. In vivo experiments were performed in a rabbit model for drug-induced proarrhythmogenesis, in anesthetized guinea pigs, and anesthetized and conscious dogs. In addition, human platelet aggregation and coronary artery contraction were studied to exclude interactions that have been suggested to mediate the cardiovascular effects of tegaserod. Effects at 5-HT4 receptors were evaluated in piglet and human atrial myocardium, and in anesthetized pigs. Finally, cardiovascular endpoints were investigated in chronic, repeated-dose toxicology studies at very high prucalopride doses in rats and dogs. No relevant effects were observed in any of the cardiovascular studies at concentrations at least 50 times the therapeutic plasma level. Only in pigs were minor and transient increases in heart rate and blood pressure noted upon first exposure to prucalopride, at plasma levels at least 10 times higher than human therapeutic plasma levels. Prucalopride may thus provide therapeutic benefit without the cardiovascular risks reported for other 5-HT4 receptor agonists.


Assuntos
Benzofuranos/farmacologia , Sistema Cardiovascular/efeitos dos fármacos , Receptores 5-HT4 de Serotonina/metabolismo , Agonistas do Receptor 5-HT4 de Serotonina/farmacologia , Animais , Cães , Relação Dose-Resposta a Droga , Fenômenos Eletrofisiológicos/efeitos dos fármacos , Cobaias , Células HEK293 , Átrios do Coração/citologia , Frequência Cardíaca/efeitos dos fármacos , Humanos , Contração Miocárdica/efeitos dos fármacos , Miocárdio/metabolismo , Coelhos
19.
Int J Toxicol ; 36(3): 229-238, 2017.
Artigo em Inglês | MEDLINE | ID: mdl-28466692

RESUMO

DE-71, a commercial mixture of polybrominated diphenyl ethers widely used in flame retardants, is a pervasive environmental contaminant due to its continuing release from waste material and its long half-life in humans. Although the genotoxic potential of DE-71 appears to be low based on bacterial mutagenicity, it remains a public health concern due to its reported involvement in tumor development. Molecular mechanisms by which DE-71 influences tumor incidence or progression remain understudied. We used liver carcinoma tissue from mice exposed to DE-71 to test the hypothesis that epigenetic alterations consistent with tumor development, specifically DNA methylation, result from long-term DE-71 exposure. We profiled DNA methylation status using the methylated-CpG island recovery assay coupled with microarray analysis of hepatocellular carcinoma DNA from animals exposed to DE-71. DE-71 exposure had little impact on global DNA methylation. However, we detected gene body-specific hypomethylation within the Tbx3 locus, a transcription factor important in liver tumorigenesis and in embryonic and cancer stem cell proliferation. This nonpromoter hypomethylation was accompanied by upregulation of Tbx3 mRNA and protein and by alterations in downstream cell cycle-associated marker expression. Thus, exposure to DE-71 may facilitate tumor development by inducing epigenetic programs that favor expansion of progenitor cell populations.


Assuntos
Metilação de DNA/efeitos dos fármacos , Retardadores de Chama/toxicidade , Éteres Difenil Halogenados/toxicidade , Proteínas com Domínio T/genética , Animais , Fígado/efeitos dos fármacos , Fígado/metabolismo , Fígado/patologia , Masculino , Camundongos , Proteínas com Domínio T/metabolismo
20.
EMBO J ; 31(2): 330-50, 2012 Jan 18.
Artigo em Inglês | MEDLINE | ID: mdl-22085927

RESUMO

Insulators help separate active chromatin domains from silenced ones. In yeast, gene promoters act as insulators to block the spread of Sir and HP1 mediated silencing while in metazoans most insulators are multipartite autonomous entities. tDNAs are repetitive sequences dispersed throughout the human genome and we now show that some of these tDNAs can function as insulators in human cells. Using computational methods, we identified putative human tDNA insulators. Using silencer blocking, transgene protection and repressor blocking assays we show that some of these tDNA-containing fragments can function as barrier insulators in human cells. We find that these elements also have the ability to block enhancers from activating RNA pol II transcribed promoters. Characterization of a putative tDNA insulator in human cells reveals that the site possesses chromatin signatures similar to those observed at other better-characterized eukaryotic insulators. Enhanced 4C analysis demonstrates that the tDNA insulator makes long-range chromatin contacts with other tDNAs and ETC sites but not with intervening or flanking RNA pol II transcribed genes.


Assuntos
Elementos Isolantes/genética , RNA de Transferência/genética , Animais , Linhagem Celular , Cromatina/genética , Cromossomos Humanos Par 17/genética , Biologia Computacional/métodos , DNA Fúngico/genética , DNA Fúngico/metabolismo , Elementos Facilitadores Genéticos/genética , Inativação Gênica , Humanos , Histona Desmetilases com o Domínio Jumonji/genética , Mamíferos/genética , Ligação Proteica , RNA Polimerase III/metabolismo , Schizosaccharomyces/genética , Alinhamento de Sequência , Sintenia , Fatores de Transcrição TFIII/metabolismo , Transcrição Gênica/genética , Transgenes
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