RESUMO
During embryogenesis, the genome shifts from transcriptionally quiescent to extensively active in a process known as Zygotic Genome Activation (ZGA). In Drosophila, the pioneer factor Zelda is known to be essential for the progression of development; still, it regulates the activation of only a small subset of genes at ZGA. However, thousands of genes do not require Zelda, suggesting that other mechanisms exist. By conducting GRO-seq, HiC and ChIP-seq in Drosophila embryos, we demonstrate that up to 65% of zygotically activated genes are enriched for the histone variant H2A.Z. H2A.Z enrichment precedes ZGA and RNA Polymerase II loading onto chromatin. In vivo knockdown of maternally contributed Domino, a histone chaperone and ATPase, reduces H2A.Z deposition at transcription start sites, causes global downregulation of housekeeping genes at ZGA, and compromises the establishment of the 3D chromatin structure. We infer that H2A.Z is essential for the de novo establishment of transcriptional programs during ZGA via chromatin reorganization.
Assuntos
Desenvolvimento Embrionário/genética , Desenvolvimento Embrionário/fisiologia , Genoma , Histonas/classificação , Histonas/genética , Histonas/metabolismo , Zigoto/metabolismo , Adenosina Trifosfatases , Animais , Divisão Celular , Cromatina , Sequenciamento de Cromatina por Imunoprecipitação , Regulação para Baixo , Drosophila , Epigenômica , Feminino , Regulação da Expressão Gênica no Desenvolvimento , Técnicas de Silenciamento de Genes , Genes Essenciais , Chaperonas de Histonas , Masculino , RNA Polimerase II , Sítio de Iniciação de Transcrição , Ativação TranscricionalRESUMO
Canonical histones (H2A, H2B, H3, and H4) are present in all eukaryotes where they package genomic DNA and participate in numerous cellular processes, such as transcription regulation and DNA repair. In addition to the canonical histones, there are many histone variants, which have different amino acid sequences, possess tissue-specific expression profiles, and function distinctly from the canonical counterparts. A number of histone variants, including both core histones (H2A/H2B/H3/H4) and linker histones (H1/H5), have been identified to date. Htz1 (H2A.Z) and CENP-A (CenH3) are present from yeasts to mammals, and H3.3 is present from Tetrahymena to humans. In addition to the prevalent variants, others like H3.4 (H3t), H2A.Bbd, and TH2B, as well as several H1 variants, are found to be specific to mammals. Among them, H2BFWT, H3.5, H3.X, H3.Y, and H4G are unique to primates (or Hominidae). In this review, we focus on localization and function of primate- or hominidae-specific histone variants.
Assuntos
Histonas/classificação , Primatas/genética , Primatas/fisiologia , Sequência de Aminoácidos , Animais , Encéfalo , Neoplasias da Mama , Nucléolo Celular , DNA , Regulação da Expressão Gênica , Histonas/genética , Humanos , Mamíferos , FilogeniaRESUMO
Nucleosomes consisting of a short piece of deoxyribonucleic acid (DNA) wrapped around an octamer of histone proteins form the fundamental unit of chromatin in eukaryotes. Their role in DNA compaction comes with regulatory functions that impact essential genomic processes such as replication, transcription, and repair. The assembly of nucleosomes obeys a precise pathway in which tetramers of histones H3 and H4 bind to the DNA first to form tetrasomes, and two dimers of histones H2A and H2B are subsequently incorporated to complete the complex. As viable intermediates, we previously showed that tetrasomes can spontaneously flip between a left-handed and right-handed conformation of DNA-wrapping. To pinpoint the underlying mechanism, here we investigated the role of the H3-H3 interface for tetramer flexibility in the flipping process at the single-molecule level. Using freely orbiting magnetic tweezers, we studied the assembly and structural dynamics of individual tetrasomes modified at the cysteines close to this interaction interface by iodoacetamide (IA) in real time. While such modification did not affect the structural properties of the tetrasomes, it caused a 3-fold change in their flipping kinetics. The results indicate that the IA-modification enhances the conformational plasticity of tetrasomes. Our findings suggest that subnucleosomal dynamics may be employed by chromatin as an intrinsic and adjustable mechanism to regulate DNA supercoiling.
Assuntos
Histonas/química , Histonas/classificação , Iodoacetamida/química , Conformação MolecularRESUMO
Post-translational modifications of histones play a key role in the regulation of gene expression during development and differentiation. Numerous studies have shown the dynamics of combinatorial regulation by transcription factors and histone modifications, in the sense that different combinations lead to distinct expression outcomes. Here, we investigated gene regulation by stable enrichment patterns of histone marks H3K4me2 and H3K4me3 in combination with the chromatin binding of the muscle tissue-specific transcription factor MyoD during myogenic differentiation of C2C12 cells. Using k-means clustering, we found that specific combinations of H3K4me2/3 profiles over and towards the gene body impact on gene expression and marks a subset of genes important for muscle development and differentiation. By further analysis, we found that the muscle key regulator MyoD was significantly enriched on this subset of genes and played a repressive role during myogenic differentiation. Among these genes, we identified the pluripotency gene Patz1, which is repressed during myogenic differentiation through direct binding of MyoD to promoter elements. These results point to the importance of integrating histone modifications and MyoD chromatin binding for coordinated gene activation and repression during myogenic differentiation.
Assuntos
Diferenciação Celular/genética , Histonas/genética , Proteína MyoD/genética , Mioblastos/metabolismo , Animais , Linhagem Celular , Células Cultivadas , Análise por Conglomerados , Fibroblastos/citologia , Fibroblastos/metabolismo , Perfilação da Expressão Gênica/métodos , Ontologia Genética , Células HEK293 , Histonas/classificação , Histonas/metabolismo , Humanos , Lisina/metabolismo , Metilação , Camundongos , Desenvolvimento Muscular/genética , Proteína MyoD/metabolismo , Mioblastos/citologia , Proteínas de Neoplasias/genética , Proteínas de Neoplasias/metabolismo , Ligação Proteica , Proteínas Repressoras/genética , Proteínas Repressoras/metabolismo , Reação em Cadeia da Polimerase Via Transcriptase ReversaRESUMO
Histone H1 is a structural component of chromatin that may have a role in the regulation of chromatin dynamics. Unlike core histones, the linker histone H1 family is evolutionarily diverse and many organisms have multiple H1 variants or subtypes, distinguishable between germ-line and somatic cells. In mammals, the H1 family includes seven somatic H1 variants with a prevalence that varies between cell types and over the course of differentiation, H1.1 to H1.5 being expressed in a replication-dependent manner, whereas H1.0 and H1X are replication-independent. Until recently, it has not been known whether the different variants had specific roles in the regulation of nuclear processes or were differentially distributed across the genome. To address this, an increasing effort has been made to investigate divergent features among H1 variants, regarding their structure, expression patterns, chromatin dynamics, post-translational modifications and genome-wide distribution. Although H1 subtypes seem to have redundant functions, several reports point to the idea that they are also differently involved in specific cellular processes. Initial studies investigating the genomic distribution of H1 variants have started to suggest that despite a wide overlap, different variants may be enriched or preferentially located at different chromatin types, but this may depend on the cell type, the relative abundance of the variants, the differentiation state of the cell, or whether cells are derived from a neoplastic process. Understanding the heterogeneity of the histone H1 family is crucial to elucidate their role in chromatin organization, gene expression regulation and other cellular processes.
Assuntos
Histonas/classificação , Animais , Cromatina/química , Regulação da Expressão Gênica , Histonas/genética , Histonas/fisiologia , Humanos , Sinais de Localização NuclearRESUMO
KEY MESSAGE: Overexpression of CsHis in tobacco promoted chromatin condensation, but did not affect the phenotype. It also conferred tolerance to low-temperature, high-salinity, ABA, drought and oxidative stress in transgenic tobacco. H1 histone, as a major structural protein of higher-order chromatin, is associated with stress responses in plants. Here, we describe the functions of the Camellia sinensis H1 Histone gene (CsHis) to illustrate its roles in plant responses to stresses. Subcellular localization and prokaryotic expression assays showed that the CsHis protein is localized in the nucleus, and its molecular size is approximately 22.5 kD. The expression levels of CsHis in C. sinensis leaves under various conditions were investigated by qRT-PCR, and the results indicated that CsHis was strongly induced by various abiotic stresses such as low-temperature, high-salinity, ABA, drought and oxidative stress. Overexpression of CsHis in tobacco (Nicotiana tabacum) promoted chromatin condensation, while there were almost no changes in the growth and development of transgenic tobacco plants. Phylogenetic analysis showed that CsHis belongs to the H1C and H1D variants of H1 histones, which are stress-induced variants and not the key variants required for growth and development. Stress tolerance analysis indicated that the transgenic tobacco plants exhibited higher tolerance than the WT plants upon exposure to various abiotic stresses; the transgenic plants displayed reduced wilting and senescence and exhibited greater net photosynthetic rate (Pn), stomatal conductance (Gs) and maximal photochemical efficiency (Fv/Fm) values. All the above results suggest that CsHis is a stress-induced gene and that its overexpression improves the tolerance to various abiotic stresses in the transgenic tobacco plants, possibly through the maintenance of photosynthetic efficiency.
Assuntos
Adaptação Fisiológica/genética , Camellia sinensis/genética , Histonas/genética , Nicotiana/genética , Proteínas de Plantas/genética , Ácido Abscísico/farmacologia , Western Blotting , Camellia sinensis/metabolismo , Núcleo Celular/metabolismo , Cromatina/genética , Cromatina/metabolismo , Temperatura Baixa , Secas , Regulação da Expressão Gênica de Plantas/efeitos dos fármacos , Histonas/classificação , Histonas/metabolismo , Microscopia Eletrônica de Transmissão , Microscopia de Fluorescência , Fotossíntese/genética , Filogenia , Reguladores de Crescimento de Plantas/farmacologia , Folhas de Planta/genética , Folhas de Planta/metabolismo , Folhas de Planta/ultraestrutura , Proteínas de Plantas/metabolismo , Plantas Geneticamente Modificadas , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Cloreto de Sódio/farmacologia , Estresse Fisiológico , Nicotiana/metabolismo , Nicotiana/ultraestruturaRESUMO
Chromatin acts as an organizer and indexer of genomic DNA and is a highly dynamic and regulated structure with properties directly related to its constituent parts. Histone variants are abundant components of chromatin that replace canonical histones in a subset of nucleosomes, thereby altering nucleosomal characteristics. The present review focuses on the H2A variant histones, summarizing current knowledge of how H2A variants can introduce chemical and functional heterogeneity into chromatin, the positions that nucleosomes containing H2A variants occupy in eukaryotic genomes, and the regulation of these localization patterns.
Assuntos
Variação Genética , Histonas/genética , Trifosfato de Adenosina/metabolismo , Sequência de Aminoácidos , Animais , Cromatina/genética , Cromatina/metabolismo , Montagem e Desmontagem da Cromatina , Feminino , Genoma , Genoma Humano , Histonas/química , Histonas/classificação , Histonas/metabolismo , Humanos , Masculino , Dados de Sequência Molecular , Família Multigênica , Nucleossomos/genética , Nucleossomos/metabolismo , Processamento de Proteína Pós-Traducional , Homologia de Sequência de Aminoácidos , Especificidade da EspécieRESUMO
Synthesized allopolyploids are valuable materials for comparative analyses of two or more distinct genomes, such as the expression changes (activation, inactivation or differential expression) of orthologous genes following allopolyploidization. CENH3 is a centromere- specific histone H3 variant and has been regarded as a central component in kinetochore formation and centromere function. In this study, interspecific hybrids of Oryza genus (AA × CC, AA × CCDD) and their backcross progenies were produced, and the genome constitutions were identified as AC, ACC, ACD, AACD, or AA(CD) by Genomic in situ hybridization (GISH). We further cloned and sequenced the CENH3 genes from O. sativa (AA), O. officinalis (CC) and O. latifolia (CCDD). Sequencing of RT-PCR products revealed that CENH3_C2 and CENH3_D, the two CENH3 alleles from O. latifolia, showed polymorphism in several sites, while CENH3_C2 and CENH3_C1 from O. officinalis were different at only two amino acids positions. Moreover, we found that the CENH3 genes from both parents are expressed in interspecific hybrids and their progenies. Specifically, based on our cDNA sequencing data, the ratio of expression level between CENH3_A and CENH3_C1 was approximately 1 in AC and 0.5 in ACC genomes, respectively. As a result, the CENH3 expression patterns shed more light on the inter-coordination between varied centromeric DNA sequences and highly conserved kinetochore protein in synthesized allopolyploids of Oryza genus.
Assuntos
Histonas/genética , Proteínas de Plantas/genética , Poaceae/genética , Poliploidia , Alelos , Sequência de Aminoácidos , Sequência de Bases , Centrômero/genética , Cruzamentos Genéticos , DNA Complementar/química , DNA Complementar/genética , Perfilação da Expressão Gênica , Regulação da Expressão Gênica de Plantas , Genoma de Planta/genética , Histonas/classificação , Hibridização Genética , Hibridização in Situ Fluorescente , Dados de Sequência Molecular , Oryza/genética , Filogenia , Proteínas de Plantas/classificação , Poaceae/classificação , Isoformas de Proteínas/genética , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Análise de Sequência de DNA , Homologia de Sequência de Aminoácidos , Homologia de Sequência do Ácido Nucleico , Especificidade da EspécieRESUMO
Chromatin-modifying factors have essential roles in DNA processing pathways that dictate cellular functions. The ability of chromatin modifiers, including the INO80 and SWR1 chromatin-remodelling complexes, to regulate transcriptional processes is well established. However, recent studies reveal that the INO80 and SWR1 complexes have crucial functions in many other essential processes, including DNA repair, checkpoint regulation, DNA replication, telomere maintenance and chromosome segregation. During these diverse nuclear processes, the INO80 and SWR1 complexes function cooperatively with their histone substrates, gamma-H2AX and H2AZ. This research reveals that INO80 and SWR1 ATP-dependent chromatin remodelling is an integral component of pathways that maintain genomic integrity.
Assuntos
Adenosina Trifosfatases/metabolismo , Montagem e Desmontagem da Cromatina , Proteínas de Saccharomyces cerevisiae/metabolismo , Sequência de Aminoácidos , Animais , Cromatina/metabolismo , Cromossomos Fúngicos/metabolismo , Cromossomos Fúngicos/ultraestrutura , Quebras de DNA de Cadeia Dupla , Reparo do DNA , Replicação do DNA , Histonas/classificação , Histonas/genética , Histonas/metabolismo , Humanos , Modelos Biológicos , Dados de Sequência Molecular , Filogenia , Saccharomyces cerevisiae/genética , Saccharomyces cerevisiae/metabolismo , Alinhamento de Sequência , Transcrição GênicaRESUMO
Although several histone variants have been studied in both animal and plant organisms, little is known about their distribution during processes that involve alterations in chromatin function, such as differentiation, dedifferentiation and hormone treatment. In this study we evaluated the ratio of each histone variant in each of the four core histone classes in the three developmental zones of maize (Zea mays L.) root and in callus cultures derived from them, in order to define possible alterations either during plant cell differentiation or dedifferentiation. We also evaluated core histone variant ratios in the developmental zones of roots treated with auxin and gibberellin in order to examine the effects of exogenously applied plant hormones to histone variant distribution. Finally, immunohistochemical detection was used to identify the root tissues containing modified forms of core histones and correlates them with the physiological status of the plant cells. According to the results presented in this study, histone variant ratios are altered in all the cases examined, i.e. in the developmental zones of maize root, in callus cultures derived from them and in the developmental zones of roots treated either with auxin or gibberellin. We propose that the alterations in linker histone variant ratios are correlated with plant cell differentiation and physiological status in each case.
Assuntos
Histonas/metabolismo , Reguladores de Crescimento de Plantas/farmacologia , Raízes de Plantas/embriologia , Zea mays/embriologia , Western Blotting , Diferenciação Celular , Densitometria , Giberelinas , Histonas/classificação , Imuno-Histoquímica , Ácidos Indolacéticos , Reguladores de Crescimento de Plantas/isolamento & purificação , Raízes de Plantas/química , Raízes de Plantas/efeitos dos fármacos , Zea mays/química , Zea mays/efeitos dos fármacosRESUMO
Phosphorylation of histone H2AX on Ser 139 (gammaH2AX) is one of the earliest events in the response to DNA double-strand breaks; however, the subsequent removal of gammaH2AX from chromatin is less understood, despite being a process tightly coordinated with DNA repair. Previous studies in yeast have identified the Pph3 phosphatase (the PP4C orthologue) as important for the dephosphorylation of gammaH2AX. By contrast, work in human cells attributed this activity to PP2A. Here, we report that PP4 contributes to the dephosphorylation of gammaH2AX, both at the sites of DNA damage and in undamaged chromatin in human cells, independently of a role in DNA repair. Furthermore, depletion of PP4C results in a prolonged checkpoint arrest, most likely owing to the persistence of mediator of DNA damage checkpoint 1 (MDC1) at the sites of DNA lesions. Taken together, these results indicate that PP4 is an evolutionarily conserved gammaH2AX phosphatase.
Assuntos
Dano ao DNA/fisiologia , Reparo do DNA/fisiologia , Histonas/metabolismo , Fosfoproteínas Fosfatases/fisiologia , Linhagem Celular Tumoral , Cromatina/metabolismo , Sequência Conservada , Evolução Molecular , Fase G2/genética , Histonas/classificação , Humanos , Mitose/genética , Fosfoproteínas Fosfatases/isolamento & purificação , Fosforilação/fisiologia , Fatores de TempoRESUMO
Although several linker histone variants have been studied in both animal and plant organisms, little is known about their distribution during processes that involve alterations in chromatin function, such as differentiation, dedifferentiation and hormone treatment. In this study, we identified linker histone variants by using specific anti-histone Hl antibodies. Each variant's ratio to total Hl in the three developmental zones of maize (Zea mays L.) root and in callus cultures derived from them was estimated in order to define possible alterations either during plant cell differentiation or during their dedifferentiation. We also evaluated linker histone variants' ratios in the developmental zones of maize roots treated with auxin in order to examine the effects of exogenous applied auxin to linker histone variant distribution. Finally, immunohistochemical detection was used to identify the root tissues containing each variant and correlate them with the physiological status of the plant cells. According to the results presented in this study, linker histone variants' ratios are altered in the developmental zones of maize root, while they are similar to the meristematic zone in samples from callus cultures and to the differentiation zone in samples from roots treated with auxin. We propose that the alterations in linker histone variants' ratios are correlated with plant cell differentiation and dedifferentiation.
Assuntos
Histonas/análise , Ácidos Indolacéticos/farmacologia , Reguladores de Crescimento de Plantas/farmacologia , Raízes de Plantas/química , Zea mays/química , Desdiferenciação Celular/efeitos dos fármacos , Diferenciação Celular/efeitos dos fármacos , Histonas/classificação , Imuno-Histoquímica , Raízes de Plantas/citologia , Raízes de Plantas/efeitos dos fármacos , Técnicas de Cultura de Tecidos , Zea mays/citologia , Zea mays/efeitos dos fármacosRESUMO
Given that cocaine induces neuroadaptations through regulation of gene expression, we investigated whether chromatin remodeling at specific gene promoters may be a key mechanism. We show that cocaine induces specific histone modifications at different gene promoters in striatum, a major neural substrate for cocaine's behavioral effects. At the cFos promoter, H4 hyperacetylation is seen within 30 min of a single cocaine injection, whereas no histone modifications were seen with chronic cocaine, consistent with cocaine's ability to induce cFos acutely, but not chronically. In contrast, at the BDNF and Cdk5 promoters, genes that are induced by chronic, but not acute, cocaine, H3 hyperacetylation was observed with chronic cocaine only. DeltaFosB, a cocaine-induced transcription factor, appears to mediate this regulation of the Cdk5 gene. Furthermore, modulating histone deacetylase activity alters locomotor and rewarding responses to cocaine. Thus, chromatin remodeling is an important regulatory mechanism underlying cocaine-induced neural and behavioral plasticity.
Assuntos
Montagem e Desmontagem da Cromatina/fisiologia , Cocaína/administração & dosagem , Corpo Estriado/efeitos dos fármacos , Inibidores da Captação de Dopamina/administração & dosagem , Plasticidade Neuronal/efeitos dos fármacos , Acetilação , Animais , Comportamento Animal/efeitos dos fármacos , Fator Neurotrófico Derivado do Encéfalo/genética , Fator Neurotrófico Derivado do Encéfalo/metabolismo , Butiratos/farmacologia , Montagem e Desmontagem da Cromatina/efeitos dos fármacos , Condicionamento Operante/efeitos dos fármacos , Corpo Estriado/fisiologia , Quinase 5 Dependente de Ciclina/genética , Quinase 5 Dependente de Ciclina/metabolismo , Esquema de Medicação , Interações Medicamentosas , Inibidores Enzimáticos/farmacologia , Regulação da Expressão Gênica/efeitos dos fármacos , Técnicas de Transferência de Genes/psicologia , Histona Desacetilases/metabolismo , Histonas/classificação , Histonas/metabolismo , Imuno-Histoquímica/métodos , Imunoprecipitação/métodos , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Transgênicos , Atividade Motora/efeitos dos fármacos , Células PC12/metabolismo , Regiões Promotoras Genéticas/fisiologia , Subunidades Proteicas , Proteínas Proto-Oncogênicas c-fos/genética , RNA Mensageiro/biossíntese , Ratos , Ratos Sprague-Dawley , Reação em Cadeia da Polimerase Via Transcriptase Reversa/métodos , Fatores de TempoRESUMO
In humans, eight types of histone H1 exist (H1.1-H1.5, H1 degrees , H1t and H1oo), all consisting of a highly conserved globular domain and less conserved N- and C-terminal tails. Although the precise functions of these isoforms are not yet understood, and H1 subtypes have been found to be dispensable for mammalian development, it is now clear that specific functions may be assigned to certain individual H1 subtypes. Moreover, microsequence variations within the isoforms, such as polymorphisms or mutations, may have biological significance because of the high degree of sequence conservation of these proteins. This study used a hydrophilic interaction liquid chromatographic method to detect sequence variants within the subtypes. Two deviations from wild-type H1 sequences were found. In K562 erythroleukemic cells, alanine at position 17 in H1.2 was replaced by valine, and, in Raji B lymphoblastoid cells, lysine at position 173 in H1.4 was replaced by arginine. We confirmed these findings by DNA sequencing of the corresponding gene segments. In K562 cells, a homozygous GCC-->GTC shift was found at codon 18, giving rise to H1.2 Ala17Val because the initial methionine is removed in H1 histones. Raji cells showed a heterozygous AAA-->AGA codon change at position 174 in H1.4, corresponding to the Lys173Arg substitution. The allele frequency of these sequence variants in a normal Swedish population was found to be 6.8% for the H1.2 GCC-->GTC shift, indicating that this is a relatively frequent polymorphism. The AAA-->AGA codon change in H1.4 was detected only in Raji cells and was not present in a normal population or in six other cell lines derived from individuals suffering from Burkitt's lymphoma. The significance of these sequence variants is unclear, but increasing evidence indicates that minor sequence variations in linker histones may change their binding characteristics, influence chromatin remodeling, and specifically affect important cellular functions.
Assuntos
Histonas/classificação , Histonas/genética , Polimorfismo Genético/genética , Sequência de Bases , Linhagem Celular , Cromatografia Líquida de Alta Pressão , Humanos , Interações Hidrofóbicas e HidrofílicasRESUMO
To investigate whether the increased rate of lymphocyte apoptosis in systemic lupus erythematosus is involved in the onset of the disease, apoptotic or necrotic T or B lymphocytes from various cell lines were injected intraperitoneally into pre-autoimmune (NZBxNZW)F1 mice (BW) and non-autoimmune BALB/c mice. The intraperitoneal production of cytokines and chemokines, the specific T cell response in the spleen, and the production of anti-histone and anti-dsDNA Ab were investigated. The onset of the disease was characterized by creatinine levels and evaluation of glomerular IgG deposits. In BW, but not in BALB/c mice, injection of apoptotic and not necrotic cells up-regulated IL-6 and IL-10 in resident macrophages. Administration of apoptotic cells augmented the number of Th2 and B lymphocytes recruited in the peritoneal cavity. Only the treatment with apoptotic B cells promoted a systemic Th2 autoimmune response to H2 histones, associated with earlier occurrence of high levels of anti-dsDNA autoantibodies, higher creatinine levels and more numerous glomerular IgG deposits than in BW controls not injected with apoptotic B cells. In genetically susceptible mice exposure to apoptotic of B, but not T, lymphocytes can elicit a Th2 response to H2 histones that helps B cell production of anti-dsDNA Ab and finally triggers the onset of lupus.
Assuntos
Apoptose , Autoantígenos/imunologia , Doenças Autoimunes/imunologia , Linfócitos B/patologia , Histonas/imunologia , Lúpus Eritematoso Sistêmico/imunologia , Animais , Anticorpos Antinucleares/análise , Anticorpos Antinucleares/imunologia , Autoanticorpos/análise , Autoanticorpos/imunologia , Doenças Autoimunes/sangue , Doenças Autoimunes/patologia , Quimiocinas/biossíntese , Creatinina/sangue , DNA/imunologia , Modelos Animais de Doenças , Progressão da Doença , Epitopos de Linfócito T/imunologia , Feminino , Histonas/classificação , Imunização , Imunoglobulina G/análise , Imunoglobulina G/imunologia , Interferon gama/biossíntese , Interferon gama/sangue , Interleucina-4/biossíntese , Interleucina-4/sangue , Glomérulos Renais/química , Glomérulos Renais/imunologia , Glomérulos Renais/patologia , Leucemia de Células T/imunologia , Leucemia de Células T/patologia , Lúpus Eritematoso Sistêmico/sangue , Lúpus Eritematoso Sistêmico/patologia , Nefrite Lúpica/sangue , Nefrite Lúpica/imunologia , Nefrite Lúpica/patologia , Cooperação Linfocítica , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Endogâmicos NZB , Fagocitose , Plasmocitoma/imunologia , Plasmocitoma/patologia , Baço/imunologia , Subpopulações de Linfócitos T/imunologia , Células Th2/imunologia , Células Tumorais CultivadasRESUMO
For nearly twenty years researchers have observed changes in the histone H1 subtype content of tissues as an organism develops into an adult. To better understand the consequences of such changes, immunofractionation of chromatin using previously characterized antibodies specific for human H1 subtypes was employed in the analysis of a fibroblast cell strain derived from a 37-year-old individual. DNAs isolated from immunoprecipitates were probed for the existence of a variety of DNA sequences. The results presented lend further support to a previously-proposed model (Parseghian et al. [2000] Chromosome Res 8:405-424) in which transcription of a sequence is accompanied by the selective depletion of subtypes. The data also suggest that there is more total H1 on actively transcribed sequences in these cells as compared to fetal fibroblasts and that there is less difference in the subtype compositions of active genes vs. inactive sequences in this strain. Specifically, the consequences of these changes appear to correlate with the attenuation of the heat shock response in aging fibroblasts. In a broader context, these results could explain why there are reductions in transcription in cells from mature tissue that approach senescence.
Assuntos
Senescência Celular/genética , Cromatina/metabolismo , Fibroblastos/fisiologia , Histonas/classificação , Histonas/metabolismo , Proteínas Tirosina Quinases , Adulto , Análise de Variância , Células Cultivadas , Centrômero/genética , Cromatina/genética , DNA Satélite/metabolismo , Proteínas de Ligação a DNA/genética , Proteínas de Ligação a DNA/metabolismo , Feto/citologia , Feto/fisiologia , Fibroblastos/metabolismo , Regulação da Expressão Gênica , Inativação Gênica , Genes , Proteínas de Choque Térmico HSP90/biossíntese , Proteínas de Choque Térmico HSP90/genética , Proteínas de Choque Térmico HSP90/metabolismo , Histonas/genética , Humanos , Regiões Constantes de Imunoglobulina/genética , Regiões Constantes de Imunoglobulina/metabolismo , Cadeias kappa de Imunoglobulina/genética , Cadeias kappa de Imunoglobulina/metabolismo , Fator 1 de Ligação ao Facilitador Linfoide , Masculino , Proteínas do Tecido Nervoso/genética , Proteínas do Tecido Nervoso/metabolismo , Proteínas Proto-Oncogênicas c-myc/genética , Proteínas Proto-Oncogênicas c-myc/metabolismo , Pseudogenes , Receptor Tipo 3 de Fator de Crescimento de Fibroblastos , Receptores de Fatores de Crescimento de Fibroblastos/genética , Receptores de Fatores de Crescimento de Fibroblastos/metabolismo , Sequências Repetitivas de Ácido Nucleico , Telômero/genética , Fatores de Transcrição/genética , Fatores de Transcrição/metabolismoRESUMO
H1 histones are involved in the formation of higher order chromatin structures and in the modulation of gene expression. Changes in chromatin structure are a characteristic initial feature of apoptosis. We therefore have investigated the histone H1 pattern of the human leukemic cell line HL60 undergoing programmed cell death, as induced by topoisomerase I inhibition. Histone H1 proteins were isolated and analyzed by high performance liquid chromatography and capillary zone electrophoresis. DNA fragmentation after apoptosis induction was monitored by agarose gel electrophoresis. The patterns of the three H1 histone subtypes extractable from apoptotic HL60 cells significantly differed from those of control cells in showing a decrease of phosphorylated H1 subtypes and an increase of the respective dephosphorylated forms. This dephosphorylation of H1 histones could be observed already 45 min after apoptosis induction and preceded internucleosomal DNA cleavage by approximately 2 h. We conclude that during apoptotic DNA fragmentation, the H1 histones become rapidly dephosphorylated by a yet unknown protein phosphatase.
Assuntos
Apoptose , Histonas/metabolismo , Fragmentação do DNA , DNA de Neoplasias/metabolismo , Desoxirribonucleases/metabolismo , Ativação Enzimática , Células HL-60 , Histonas/classificação , Humanos , Toxinas Marinhas , Nucleossomos/metabolismo , Oxazóis/farmacologia , Fosfoproteínas Fosfatases/antagonistas & inibidores , Fosforilação , Proteína Quinase C/antagonistas & inibidores , Proteína Quinase C/metabolismo , Estaurosporina/farmacologia , Acetato de Tetradecanoilforbol/farmacologia , Topotecan/farmacologiaRESUMO
Using a poly(ADP-ribose) binding assay on protein blots we examined the ability of rat testis histone H1 variants to establish non-covalent interactions with the polymer. All the H1 variants bound ADP-ribose polymers; the binding was salt resistant and highly specific, occurring even in the presence of a large excess of competitor DNA. A comparison among the H1 variants showed that H1t has the highest affinity for poly(ADP-ribose). Long and branched poly(ADP-ribose) molecules were found to be preferentially involved in the interaction with the histone variants. The results further corroborate the concept that non-covalent interactions of poly(ADP-ribose) with target proteins may constitute an important mechanism to modulate chromatin structure.
Assuntos
Histonas/química , Poli Adenosina Difosfato Ribose/química , DNA/química , Eletroforese em Gel de Poliacrilamida , Endopeptidase K , Histonas/classificação , Poli Adenosina Difosfato Ribose/síntese químicaRESUMO
MacroH2A is a novel nucleosomal core histone that contains a large nonhistone region and a region that closely resembles a full length histone H2A. We have cloned a cDNA that contains the entire coding region of macroH2A1.2, one of the two identified subtypes of macroH2A1. MacroH2A1.2 was found to differ from the other known subtype, macroH2A1.1, in a single segment of the nonhistone region. MacroH2A1 specific antibodies revealed relatively high levels of both subtypes in adult liver and kidney. MacroH2A1.1 was much lower in fetal liver and kidney in comparison to their adult counterparts, and was not detected in adult thymus and testis, tissues with active cell division and differentiation. Both subtypes were present at very low levels or absent from mouse embryonic stem cells maintained in an undifferentiated state by growth in the presence of leukemia inhibitory factor. MacroH2A1.2 increased when the embryonic stem cells were induced to differentiate in vitro, while macroH2A1.1 remained undetectable. These results support the idea that macroH2A1.1 and macroH2A1.2 are functionally distinct, and suggest that changes in their expression may play a role in developmentally regulated changes in chromatin structure and function.
Assuntos
Histonas/metabolismo , Animais , Clonagem Molecular , Regulação da Expressão Gênica no Desenvolvimento , Histonas/classificação , Camundongos , Nucleossomos/ultraestrutura , Coelhos , Ratos , Distribuição TecidualRESUMO
The H1 class of histones comprises several main-type, S-phase dependent isoforms and, in addition, a sperm-specific H1t and a peculiar subtype, H1 zero, which is confined to highly differentiated cells. In contrast to main type histone genes, the H1 zero gene expression does not strictly depend on DNA replication. Also in contrast to the other H1 subtype genes, the mammalian H1 zero gene is not included in a histone gene cluster and its mRNA differs in structure, size and mode of processing from other histone mRNAs. The regulation of expression of the H1 zero gene varies from main type H1 genes in several respects. This is manifested in the promoter structure which contains sequence elements that are also found in main type H1 promoters, but also shows regulatory motifs which appear to be involved in a developmental regulation of the H1 zero gene, such as a retinoic acid receptor binding site, which has been described in the mammalian H1 zero gene promoter.