RESUMO
The heat shock factor 1 (HSF1)-dependent transcriptional activation of human pericentric heterochromatin in heat-shocked cells is the most striking example of transcriptional activation of heterochromatin. Until now, pericentric heterochromatin of chromosome 9 has been identified as the primary target of HSF1, in both normal and tumor heat-shocked cells. Transcriptional awakening of this large genomic region results in the nuclear accumulation of satellite III (SATIII) noncoding RNAs (ncRNAs) and the formation in cis of specific structures known as nuclear stress bodies (nSBs). Here, we show that, in four different male cell lines, including primary human fibroblasts and amniocytes, pericentric heterochromatin of chromosome Y can also serve as a unique primary site of HSF1-dependent heterochromatin transcriptional activation, production of SATIII ncRNA, and nucleation of nuclear stress bodies (nSBs) upon heat shock. Our observation suggests that the chromosomal origin of SATIII transcripts in cells submitted to heat shock is not a determinant factor as such, but that transcription of SATIII repetitive units or the SATIII ncRNA molecules is the critical element of HSF1-dependent transcription activation of constitutive heterochromatin.
Assuntos
Cromossomos Humanos Y/genética , DNA Satélite/genética , Fibroblastos/metabolismo , Fatores de Transcrição de Choque Térmico/metabolismo , Heterocromatina/genética , Fatores de Processamento de Serina-Arginina/metabolismo , Estresse Fisiológico , Feminino , Fibroblastos/citologia , Fatores de Transcrição de Choque Térmico/genética , Resposta ao Choque Térmico , Humanos , Masculino , Fatores de Processamento de Serina-Arginina/genética , Transcrição GênicaRESUMO
In response to metabolic or environmental stress, cells activate powerful defense mechanisms to prevent the formation and accumulation of toxic protein aggregates. The main orchestrator of this cellular response is HSF1 (heat shock factor 1), a transcription factor involved in the up-regulation of protein-coding genes with protective roles. It has become very clear that HSF1 has a broader function than initially expected. Indeed, our previous work demonstrated that, upon stress, HSF1 activates the transcription of a non-coding RNA, named Satellite III, at pericentromeric heterochromatin. Here, we observe that the function of HSF1 extends to telomeres and identify subtelomeric DNA as a new genomic target of HSF1. We show that the binding of HSF1 to subtelomeric regions plays an essential role in the upregulation of non-coding TElomeric Repeat containing RNA (TERRA) transcription upon heat shock. Importantly, our data show that telomere integrity is impacted by heat shock and that telomeric DNA damages are markedly enhanced in HSF1 deficient cells. Altogether, our findings reveal a new direct and essential function of HSF1 in the transcriptional activation of TERRA and in telomere protection upon stress.
Assuntos
Fatores de Transcrição de Choque Térmico/fisiologia , RNA Longo não Codificante/genética , RNA Longo não Codificante/metabolismo , Telômero/metabolismo , Células HeLa , Resposta ao Choque Térmico , Humanos , Estabilidade de RNA , Homeostase do Telômero , Transcrição Gênica , Ativação TranscricionalRESUMO
Heat shock triggers a transient and ubiquitous response, the function of which is to protect cells against stress-induced damage. The heat-shock response is controlled by a key transcription factor known as heat shock factor 1 (HSF1). We have developed a multiconfocal fluorescence correlation spectroscopy setup to measure the dynamics of HSF1 during the course of the heat-shock response. The system combines a spatial light modulator, to address several points of interest, and an electron-multiplying charge-coupled camera for fast multiconfocal recording of the photon streams. Autocorrelation curves with a temporal resolution of 14 µs were analyzed before and after heat shock on eGFP and HSF1-eGFP-expressing cells. Evaluation of the dynamic parameters of a diffusion-and-binding model showed a slower HSF1 diffusion after heat shock. It is also observed that the dissociation rate decreases after heat shock, whereas the association rate is not affected. In addition, thanks to the multiconfocal fluorescence correlation spectroscopy system, up to five spots could be simultaneously located in each cell nucleus. This made it possible to quantify the intracellular variability of the diffusion constant of HSF1, which is higher than that of inert eGFP molecules and increases after heat shock. This finding is consistent with the fact that heat-shock response is associated with an increase of HSF1 interactions with DNA and cannot be explained even partially by heat-induced modifications of nuclear organization.
Assuntos
Resposta ao Choque Térmico , Espectrometria de Fluorescência/métodos , Sítios de Ligação , Calibragem , Linhagem Celular Tumoral , Sobrevivência Celular , Corantes Fluorescentes/metabolismo , Proteínas de Fluorescência Verde/metabolismo , Humanos , Cinética , FotodegradaçãoRESUMO
In eukaryotes, the heat shock response is orchestrated by a transcription factor named Heat Shock Factor 1 (HSF1). HSF1 is mostly characterized for its role in activating the expression of a repertoire of protein-coding genes, including the heat shock protein (HSP) genes. Remarkably, a growing set of reports indicate that, upon heat shock, HSF1 also targets various non-coding regions of the genome. Focusing primarily on mammals, this review aims at reporting the identity of the non-coding genomic sites directly bound by HSF1, and at describing the molecular function of the long non-coding RNAs (lncRNAs) produced in response to HSF1 binding. The described non-coding genomic targets of HSF1 are pericentric Satellite DNA repeats, (sub)telomeric DNA repeats, Short Interspersed Nuclear Element (SINE) repeats, transcriptionally active enhancers and the NEAT1 gene. This diverse set of non-coding genomic sites, which already appears to be an integral part of the cellular response to stress, may only represent the first of many. Thus, the study of the evolutionary conserved heat stress response has the potential to emerge as a powerful cellular context to study lncRNAs, produced from repeated or unique DNA regions, with a regulatory function that is often well-documented but a mode of action that remains largely unknown.
Assuntos
RNA Longo não Codificante , Animais , DNA , Fatores de Transcrição de Choque Térmico/genética , Fatores de Transcrição de Choque Térmico/metabolismo , Resposta ao Choque Térmico/genética , Mamíferos/genética , RNA Longo não Codificante/genética , RNA Longo não Codificante/metabolismo , Fatores de Transcrição/metabolismoRESUMO
Heat stress (HS) induces a cellular response leading to profound changes in gene expression. Here, we show that human YTHDC1, a reader of N6-methyladenosine (m6A) RNA modification, mostly associates to the chromatin fraction and that HS induces a redistribution of YTHDC1 across the genome, including to heat-induced heat shock protein (HSP) genes. YTHDC1 binding to m6A-modified HSP transcripts co-transcriptionally promotes expression of HSPs. In parallel, hundreds of the genes enriched in YTHDC1 during HS have their transcripts undergoing YTHDC1- and m6A-dependent intron retention. Later, YTHDC1 concentrates within nuclear stress bodies (nSBs) where it binds to m6A-modified SATIII non-coding RNAs, produced in an HSF1-dependent manner upon HS. These findings reveal that YTHDC1 plays a central role in a chromatin-associated m6A-based reprogramming of gene expression during HS. Furthermore, they support the model where the subsequent and temporary sequestration of YTHDC1 within nSBs calibrates the timing of this YTHDC1-dependent gene expression reprogramming.
Assuntos
Cromatina , Resposta ao Choque Térmico , Humanos , Resposta ao Choque Térmico/genética , Proteínas de Choque Térmico/metabolismo , Expressão Gênica , Fatores de Processamento de RNA/metabolismo , Proteínas do Tecido Nervoso/metabolismoRESUMO
Centromeric and pericentric regions have long been regarded as transcriptionally inert portions of chromosomes. A number of studies in the past 10 years disproved this dogma and provided convincing evidence that centromeric and pericentric sequences are transcriptionally active in several biological contexts.In this chapter, we provide a comprehensive picture of the various contexts (cell growth and differentiation, stress, effect of chromatin organization) in which these sequences are expressed in mouse and human cells and discuss the possible functional implications of centromeric and pericentric sequences activation and/or of the resulting noncoding RNAs. Moreover, we provide an overview of the molecular mechanisms underlying the activation of centromeric and pericentromeric sequences as well as the structural features of encoded RNAs.
Assuntos
Centrômero , DNA Satélite , Animais , Humanos , Mamíferos/genéticaRESUMO
Cells respond to stress by activating the synthesis of heat shock proteins (HSPs) which protect the cells against the deleterious effects of stress. This mechanism is controlled by the heat shock factor 1 (HSF1). In parallel to HSP gene transcription, in human cells, HSF1 also binds to and transcribes satellite III repeated sequences present in numerous copies in the 9q12 pericentromeric region of chromosome 9. These HSF1 accumulation sites are termed nuclear stress bodies (nSBs). In tumor cells, however, the number of nSBs is higher than the number of 9q12 copies, suggesting the existence of other HSF1 targets. In this paper, we were interested in characterizing these other HSF1 binding sites. We show that HSF1 indeed binds to the pericentromeric region of 14 chromosomes, thereby directing the formation of 'secondary nSBs'. The appearance of secondary nSBs depends on the number of satellite sequences present in the target locus, and on the cellular amount of HSF1 protein. Moreover, secondary nSBs also correspond to transcription sites, thus demonstrating that heat shock induces a genome-wide transcription of satellite sequences. Finally, by analyzing published transcriptomic data, we show that the derepression of these large heterochromatic blocks does not significantly affect the transcription of neighboring genes.
Assuntos
DNA Satélite/genética , DNA Satélite/metabolismo , Proteínas de Ligação a DNA/metabolismo , Resposta ao Choque Térmico/genética , Resposta ao Choque Térmico/fisiologia , Fatores de Transcrição/metabolismo , Animais , Sítios de Ligação/genética , Linhagem Celular , Centrômero/genética , Centrômero/metabolismo , Cromossomos Humanos/genética , Cromossomos Humanos/metabolismo , Primers do DNA/genética , Proteínas de Ligação a DNA/genética , Células HeLa , Fatores de Transcrição de Choque Térmico , Humanos , Células Híbridas , Hibridização in Situ Fluorescente , Camundongos , Proteínas Recombinantes de Fusão/genética , Proteínas Recombinantes de Fusão/metabolismo , Fatores de Transcrição/genética , Transcrição GênicaRESUMO
Although there is now evidence that the expression of centromeric (CT) and pericentric (PCT) sequences are key players in major genomic functions, their transcriptional status in human cells is still poorly known. The main reason for this lack of data is the complexity and high level of polymorphism of these repeated sequences, which hampers straightforward analyses by available transcriptomic approaches. Here a transcriptomic macro-array dedicated to the analysis of CT and PCT expression is developed and validated in heat-shocked (HS) HeLa cells. For the first time, the expression status of CT and PCT sequences is analyzed in a series of normal and cancer human cells and tissues demonstrating that they are repressed in all normal tissues except in the testis, where PCT transcripts are found. Moreover, PCT sequences are specifically expressed in HS cells in a Heat-Shock Factor 1 (HSF1)-dependent fashion, and we show here that another independent pathway, involving DNA hypo-methylation, can also trigger their expression. Interestingly, CT and PCT were found illegitimately expressed in somatic cancer samples, whereas PCT were repressed in testis cancer, suggesting that the expression of CT and PCT sequences may represent a good indicator of epigenetic deregulations occurring in response to environmental changes or in cell transformation.
Assuntos
Centrômero/metabolismo , Linhagem Celular Tumoral , Centrômero/química , Montagem e Desmontagem da Cromatina , Perfilação da Expressão Gênica , Células HeLa , Resposta ao Choque Térmico , Humanos , Análise de Sequência com Séries de Oligonucleotídeos , Ribonuclease III/metabolismoRESUMO
Activation of the mammalian heat shock transcription factor (HSF)1 by stress is a multistep process resulting in the transcription of heat shock genes. Coincident with these events is the rapid and reversible redistribution of HSF1 to discrete nuclear structures termed HSF1 granules, whose function is still unknown. Key features are that the number of granules correlates with cell ploidy, suggesting the existence of a chromosomal target. Here we show that in humans, HSF1 granules localize to the 9q11-q12 heterochromatic region. Within this locus, HSF1 binds through direct DNA-protein interaction with a nucleosome-containing subclass of satellite III repeats. HSF1 granule formation only requires the DNA binding competence and the trimerization of the factor. This is the first example of a transcriptional activator that accumulates transiently and reversibly on a chromosome-specific heterochromatic locus.
Assuntos
Cromossomos Humanos Par 9/genética , Grânulos Citoplasmáticos/genética , Proteínas de Ligação a DNA/genética , Proteínas de Choque Térmico/genética , Heterocromatina/genética , Estresse Fisiológico/genética , Sítios de Ligação/genética , Núcleo Celular/genética , Núcleo Celular/metabolismo , Células Cultivadas , Grânulos Citoplasmáticos/metabolismo , Proteínas de Ligação a DNA/metabolismo , Feminino , Imunofluorescência , Fatores de Transcrição de Choque Térmico , Proteínas de Choque Térmico/metabolismo , Humanos , Repetições de Microssatélites/genética , Estrutura Terciária de Proteína/genética , Estresse Fisiológico/metabolismo , Fatores de TranscriçãoRESUMO
Exposure of mammalian cells to stress induces the activation of heat shock transcription factor 1 (HSF1) and the subsequent transcription of heat shock genes. Activation of the heat shock response also correlates with a rapid relocalization of HSF1 within a few nuclear structures termed nuclear stress granules. These stress-induced structures, which form primarily on the 9q12 region in humans through direct binding of HSF1 to satellite III repeats, do not colocalize with transcription sites of known hsp genes. In this paper, we show that nuclear stress granules correspond to RNA polymerase II transcription factories where satellite III repeats are transcribed into large and stable RNAs that remain associated with the 9q12 region, even throughout mitosis. This work not only reveals the existence of a new major heat-induced transcript in human cells that may play a role in chromatin structure, but also provides evidence for a transcriptional activity within a locus considered so far as heterochromatic and silent.
Assuntos
Estruturas do Núcleo Celular/genética , DNA Satélite/genética , Proteínas de Ligação a DNA/genética , RNA Polimerase II/biossíntese , Estresse Fisiológico/genética , Ativação Transcricional/genética , Acetilação , Proteína de Ligação a CREB , Estruturas do Núcleo Celular/metabolismo , Estruturas do Núcleo Celular/ultraestrutura , Cromossomos Humanos Par 9/genética , Proteínas de Ligação a DNA/metabolismo , Células HeLa , Fatores de Transcrição de Choque Térmico , Heterocromatina/genética , Histonas/genética , Histonas/metabolismo , Humanos , Proteínas Nucleares/genética , Proteínas Nucleares/metabolismo , Transporte Proteico/genética , RNA/biossíntese , RNA/genética , Estresse Fisiológico/metabolismo , Transativadores/genética , Transativadores/metabolismo , Fatores de TranscriçãoRESUMO
The heat shock response is characterized by the transcriptional activation of both hsp genes and noncoding and repeated satellite III DNA sequences located at pericentric heterochromatin. Both events are under the control of Heat Shock Factor I (HSF1). Here we show that under heat shock, HSF1 recruits major cellular acetyltransferases, GCN5, TIP60 and p300 to pericentric heterochromatin leading to a targeted hyperacetylation of pericentric chromatin. Redistribution of histone acetylation toward pericentric region in turn directs the recruitment of Bromodomain and Extra-Terminal (BET) proteins BRD2, BRD3, BRD4, which are required for satellite III transcription by RNAP II. Altogether we uncover here a critical role for HSF1 in stressed cells relying on the restricted use of histone acetylation signaling over pericentric heterochromatin (HC).
Assuntos
Resposta ao Choque Térmico , Heterocromatina/genética , Transdução de Sinais/genética , Ativação Transcricional , Animais , Células COS , Proteínas de Ciclo Celular , Chlorocebus aethiops , Células HeLa , Fatores de Transcrição de Choque Térmico/genética , Fatores de Transcrição de Choque Térmico/metabolismo , Heterocromatina/metabolismo , Histona Acetiltransferases/metabolismo , Histonas/metabolismo , Humanos , Proteínas Nucleares/genética , Proteínas Nucleares/metabolismo , Proteínas Serina-Treonina Quinases/genética , Proteínas Serina-Treonina Quinases/metabolismo , RNA Polimerase II/metabolismo , Proteínas de Ligação a RNA/genética , Proteínas de Ligação a RNA/metabolismo , Fatores de Transcrição/genética , Fatores de Transcrição/metabolismoRESUMO
The histones H2A, H2B, H3 and H4 are very conserved basic proteins that wrap almost two turns of DNA to form the nucleosome core. Conventional histones can be replaced with histone variants that are found in all eukaryotic organisms. Together with other nucleosome modification pathways, histone variants participate in the functional specialization of chromatin. In this review, we focus on three major H2A histone variants: H2A.X, H2A.Z and macroH2A. Recent discoveries highlight their involvement in crucial events such as DNA repair and transcriptional regulation.
Assuntos
Histonas/farmacologia , Nucleossomos/fisiologia , Animais , Reparo do DNA , Humanos , Transcrição GênicaRESUMO
After heat shock, HSF1 controls a major cellular transcriptional response involving the activation of early (HSP70) and late (HSP25) heat shock gene expression. Here we show that a full response to heat shock (activation of both HSP70 and HSP25) depends on the duration of HSF1 activation, which is itself controlled by HDAC6, a unique deacetylase known to bind monoubiquitin and polyubiquitin with high affinity. On the basis of a comparative analysis of the heat shock response in cells knocked out for HDAC6 or expressing HDAC6 mutants, we show that HDAC6 binding to ubiquitinated proteins controls the duration of HSF1 activation after heat shock. In cells expressing HDAC6 mutated in the ubiquitin-binding domain, the AAA ATPase factor p97/VCP mediates rapid inactivation of HSF1, precluding late activation of the HSP25 gene. In these cells, knockdown of p97/VCP rescues HSF1 from this rapid inactivation and restores HSP25 expression. We present here a new regulatory circuit that adjusts the duration of the heat shock response to the extent of protein ubiquitination after heat shock.
Assuntos
Proteínas de Ligação a DNA/metabolismo , Resposta ao Choque Térmico , Histona Desacetilases/fisiologia , Fatores de Transcrição/metabolismo , Ubiquitina/metabolismo , Células 3T3 , Adenosina Trifosfatases/metabolismo , Animais , Proteínas de Ciclo Celular/metabolismo , Regulação da Expressão Gênica , Fatores de Transcrição de Choque Térmico , Proteínas de Choque Térmico/metabolismo , Desacetilase 6 de Histona , Camundongos , Chaperonas Moleculares , Proteínas de Neoplasias/metabolismo , Regiões Promotoras Genéticas , Ligação Proteica , Domínios e Motivos de Interação entre Proteínas , Ubiquitinação , Proteína com ValosinaRESUMO
Heat shock factor 1 is the key transcription factor of the heat shock response. Its function is to protect the cell against the deleterious effects of stress. Upon stress, HSF1 binds to and transcribes hsp genes and repeated satellite III (sat III) sequences present at the 9q12 locus. HSF1 binding to pericentric sat III sequences forms structures known as nuclear stress bodies (nSBs). nSBs represent a natural amplification of RNA pol II dependent transcription sites. Dynamics of HSF1 and of deletion mutants were studied in living cells using multi-confocal Fluorescence Correlation Spectroscopy (mFCS) and Fluorescence Recovery After Photobleaching (FRAP). In this paper, we show that HSF1 dynamics modifications upon heat shock result from both formation of high molecular weight complexes and increased HSF1 interactions with chromatin. These interactions involve both DNA binding with Heat Shock Element (HSE) and sat III sequences and a more transient sequence-independent binding likely corresponding to a search for more specific targets. We find that the trimerization domain is required for low affinity interactions with chromatin while the DNA binding domain is required for site-specific interactions of HSF1 with DNA.
Assuntos
Proteínas de Ligação a DNA/genética , Proteínas de Ligação a DNA/metabolismo , Proteínas Mutantes/metabolismo , Mutação/genética , Fatores de Transcrição/genética , Fatores de Transcrição/metabolismo , Animais , Permeabilidade da Membrana Celular , Estruturas do Núcleo Celular/metabolismo , Fracionamento Químico , DNA/metabolismo , Proteínas de Ligação a DNA/química , Difusão , Recuperação de Fluorescência Após Fotodegradação , Técnicas de Silenciamento de Genes , Proteínas de Fluorescência Verde/metabolismo , Proteínas de Choque Térmico HSP70/metabolismo , Células HeLa , Fatores de Transcrição de Choque Térmico , Resposta ao Choque Térmico/genética , Humanos , Espaço Intracelular/metabolismo , Camundongos , Peso Molecular , Proteínas Mutantes/química , Ligação Proteica , Estrutura Terciária de Proteína , Transporte Proteico , RNA/metabolismo , Espectrometria de Fluorescência , Estresse Fisiológico , Frações Subcelulares/metabolismo , Fatores de Transcrição/química , Ativação Transcricional/genéticaRESUMO
Nuclear stress bodies (nSBs) are unique subnuclear organelles which form in response to heat shock. They are initiated through a direct interaction between heat shock transcription factor 1 (HSF1) and pericentric tandem repeats of satellite III sequences and correspond to active transcription sites for noncoding satellite III transcripts. Given their unusual features, nSBs are distinct from other known transcription sites. In stressed cells, they are thought to participate in rapid, transient, and global reprogramming of gene expression through different types of mechanisms including chromatin remodeling and trapping of transcription and splicing factors. The analysis of these atypical and intriguing structures uncovers new facets of the relationship between nuclear organization and nuclear function.
Assuntos
Espaço Intranuclear , Organelas , Estresse Fisiológico/fisiologia , Animais , DNA Satélite , Transcrição GênicaRESUMO
Epigenetic perturbations are increasingly described in cancer cells where they are thought to contribute to deregulated gene expression and genome instability. Here, we report the first evidence that a distinct category of chromosomal translocations observed in human tumours--those targeting 1q12 satellite DNA--can directly mediate such perturbations by promoting the formation of aberrant heterochromatic foci (aHCF). By detailed investigations of a 1q12 translocation to chromosome 2p, in a case of human B cell lymphoma, aberrant aHCF were shown to be localized to the nuclear periphery and to arise as a consequence of long range 'pairing' between the translocated 1q12 and chromosome 2 centromeric regions. Remarkably, adjacent 2p sequences showed increased levels of repressive histone modifications, including H4K20me3 and H3K9me3, and were bound by HP1. aHCF were associated to aberrant spatial localization and deregulated expression of a novel 2p gene (GMCL1) that was found to have prognostic impact in diffuse large B cell lymphoma. Thus constitutive heterochromatin rearrangements can contribute to tumourigenesis by perturbing gene expression via long range epigenetic mechanisms.
Assuntos
Núcleo Celular/genética , Cromossomos Humanos Par 1/genética , Regulação Neoplásica da Expressão Gênica , Heterocromatina/genética , Linfoma de Células B/genética , Translocação Genética , Cromossomos Humanos Par 2/genética , HumanosRESUMO
In the fission yeast, S. Pombe, small dsRNA generated by RNAi-dependent mechanisms are involved in the establishment and maintenance of heterochromatic regions. The existence of conserved features within the general organization of centromeric and pericentromeric repeats in yeast, mouse and human argues in favor of a conserved role for centromeric and pericentromeric-derived transcripts across these species. In support of this, evidence is accumulating that centromeric and pericentromeric sequences are transcriptionally competent in diverse biological contexts in mammalian cells. Given the importance of centromeric and pericentromeric regions, not only with respect to centromere function, but also to gene regulation, this review examines the biological contexts in which mouse and human centromeric and pericentromeric-specific transcripts have been observed. The structure of the transcripts generated, the molecular mechanisms underlying their expression and their supposed functions will be discussed.
Assuntos
Centrômero/genética , Heterocromatina/genética , Sequências Repetitivas de Ácido Nucleico/genética , Transcrição Gênica , Animais , Sequência de Bases , Regulação da Expressão Gênica , Humanos , Modelos Biológicos , Dados de Sequência MolecularRESUMO
A major regulatory function has been evidenced here for HSF1, the key transcription factor of the heat-shock response, in a large-scale remodeling of the cell epigenome. Indeed, upon heat shock, HSF1, in addition to its well-known transactivating activities, mediates a genome-wide and massive histone deacetylation. Investigating the underlying mechanisms, we show that HSF1 specifically associates with and uses HDAC1 and HDAC2 to trigger this heat-shock-dependent histone deacetylation. This work therefore identifies HSF1 as a master regulator of global chromatin acetylation and reveals a cross-talk between HSF1 and histone deacetylases in the general control of genome organization in response to heat shock.
Assuntos
Proteínas de Ligação a DNA/metabolismo , Regulação da Expressão Gênica , Genoma , Resposta ao Choque Térmico/genética , Fatores de Transcrição/metabolismo , Acetilação , Animais , Linhagem Celular , Cromatina/metabolismo , Proteínas de Ligação a DNA/genética , Fatores de Transcrição de Choque Térmico , Histona Desacetilase 1/genética , Histona Desacetilase 1/metabolismo , Histona Desacetilase 2/genética , Histona Desacetilase 2/metabolismo , Histonas/genética , Histonas/metabolismo , Humanos , Camundongos , RNA Interferente Pequeno/genética , RNA Interferente Pequeno/metabolismo , Fatores de Transcrição/genéticaRESUMO
Organisms respond to circumstances threatening the cellular protein homeostasis by activation of heat-shock transcription factors (HSFs), which play important roles in stress resistance, development, and longevity. Of the four HSFs in vertebrates (HSF1-4), HSF1 is activated by stress, whereas HSF2 lacks intrinsic stress responsiveness. The mechanism by which HSF2 is recruited to stress-inducible promoters and how HSF2 is activated is not known. However, changes in the HSF2 expression occur, coinciding with the functions of HSF2 in development. Here, we demonstrate that HSF1 and HSF2 form heterotrimers when bound to satellite III DNA in nuclear stress bodies, subnuclear structures in which HSF1 induces transcription. By depleting HSF2, we show that HSF1-HSF2 heterotrimerization is a mechanism regulating transcription. Upon stress, HSF2 DNA binding is HSF1 dependent. Intriguingly, when the elevated expression of HSF2 during development is mimicked, HSF2 binds to DNA and becomes transcriptionally competent. HSF2 activation leads to activation of also HSF1, revealing a functional interdependency that is mediated through the conserved trimerization domains of these factors. We propose that heterotrimerization of HSF1 and HSF2 integrates transcriptional activation in response to distinct stress and developmental stimuli.
Assuntos
Proteínas de Ligação a DNA/metabolismo , Proteínas de Choque Térmico/metabolismo , Resposta ao Choque Térmico/fisiologia , Fatores de Transcrição/metabolismo , Ativação Transcricional , Animais , Linhagem Celular , DNA Satélite/metabolismo , Proteínas de Ligação a DNA/genética , Proteínas de Ligação a DNA/fisiologia , Fatores de Transcrição de Choque Térmico , Proteínas de Choque Térmico/genética , Proteínas de Choque Térmico/fisiologia , Humanos , Masculino , Camundongos , Testículo/metabolismo , Fatores de Transcrição/genética , Fatores de Transcrição/fisiologia , Transcrição GênicaRESUMO
Half of the human genome consists of repetitive DNA sequences. Recent studies in various organisms highlight the role of chromatin regulation of repetitive DNA in gene regulation as well as in maintainance of chromosomes and genome integrity. Hence, repetitive DNA sequences might be potential "sensors" for chromatin changes associated with pathogenesis. Therefore, we developed a new genomic tool called RepArray. RepArray is a repeat-specific microarray composed of a representative set of human repeated sequences including transposon-derived repeats, simple sequences repeats, tandemly repeated sequences such as centromeres and telomeres. We showed that combined to anti-methylcytosine immunoprecipitation assay, the RepArray can be used to generate repeat-specific methylation maps. Using cell lines impaired chemically or genetically for DNA methyltransferases activities, we were able to distinguish different epigenomes demonstrating that repeats can be used as markers of genome-wide methylation changes. Besides, using a well-documented system model, the thermal stress, we demonstrated that RepArray is also a fast and reliable tool to obtain an overview of overall transcriptional activity on whole repetitive compartment in a given cell type. Thus, the RepArray represents the first valuable tool for systematic and genome-wide analyses of the methylation and transcriptional status of the repetitive counterpart of the human genome.