RESUMO
Thyroid hormones (THs) are important regulators of growth, development, and homeostasis of all vertebrates. There are many environmental contaminants that are known to disrupt TH action, yet their mechanisms are only partially understood. While the effects of Endocrine Disrupting Chemicals (EDCs) are mostly studied as "hormone system silos", the present critical review highlights the complexity of EDCs interfering with TH function through their interactions with other hormonal axes involved in reproduction, stress, and energy metabolism. The impact of EDCs on components that are shared between hormone signaling pathways or intersect between pathways can thus extend beyond the molecular ramifications to cellular, physiological, behavioral, and whole-body consequences for exposed organisms. The comparatively more extensive studies conducted in mammalian models provides encouraging support for expanded investigation and highlight the paucity of data generated in other non-mammalian vertebrate classes. As greater genomics-based resources become available across vertebrate classes, better identification and delineation of EDC effects, modes of action, and identification of effective biomarkers suitable for HPT disruption is possible. EDC-derived effects are likely to cascade into a plurality of physiological effects far more complex than the few variables tested within any research studies. The field should move towards understanding a system of hormonal systems' interactions rather than maintaining hormone system silos.
Assuntos
Disruptores Endócrinos , Animais , Disruptores Endócrinos/toxicidade , Sistema Endócrino , Humanos , Reprodução , Glândula Tireoide , Hormônios TireóideosRESUMO
Methyl-CpG-binding protein 2 (MeCP2) is a chromatin-binding protein that mediates transcriptional regulation, and is highly abundant in brain. The nature of its binding to reconstituted templates has been well characterized in vitro. However, its interactions with native chromatin are less understood. Here we show that MeCP2 displays a distinct distribution within fractionated chromatin from various tissues and cell types. Artificially induced global changes in DNA methylation by 3-aminobenzamide or 5-aza-2'-deoxycytidine, do not significantly affect the distribution or amount of MeCP2 in HeLa S3 or 3T3 cells. Most MeCP2 in brain is chromatin-bound and localized within highly nuclease-accessible regions. We also show that, while in most tissues and cell lines, MeCP2 forms stable complexes with nucleosome, in brain, a fraction of it is loosely bound to chromatin, likely to nucleosome-depleted regions. Finally, we provide evidence for novel associations of MeCP2 with mononucleosomes containing histone H2A.X, H3K9me(2) and H3K27me(3) in different chromatin fractions from brain cortex and in vitro. We postulate that the functional compartmentalization and tissue-specific distribution of MeCP2 within different chromatin types may be directed by its association with nucleosomes containing specific histone variants, and post-translational modifications.
Assuntos
Encéfalo/metabolismo , Cromatina/metabolismo , Histonas/metabolismo , Proteína 2 de Ligação a Metil-CpG/metabolismo , Nucleossomos/metabolismo , Animais , Núcleo Celular/metabolismo , Cromatina/ultraestrutura , DNA/metabolismo , Metilação de DNA , Desoxirribonucleases , Células HeLa , Histonas/química , Humanos , Neurônios/metabolismo , Neurônios/ultraestrutura , Ligação Proteica , Processamento de Proteína Pós-Traducional , RatosRESUMO
Oil spills that occur in high traffic coastal environments can have profound consequences for the health of marine ecosystems and the commercial and social interests that are dependent upon these habitats. Given that the global reliance on marine fuels is not abating, it is imperative to develop sensitive and robust tools to monitor oil contamination and remediation in a timely manner. Such tools are increasingly important for ascertaining the immediate and long-term effects of oil contamination on species of interest and local habitats as water-soluble components of oils, such as polycyclic aromatic hydrocarbons (PAHs), can persist post-remediation. We previously demonstrated that 3-methylcholanthrene responsive cytochrome P450-1a (cyp1a1) transcript abundance in the liver and caudal fin of coho salmon smolts (Onchorhynchus kisutch) was sensitive to exposure to low sulfur marine diesel (LSMD) seawater accommodated fractions (seaWAF) in cold water. We expanded upon this paradigm by assessing the utility of the cyp1a1 transcript to track both exposure to LSMD seaWAF and recovery from exposure by measuring cyp1a1 abundance in coho smolts using quantitative polymerase chain reaction (qPCR). Smolts were exposed to either 100 mg/L LSMD seaWAF or clean seawater (control) for 4 days. Fish were then transferred to clean seawater for depuration and tissues sampled at 0, 1, 2, 4, and 8 days from both treatments. Livers and caudal fins were dissected from 40 smolts per group (ntotal = 400 smolts). The LSMD seaWAF-induced cyp1a1 transcript levels significantly decreased one day after depuration in the liver and caudal fin in a sex-independent manner in genotyped females and males. After four days of depuration, cyp1a1 transcript abundance decreased to baseline control levels, regardless of tissue or sex. The present study demonstrates the value of using the caudal fin as a reliable, sensitive, and non-lethal sampling and monitoring tool.
Assuntos
Oncorhynchus kisutch , Poluentes Químicos da Água , Animais , Masculino , Feminino , Água , Oncorhynchus kisutch/genética , Ecossistema , Poluentes Químicos da Água/toxicidade , Sistema Enzimático do Citocromo P-450 , EnxofreRESUMO
Thyroid hormones (THs) are important developmental regulators in vertebrates, including during the metamorphosis of a tadpole into a frog. Metamorphosis is a post-embryonic developmental period initiated by TH production in the tadpole thyroid gland. The two main bioactive forms of TH are L-thyroxine (T4) and 3,5,3'-triiodothyronine (T3); these hormones have overlapping but distinct mechanisms of action. Premetamorphic tadpoles are highly responsive to TH and can be induced to metamorphose through exogenous TH exposure, making them an important model for both the study of vertebrate TH signaling and endocrine disrupting chemicals (EDCs). It is important to differentiate TH-mediated responses from estrogenic responses in premetamorphic tadpoles when assessing dysregulation by EDCs as crosstalk between the two endocrine systems is well-documented. Herein, we compare the RNA-sequencing-derived transcriptomic profiles of three TH-responsive tissues (liver, olfactory epithelium, and tail fin) in premetamorphic bullfrog (Rana [Lithobates] catesbeiana) tadpoles exposed to T3, T4, and estradiol (E2). These profiles were generated using the latest available genome assembly for the species. The data indicate that there is a clear distinction, and little overlap, between the transcriptomic responses elicited by E2 and the THs. In contrast, within the THs, the T3- and T4-induced transcriptomic profiles generally show considerable overlap; however, the degree of overlap is highly tissue-dependent, illustrating the importance of distinguishing the two THs and the affected signaling pathways within the target tissue type when evaluating hormone active agents. The data herein also show that E2 and TH treatment can uniquely induce significant changes in expression of their respective "classic" bioindicator transcripts vtg (E2) and thra, thrb, and thibz (THs). However, care must be taken in the interpretation of increased vep or esr1 transcripts as a change in transcript levels can be induced by THs rather than solely E2.
Assuntos
Disruptores Endócrinos , Poluentes Químicos da Água , Animais , Larva/genética , Larva/metabolismo , Transcriptoma , Poluentes Químicos da Água/toxicidade , Hormônios Tireóideos/metabolismo , Tri-Iodotironina/metabolismo , Ranidae/metabolismo , Estrogênios/toxicidade , Estrogênios/metabolismo , Disruptores Endócrinos/toxicidade , Disruptores Endócrinos/metabolismo , Mucosa Olfatória , Fígado/metabolismoRESUMO
Mutations in the methyl-CpG-binding protein 2 (MeCP2) cause Rett syndrome, a severe neurodevelopmental disease associated with ataxia and other post-natal symptoms similar to autism. Much research interest has focussed on the implications of MeCP2 in disease and neuron physiology. However, little or no attention has been paid to how MeCP2 turnover is regulated. The post-translational control of MeCP2 is of critical importance, especially as subtle increases or decreases in MeCP2 amounts can affect neuron morphology and function. The latter point is of particular importance for gene therapeutic approaches in which exogenous wild-type MeCP2 is being introduced into diseased neurons. Further to this, we propose two hypotheses. The first hypothesis discusses the poly-ubiquitin-mediated post-translational regulation of MeCP2 through its two PEST domains. The second hypothesis explores the use of histone deacetylase inhibitors to modulate the amounts of MeCP2 expressed in conjunction with the aforementioned therapeutic approaches.
Assuntos
Proteína 2 de Ligação a Metil-CpG/metabolismo , Processamento de Proteína Pós-Traducional/fisiologia , Síndrome de Rett/metabolismo , Sequência de Aminoácidos , Animais , Humanos , Proteína 2 de Ligação a Metil-CpG/química , Proteína 2 de Ligação a Metil-CpG/genética , Dados de Sequência Molecular , Fosforilação/genética , Fosforilação/fisiologia , Processamento de Proteína Pós-Traducional/genética , Estrutura Terciária de Proteína/genética , Estrutura Terciária de Proteína/fisiologia , Homologia de Sequência de Aminoácidos , Ubiquitinação/genética , Ubiquitinação/fisiologiaRESUMO
There has been a hotbed of activity surrounding MeCP2 research in the past number of years. Despite better characterizing the functions and nature of this protein, it has become abundantly clear that MeCP2 is involved in far more complex activities than perhaps initially anticipated. Recent publications have shown that MeCP2 is dynamically post-translationally modified, and it is possible that these marks permit MeCP2 to inhabit very diverse chromatin environments. It is also of interest to consider how nucleosome composition differs in these varying chromatin regions, and how the chromatin template itself contributes to diversifying the regulatory roles of MeCP2. These will be critical points to examine when seeking to understand how MeCP2 behaviour differentiates in tissues other than the brain. By understanding the chromatin and (or) tissue context in which MeCP2 interacts, it may be possible to discern the specific etiology of diseases linked to MeCP2 dysfunction.
Assuntos
Regulação da Expressão Gênica/fisiologia , Proteína 2 de Ligação a Metil-CpG/fisiologia , Nucleossomos/fisiologia , Animais , Sítios de Ligação/genética , Humanos , Proteína 2 de Ligação a Metil-CpG/metabolismo , Modelos Biológicos , Nucleossomos/química , Nucleossomos/genética , Nucleossomos/metabolismo , Sequências Reguladoras de Ácido Nucleico/genética , Sequências Reguladoras de Ácido Nucleico/fisiologia , Síndrome de Rett/genética , Síndrome de Rett/metabolismo , Fatores de TempoRESUMO
[This corrects the article DOI: 10.3389/fendo.2019.00276.].
RESUMO
Aquatic and terrestrial environments are increasingly contaminated by anthropogenic sources that include pharmaceuticals, personal care products, and industrial and agricultural chemicals (i. e., pesticides). Many of these substances have the potential to disrupt endocrine function, yet their effect on thyroid hormone (TH) action has garnered relatively little attention. Anuran postembryonic metamorphosis is strictly dependent on TH and perturbation of this process can serve as a sensitive barometer for the detection and mechanistic elucidation of TH disrupting activities of chemical contaminants and their complex mixtures. The ecological threats posed by these contaminants are further exacerbated by changing environmental conditions such as temperature, photoperiod, pond drying, food restriction, and ultraviolet radiation. We review the current knowledge of several chemical and environmental factors that disrupt TH-dependent metamorphosis in amphibian tadpoles as assessed by morphological, thyroid histology, behavioral, and molecular endpoints. Although the molecular mechanisms for TH disruption have yet to be determined for many chemical and environmental factors, several affect TH synthesis, transport or metabolism with subsequent downstream effects. As molecular dysfunction typically precedes phenotypic or histological pathologies, sensitive assays that detect changes in transcript, protein, or metabolite abundance are indispensable for the timely detection of TH disruption. The emergence and application of 'omics techniques-genomics, transcriptomics, proteomics, metabolomics, and epigenomics-on metamorphosing tadpoles are powerful emerging assets for the rapid, proxy assessment of toxicant or environmental damage for all vertebrates including humans. Moreover, these highly informative 'omics techniques will complement morphological, behavioral, and histological assessments, thereby providing a comprehensive understanding of how TH-dependent signal disruption is propagated by environmental contaminants and factors.
RESUMO
Methyl CpG binding protein 2 (MeCP2) is a basic protein that contains a DNA methyl binding domain. The mechanism by which the highly positive charge of MeCP2 and its ability to bind methylated DNA contribute to the specificity of its binding to chromatin has long remained elusive. In this paper, we show that MeCP2 binds to nucleosomes in a very similar way to linker histones both in vitro and in vivo. However, its binding specificity strongly depends on DNA methylation. We also observed that as with linker histones, this binding is independent of the core histone H3 N-terminal tail and is not affected by histone acetylation.
Assuntos
Cromatina/metabolismo , Metilação de DNA , Histonas/metabolismo , Proteína 2 de Ligação a Metil-CpG/metabolismo , Acetilação , Sequência de Bases , Sítios de Ligação , DNA/química , Células HeLa , Histonas/química , Humanos , Proteína 2 de Ligação a Metil-CpG/análise , Dados de Sequência Molecular , Nucleossomos/química , Nucleossomos/metabolismo , Ligação ProteicaRESUMO
PURPOSE: Since physical exertion is known to exacerbate the symptoms of chronic fatigue syndrome (CFS) and metabolic changes and including oxidative stress can modulate heat shock protein (HSP) expression responses, we sought to determine whether HSP expression is altered in CFS patients before and after exercise. Heat shock proteins (HSPs) in peripheral blood mononuclear cells (PBMC) were examined from 6 chronic fatigue syndrome (CFS) patients and 7 controls before and after a standardized treadmill exercise. Basal hsp27 was significantly higher among CFS patients compared to controls, and decreased immediately post-exercise, remaining below basal levels even at 7 days. A similar pattern was observed for HSP60, which gradually decreased in CFS patients but increased in controls post-exercise. These findings suggest an abnormal adaptive response to oxidative stress in CFS, and raise the possibility that HSP profiling may provide a more objective biologic marker for this illness. METHODS: HSP27, HSP60, HSP70 and HSP90 expression from 6 CFS patients and 7 age- and sex-matched controls were examined by western blot analysis of peripheral blood mononuclear cells immediately before, after, and at 1 day and 7 days following a standardized treadmill exercise. RESULTS: Basal HSP27 was higher among CFS patients than in controls (0.54 +/- 0.13 vs. 0.19 +/- 0.06, mean +/- SEM; P < 0.01). In addition, these levels in CFS patients decreased immediately post-exercise (0.25 +/- 0.09; P < 0.05) and remained below basal levels at day 1 post-exercises (0.18 +/- 0.05; P < 0.05). P < 0.05). This declining expression of HSP27 during the post-exercise period among CFS patients was confirmed by one-way ANOVA analysis with repeated measures (P < 0.05). In contrast, HSP27 levels remained relatively constant following exercise among control subjects. Similar patterns of declining HSP levels in CFS patients were also observed for HSP60 (0.94 +/- 0.40 vs. 1.32 +/- 0.46; P < 0.05), and for HSP90 (0.34 +/- 0.09 vs. 0.49 +/- 0.10; P < 0.05) at day 7 post-exercise compared with basal levels, respectively. In contrast, HSP60 levels in control subjects increased at day 1 (1.09 +/- 0.27) and day 7 (1.24 +/- 0.50) post-exercise compared to corresponding levels immediately post-exercise (0.55 +/- 0.06) (P < 0.05, respectively). CONCLUSION: These preliminary findings suggest an abnormal or defective adaptive response to oxidative stress in CFS, and raise the possibility that HSP profiling may provide a more objective biologic marker for this illness.
Assuntos
Exercício Físico/fisiologia , Síndrome de Fadiga Crônica/sangue , Proteínas de Choque Térmico/sangue , Leucócitos Mononucleares/metabolismo , Biomarcadores/sangue , Western Blotting , Chaperonina 60/sangue , Teste de Esforço , Síndrome de Fadiga Crônica/fisiopatologia , Feminino , Proteínas de Choque Térmico HSP27/sangue , Proteínas de Choque Térmico HSP70/sangue , Proteínas de Choque Térmico HSP90/sangue , Humanos , Leucócitos Mononucleares/citologia , Masculino , Pessoa de Meia-IdadeRESUMO
MeCP2 binds to methylated DNA in a chromatin context and has an important role in cancer and brain development and function. Histone deacetylase (HDAC) inhibitors are currently being used to palliate many cancer and neurological disorders. Yet, the molecular mechanisms involved are not well known for the most part and, in particular, the relationship between histone acetylation and MeCP2 is not well understood. In this paper, we study the effect of the HDAC inhibitor trichostatin A (TSA) on MeCP2, a protein whose dysregulation plays an important role in these diseases. We find that treatment of cells with TSA decreases the phosphorylation state of this protein and appears to result in a higher MeCP2 chromatin binding affinity. Yet, the binding dynamics with which the protein binds to DNA appear not to be significantly affected despite the chromatin reorganization resulting from the high levels of acetylation. HDAC inhibition also results in an overall decrease in MeCP2 levels of different cell lines. Moreover, we show that miR132 increases upon TSA treatment, and is one of the players involved in the observed downregulation of MeCP2.
Assuntos
Cromatina/metabolismo , Inibidores de Histona Desacetilases/farmacologia , Ácidos Hidroxâmicos/farmacologia , Proteína 2 de Ligação a Metil-CpG/genética , Processamento de Proteína Pós-Traducional/efeitos dos fármacos , Células 3T3 , Animais , Células HEK293 , Células HeLa , Humanos , Proteína 2 de Ligação a Metil-CpG/metabolismo , Camundongos , Fosforilação , Ligação Proteica/efeitos dos fármacosRESUMO
Structural variability within histone families, such as H2A, can be achieved through 2 primary mechanisms: the expression of histone variants and the incorporation of chemical modifications. The histone H2A family contains several variants in addition to the canonical H2A forms. In this review, recent developments in the study of the heteromorphous variants H2A.X, H2A.Z, and macroH2A will be discussed. Particular focus will be given to the post-translational modifications (PTMs) of these variants, including phosphorylation, ubiquitination, acetylation, and methylation. The combination of the newly identified N- and C-terminal tail PTMs expands the multiplicity of roles that the individual H2A variants can perform. It is of additional interest that analogous sites within these different histone variants can be similarly modified. Whether this is a redundant function or a finely tuned one, designed to meet specific needs, remains to be elucidated.
Assuntos
Histonas/química , Histonas/metabolismo , Processamento de Proteína Pós-Traducional , Animais , Instabilidade Genômica , Humanos , Isoformas de Proteínas/química , Isoformas de Proteínas/metabolismoRESUMO
The functional and structural chromatin roles of H2A.Z are still controversial. This work represents a further attempt to resolve the current functional and structural dichotomy by characterizing chromatin structures containing native H2A.Z. We have analyzed the role of this variant in mediating the stability of the histone octamer in solution using gel-filtration chromatography at different pH. It was found that decreasing the pH from neutral to acidic conditions destabilized the histone complex. Furthermore, it was shown that the H2A.Z-H2B dimer had a reduced stability. Sedimentation velocity analysis of nucleosome core particles (NCPs) reconstituted from native H2A.Z-containing octamers indicated that these particles exhibit a very similar behavior to that of native NCPs consisting of canonical H2A. Sucrose gradient fractionation of native NCPs under different ionic strengths indicated that H2A.Z had a subtle tendency to fractionate with more stabilized populations. An extensive analysis of the salt-dependent dissociation of histones from hydroxyapatite-adsorbed chromatin revealed that, whereas H2A.Z co-elutes with H3-H4, hyperacetylation of histones (by treatment of chicken MSB cells with sodium butyrate) resulted in a significant fraction of this variant eluting with the canonical H2A. These studies also showed that the late elution of this variant (correlated to enhanced binding stability) was independent of the chromatin size and of the presence or absence of linker histones.
Assuntos
Cromatina/metabolismo , Histonas/metabolismo , Acetilação , Animais , Galinhas , Dicroísmo Circular , Dimerização , Eritrócitos/metabolismo , Histonas/sangue , Histonas/química , Histonas/isolamento & purificação , Concentração de Íons de Hidrogênio , Mardivirus/genética , Nucleossomos/metabolismoRESUMO
MacroH2A (mH2A) is a histone variant that is enriched in the inactivated X-chromosomes of mammalian females. To characterize the role of this protein in other nuclear processes we isolated chromatin particles from chicken liver, a vertebrate system that does not undergo X-inactivation. Chromatin digestion and fractionation studies determined that mH2A is evenly distributed at several levels of chromatin structure and stabilizes the nucleosome core particle in solution. However, at the level of the chromatosome, selective salt precipitation showed the existence of a mutually exclusive relationship between mH2A and H1, which may reveal functional redundancy between these proteins. Two-dimensional gel electrophoresis demonstrated the presence of one major population of mH2A containing nucleosomes, which may become ADP-ribosylated. This report provides new clues into how mH2A distribution and a previously unidentified post-translational modification may help regulate the repression of autosomal chromatin.
Assuntos
Galinhas/metabolismo , Cromatina/metabolismo , Histonas/metabolismo , Processamento de Proteína Pós-Traducional/fisiologia , Animais , Mecanismo Genético de Compensação de Dose , Eletroforese em Gel Bidimensional , Hepatócitos/metabolismo , Histonas/genética , Nucleossomos/metabolismo , Processamento de Proteína Pós-Traducional/genéticaRESUMO
Integration of histone variants into chromatin organization allows for functional specification of chromatin regions. Recent functional analyses of H2A.Z have ascribed to it a multiplicity of complex and often opposing roles in developmental and homeostatic regulation. However, although the manner in which this essential histone variant is able to mediate its effects is not entirely well understood, current work has sought to investigate its mode of action. It is becoming increasingly clear that H2A.Z does not necessarily act independently, but rather, in conjunction with trans-acting factors to elicit chromatin changes. The nature of these structural changes has remained somewhat controversial but nevertheless exemplifies the seemingly multifaceted nature of H2A.Z.
Assuntos
Histonas/fisiologia , Proteínas de Saccharomyces cerevisiae/fisiologia , Sequência de Aminoácidos , Animais , Núcleo Celular/metabolismo , Cromatina/química , Dimerização , Heterocromatina/química , Histonas/química , Humanos , Imunoprecipitação , Modelos Biológicos , Modelos Moleculares , Dados de Sequência Molecular , Ligação Proteica , Conformação Proteica , Estrutura Terciária de Proteína , Saccharomyces cerevisiae/fisiologia , Proteínas de Saccharomyces cerevisiae/química , Relação Estrutura-AtividadeRESUMO
Daunomycin is an anticancer drug that is well-known to interact with DNA in chromatin. Using a compositionally defined chicken erythrocyte chromatin fraction, we have obtained conclusive evidence that the drug is also able to interact with chromatin-bound linker histones without any noticeable binding to core histones. The drug can interact in an equal fashion with both histone H1 and H5 and to a greater extent with core histones H3/H4 and H2A/H2B as free proteins in solution. Thus, the binding of daunomycin to linker histones in the chromatin fiber is most likely due to the well-known higher accessibility of these histones to the surrounding environment of the fiber. Binding of daunomycin to linker histones appears to primarily involve the trypsin-resistant (winged-helix) domain of these proteins. The studies described here reveal the occurrence of a previously undisclosed mechanism for the antitumor activity of anthracycline drugs at the chromatin level.