RESUMO
We previously showed that fluorizoline, a fluorinated thiazoline compound, binds to both subunits of the mitochondrial prohibitin (PHB) complex, PHB1 and PHB2, being the expression of these proteins required for fluorizoline-induced apoptosis in mouse embryonic fibroblasts. To investigate the conservation of this apoptotic mechanism, we studied the effect of PHB downregulation on fluorizoline activity on two human cell lines, HEK293T and U2OS. Then, we asked whether PHBs mediate the effect of fluorizoline in a multicellular organism. Interestingly, reduced levels of PHBs in the human cells impaired the induction of apoptosis by fluorizoline. We observed that fluorizoline has a detrimental dose-dependent effect on the development and survival of the nematode model Caenorhabditis elegans. Besides, such effects of fluorizoline treatment in living nematodes were absent in PHB mutants. Finally, we further explored the apoptotic pathway triggered by fluorizoline in human cell lines. We found that the BH3-only proteins NOXA, BIM and PUMA participate in fluorizoline-induced apoptosis and that the induction of NOXA and PUMA is dependent on PHB expression.
Assuntos
Apoptose/efeitos dos fármacos , Proteínas de Caenorhabditis elegans/metabolismo , Caenorhabditis elegans/citologia , Proteínas Repressoras/metabolismo , Tiazolidinas/farmacologia , Animais , Caenorhabditis elegans/efeitos dos fármacos , Caenorhabditis elegans/genética , Caenorhabditis elegans/metabolismo , Proteínas de Caenorhabditis elegans/genética , Células HEK293 , Humanos , Proibitinas , Proteínas Repressoras/genética , Tiazolidinas/químicaRESUMO
BACKGROUND: Advances in automated image-based microscopy platforms coupled with high-throughput liquid workflows have facilitated the design of large-scale screens utilising multicellular model organisms such as Caenorhabditis elegans to identify genetic interactions, therapeutic drugs or disease modifiers. However, the analysis of essential genes has lagged behind because lethal or sterile mutations pose a bottleneck for high-throughput approaches, and a systematic way to analyse genetic interactions of essential genes in multicellular organisms has been lacking. RESULTS: In C. elegans, non-conditional lethal mutations can be maintained in heterozygosity using chromosome balancers, commonly expressing green fluorescent protein (GFP) in the pharynx. However, gene expression or function is typically monitored by the use of fluorescent reporters marked with the same fluorophore, presenting a challenge to sort worm populations of interest, particularly at early larval stages. Here, we develop a sorting strategy capable of selecting homozygous mutants carrying a GFP stress reporter from GFP-balanced animals at the second larval stage. Because sorting is not completely error-free, we develop an automated high-throughput image analysis protocol that identifies and discards animals carrying the chromosome balancer. We demonstrate the experimental usefulness of combining sorting of homozygous lethal mutants and automated image analysis in a functional genomic RNA interference (RNAi) screen for genes that genetically interact with mitochondrial prohibitin (PHB). Lack of PHB results in embryonic lethality, while homozygous PHB deletion mutants develop into sterile adults due to maternal contribution and strongly induce the mitochondrial unfolded protein response (UPRmt). In a chromosome-wide RNAi screen for C. elegans genes having human orthologues, we uncover both known and new PHB genetic interactors affecting the UPRmt and growth. CONCLUSIONS: The method presented here allows the study of balanced lethal mutations in a high-throughput manner. It can be easily adapted depending on the user's requirements and should serve as a useful resource for the C. elegans community for probing new biological aspects of essential nematode genes as well as the generation of more comprehensive genetic networks.
Assuntos
Proteínas de Caenorhabditis elegans/metabolismo , Caenorhabditis elegans/metabolismo , Citometria de Fluxo/métodos , Animais , Animais Geneticamente Modificados , Caenorhabditis elegans/genética , Proteínas de Caenorhabditis elegans/genética , Proteínas de Fluorescência Verde/genética , Proteínas de Fluorescência Verde/metabolismo , Mitocôndrias/metabolismo , Mutação , ProibitinasRESUMO
The mitochondrial prohibitin complex, composed of two proteins, PHB-1 and PHB-2, is a context-dependent modulator of longevity. Specifically, prohibitin deficiency shortens the lifespan of otherwise wild type worms, while it dramatically extends the lifespan under compromised metabolic conditions. This extremely intriguingly phenotype has been linked to alterations in mitochondrial function and in fat metabolism. However, the true function of the mitochondrial prohibitin complex remains elusive. Here, we used gas chromatography coupled to a flame ionization detector (GC/FID) and ¹H NMR spectroscopy to gain molecular insights into the effect of prohibitin depletion on the Caenorhabditis elegans metabolome. We analysed the effect of prohibitin deficiency in two different developmental stages and under two different conditions, which result in opposing longevity phenotypes, namely wild type worms and daf-2(e1370) insulin signalling deficient mutants. Prohibitin depletion was shown to alter the fatty acid (GC/FID) and ¹H NMR metabolic profiles of wild type animals both at the fourth larval stage of development (L4) and at the young adult (YA) stage, while being more pronounced at the later stage. Furthermore, wild type and the diapause mutant daf-2(e1370), either expressing or not prohibitin, were clearly distinguishable based on their metabolic profiles, revealing changes in fatty acid composition, as well as in carbohydrate and amino acid metabolism. Moreover, the metabolic data indicate that daf-2(e1370) mutants are more robust than the wild type animals to changes induced by prohibitin depletion. The impact of prohibitin depletion on the C. elegans metabolome will be discussed herein in the scope of its effect on longevity. This article is part of a Special Issue entitled: Mitochondrial Dysfunction in Aging. Guest Editor: Aleksandra Trifunovic.
Assuntos
Caenorhabditis elegans/metabolismo , Longevidade , Metaboloma , Mitocôndrias/fisiologia , Proteínas Repressoras/fisiologia , Animais , Proteínas de Caenorhabditis elegans , Ácidos Graxos/análise , Espectroscopia de Ressonância Magnética , ProibitinasRESUMO
Marked alterations in cellular energy metabolism are a universal hallmark of the ageing process. The biogenesis and function of mitochondria, the energy-generating organelles in eukaryotic cells, are primary longevity determinants. Genetic or pharmacological manipulations of mitochondrial activity profoundly affect the lifespan of diverse organisms. However, the molecular mechanisms regulating mitochondrial biogenesis and energy metabolism during ageing are poorly understood. Prohibitins are ubiquitous, evolutionarily conserved proteins, which form a ring-like, high-molecular-mass complex at the inner membrane of mitochondria. Here, we show that the mitochondrial prohibitin complex promotes longevity by modulating mitochondrial function and fat metabolism in the nematode Caenorhabditis elegans. We found that prohibitin deficiency shortens the lifespan of otherwise wild-type animals. Notably, knockdown of prohibitin promotes longevity in diapause mutants or under conditions of dietary restriction. In addition, prohibitin deficiency extends the lifespan of animals with compromised mitochondrial function or fat metabolism. Depletion of prohibitin influences ATP levels, animal fat content and mitochondrial proliferation in a genetic-background- and age-specific manner. Together, these findings reveal a novel mechanism regulating mitochondrial biogenesis and function, with opposing effects on energy metabolism, fat utilization and ageing in C. elegans. Prohibitin may have a similar key role in modulating energy metabolism during ageing in mammals.
Assuntos
Envelhecimento/fisiologia , Caenorhabditis elegans/crescimento & desenvolvimento , Caenorhabditis elegans/metabolismo , Mitocôndrias/metabolismo , Proteínas Repressoras/metabolismo , Transdução de Sinais , Trifosfato de Adenosina/metabolismo , Animais , Caenorhabditis elegans/genética , Caenorhabditis elegans/fisiologia , Proteínas de Caenorhabditis elegans , Restrição Calórica , Metabolismo Energético/genética , Metabolismo Energético/fisiologia , Técnicas de Silenciamento de Genes , Mucosa Intestinal/metabolismo , Metabolismo dos Lipídeos/genética , Metabolismo dos Lipídeos/fisiologia , Longevidade/genética , Longevidade/fisiologia , Mitocôndrias/fisiologia , Mutação/genética , Proibitinas , Proteínas Repressoras/genéticaRESUMO
Mitochondrial function relies on the coordinated transcription of mitochondrial and nuclear genomes to assemble respiratory chain complexes. Across species, the SIN3 coregulator influences mitochondrial functions, but how its loss impacts mitochondrial homeostasis and metabolism in the context of a whole organism is unknown. Exploring this link is important because SIN3 haploinsufficiency causes intellectual disability/autism syndromes and SIN3 plays a role in tumor biology. Here we show that loss of C. elegans SIN-3 results in transcriptional deregulation of mitochondrial- and nuclear-encoded mitochondrial genes, potentially leading to mito-nuclear imbalance. Consistent with impaired mitochondrial function, sin-3 mutants show extensive mitochondrial fragmentation by transmission electron microscopy (TEM) and in vivo imaging, and altered oxygen consumption. Metabolomic analysis of sin-3 mutant animals revealed a mitochondria stress signature and deregulation of methionine flux, resulting in decreased S-adenosyl methionine (SAM) and increased polyamine levels. Our results identify SIN3 as a key regulator of mitochondrial dynamics and metabolic flux, with important implications for human pathologies.
RESUMO
Heterochromatin is characterized by an enrichment of repetitive elements and low gene density and is often maintained in a repressed state across cell division and differentiation. The silencing is mainly regulated by repressive histone marks such as H3K9 and H3K27 methylated forms and the heterochromatin protein 1 (HP1) family. Here, we analyzed in a tissue-specific manner the binding profile of the two HP1 homologs in Caenorhabditis elegans, HPL-1 and HPL-2, at the L4 developmental stage. We identified the genome-wide binding profile of intestinal and hypodermal HPL-2 and intestinal HPL-1 and compared them with heterochromatin marks and other features. HPL-2 associated preferentially to the distal arms of autosomes and correlated positively with the methylated forms of H3K9 and H3K27. HPL-1 was also enriched in regions containing H3K9me3 and H3K27me3 but exhibited a more even distribution between autosome arms and centers. HPL-2 showed a differential tissue-specific enrichment for repetitive elements conversely with HPL-1, which exhibited a poor association. Finally, we found a significant intersection of genomic regions bound by the BLMP-1/PRDM1 transcription factor and intestinal HPL-1, suggesting a corepressive role during cell differentiation. Our study uncovers both shared and singular properties of conserved HP1 proteins, providing information about genomic binding preferences in relation to their role as heterochromatic markers.
Assuntos
Proteínas de Caenorhabditis elegans , Caenorhabditis elegans , Animais , Caenorhabditis elegans/genética , Caenorhabditis elegans/metabolismo , Homólogo 5 da Proteína Cromobox , Heterocromatina/genética , Heterocromatina/metabolismo , Proteínas de Caenorhabditis elegans/metabolismo , Proteínas Cromossômicas não Histona/genética , Proteínas Cromossômicas não Histona/metabolismo , Regulação da Expressão GênicaRESUMO
The incidence of age-related neurodegenerative disorders has risen with the increase of life expectancy. Unfortunately, the diagnosis of such disorders is in most cases only possible when the neurodegeneration status is already advanced, and symptoms are evident. Although age-related neurodegeneration is a common phenomenon in living animals, the cellular and molecular mechanisms behind remain poorly understood. Pathways leading to neurodegeneration usually diverge from a common starting point, mitochondrial stress, which can serve as a potential target for early diagnosis and treatments. Interestingly, the evolutionarily conserved mitochondrial prohibitin (PHB) complex is a key regulator of ageing and metabolism that has been associated with neurodegenerative diseases. However, its role in neurodegeneration is still not well characterized. The PHB complex shows protective or toxic effects in different genetic and physiological contexts, while mitochondrial and cellular stress promote both up and downregulation of PHB expression. With this review we aim to shed light into the complex world of PHB's function in neurodegeneration by putting together the latest advances in neurodegeneration and mitochondrial homeostasis associated with PHB. A better understanding of the role of PHB in neurodegeneration will add knowledge to neuron deterioration during ageing and help to identify early molecular markers of mitochondrial stress. This review will deepen our understanding of age-related neurodegeneration and provide questions to be addressed, relevant to human health and to improve the life quality of the elderly.
RESUMO
Necrotic cell death is defined by distinctive morphological characteristics that are displayed by dying cells (Walker, N.I., B.V. Harmon, G.C. Gobe, and J.F. Kerr. 1988. Methods Achiev. Exp. Pathol. 13:18-54). The cellular events that transpire during necrosis to generate these necrotic traits are poorly understood. Recent studies in the nematode Caenorhabditis elegans show that cytoplasmic acidification develops during necrosis and is required for cell death (Syntichaki, P., C. Samara, and N. Tavernarakis. 2005. Curr. Biol. 15:1249-1254). However, the origin of cytoplasmic acidification remains elusive. We show that the alkalization of endosomal and lysosomal compartments ameliorates necrotic cell death triggered by diverse stimuli. In addition, mutations in genes that result in altered lysosomal biogenesis and function markedly affect neuronal necrosis. We used a genetically encoded fluorescent marker to follow lysosome fate during neurodegeneration in vivo. Strikingly, we found that lysosomes fuse and localize exclusively around a swollen nucleus. In the advanced stages of cell death, the nucleus condenses and migrates toward the periphery of the cell, whereas green fluorescent protein-labeled lysosomal membranes fade, indicating lysosomal rupture. Our findings demonstrate a prominent role for lysosomes in cellular destruction during necrotic cell death, which is likely conserved in metazoans.
Assuntos
Caenorhabditis elegans/fisiologia , Lisossomos/fisiologia , Necrose , Animais , Ácido Aspártico Endopeptidases/deficiência , Ácido Aspártico Endopeptidases/genética , Ácido Aspártico Endopeptidases/fisiologia , Sequência de Bases , Calpaína/genética , Calpaína/fisiologia , Morte Celular , Endossomos/fisiologia , Microscopia Confocal , Dados de Sequência Molecular , PlasmídeosRESUMO
The mitochondrial prohibitin (PHB) complex, composed of PHB-1 and PHB-2, is an evolutionarily conserved context-dependent modulator of longevity. This extremely intriguing phenotype has been linked to alterations in mitochondrial function and lipid metabolism. The true biochemical function of the mitochondrial PHB complex remains elusive, but it has been shown to affect membrane lipid composition. Recent work, using large-scale biochemical approaches, has highlighted a broad effect of PHB on the C. elegans metabolic network. Collectively, the biochemical data support the notion that PHB modulates, at least partially, worm longevity through the moderation of fat utilisation and energy production via the mitochondrial respiratory chain. Herein, we review, in a systematic manner, recent biochemical insights into the impact of PHB on the C. elegans metabolome.
RESUMO
Mitochondrial prohibitins (PHB) are highly conserved proteins with a peculiar effect on lifespan. While PHB depletion shortens lifespan of wild-type animals, it enhances longevity of a plethora of metabolically compromised mutants, including target of rapamycin complex 2 (TORC2) mutants sgk-1 and rict-1. Here, we show that sgk-1 mutants have impaired mitochondrial homeostasis, lipogenesis and yolk formation, plausibly due to alterations in membrane lipid and sterol homeostasis. Remarkably, all these features are suppressed by PHB depletion. Our analysis shows the requirement of SRBP1/SBP-1 for the lifespan extension of sgk-1 mutants and the further extension conferred by PHB depletion. Moreover, although the mitochondrial unfolded protein response (UPRmt ) and autophagy are induced in sgk-1 mutants and upon PHB depletion, they are dispensable for lifespan. However, the enhanced longevity caused by PHB depletion in sgk-1 mutants requires both, the UPRmt and autophagy, but not mitophagy. We hypothesize that UPRmt induction upon PHB depletion extends lifespan of sgk-1 mutants through autophagy and probably modulation of lipid metabolism.
Assuntos
Autofagia , Proteínas de Caenorhabditis elegans/genética , Longevidade/fisiologia , Mitocôndrias/fisiologia , Proibitinas/fisiologia , Proteínas Serina-Treonina Quinases/genética , Resposta a Proteínas não Dobradas , Animais , Caenorhabditis elegans/genética , Caenorhabditis elegans/metabolismo , Caenorhabditis elegans/ultraestrutura , Metabolismo dos Lipídeos/genética , Lipogênese , Lisossomos/metabolismo , Alvo Mecanístico do Complexo 2 de Rapamicina/genética , Mitocôndrias/ultraestrutura , Mitofagia , Esteróis/metabolismoRESUMO
Metabolic disorders are frequently associated with physiological changes that occur during ageing. The mitochondrial prohibitin complex (PHB) is an evolutionary conserved context-dependent modulator of longevity, which has been linked to alterations in lipid metabolism but which biochemical function remains elusive. In this work we aimed at elucidating the molecular mechanism by which depletion of mitochondrial PHB shortens the lifespan of wild type animals while it extends that of insulin signaling receptor (daf-2) mutants. A liquid chromatography coupled with mass spectrometry approach was used to characterize the worm lipidome of wild type and insulin deficient animals upon PHB depletion. Toward a mechanistic interpretation of the insights coming from this analysis, we used a combination of biochemical, microscopic, and lifespan analyses. We show that PHB depletion perturbed glycerophospholipids and glycerolipids pools differently in short- versus long-lived animals. Interestingly, PHB depletion in otherwise wild type animals induced the endoplasmic reticulum (ER) unfolded protein response (UPR), which was mitigated in daf-2 mutants. Moreover, depletion of DNJ-21, which functionally interacts with PHB in mitochondria, mimicked the effect of PHB deficiency on the UPRER and on the lifespan of wild type and insulin signaling deficient mutants. Our work shows that PHB differentially modulates lipid metabolism depending on the worm's metabolic status and provides evidences for a new link between PHB and ER homeostasis in ageing regulation.
RESUMO
Aging and fertility are two interconnected processes. From invertebrates to mammals, absence of the germline increases longevity. Here we show that loss of function of sul-2, the Caenorhabditis elegans steroid sulfatase (STS), raises the pool of sulfated steroid hormones, increases longevity and ameliorates protein aggregation diseases. This increased longevity requires factors involved in germline-mediated longevity (daf-16, daf-12, kri-1, tcer-1 and daf-36 genes) although sul-2 mutations do not affect fertility. Interestingly, sul-2 is only expressed in sensory neurons, suggesting a regulation of sulfated hormones state by environmental cues. Treatment with the specific STS inhibitor STX64, as well as with testosterone-derived sulfated hormones reproduces the longevity phenotype of sul-2 mutants. Remarkably, those treatments ameliorate protein aggregation diseases in C. elegans, and STX64 also Alzheimer's disease in a mammalian model. These results open the possibility of reallocating steroid sulfatase inhibitors or derivates for the treatment of aging and aging related diseases.
Assuntos
Doença de Alzheimer/enzimologia , Doença de Alzheimer/patologia , Proteínas de Caenorhabditis elegans/metabolismo , Caenorhabditis elegans/enzimologia , Caenorhabditis elegans/fisiologia , Longevidade , Esteril-Sulfatase/metabolismo , Sulfatases/metabolismo , Animais , Modelos Animais de Doenças , Epistasia Genética , Gônadas/metabolismo , Camundongos , Fenótipo , Células Receptoras Sensoriais/metabolismo , Esteroides/metabolismoRESUMO
In a population, chemical communication determines the response of animals to changing environmental conditions, what leads to an enhanced resistance against stressors. In response to starvation, the nematode Caenorhabditis elegans arrest post-embryonic development at the first larval stage (L1) right after hatching. As arrested L1 larvae, C. elegans become more resistant to diverse stresses, allowing them to survive for several weeks expecting to encounter more favorable conditions. L1 arrested at high densities display an enhanced resistance to starvation, dependent on soluble compounds released beyond hatching and the first day of arrest. Here, we show that this chemical communication also influences recovery after prolonged periods in L1 arrest. Animals at high density recovered faster than animals at low density. We found that the density effect on survival depends on the final effector of the insulin signaling pathway, the transcription factor DAF-16. Moreover, DAF-16 activation was higher at high density, consistent with a lower expression of the insulin-like peptide DAF-28 in the neurons. The improved recovery of animals after arrest at high density depended on soluble compounds present in the media of arrested L1s. In an effort to find the nature of these compounds, we investigated the disaccharide trehalose as putative signaling molecule, since its production is enhanced during L1 arrest and it is able to activate DAF-16. We detected the presence of trehalose in the medium of arrested L1 larvae at a low concentration. The addition of this concentration of trehalose to animals arrested at low density was enough to rescue DAF-28 production and DAF-16 activation to the levels of animals arrested at high density. However, despite activating DAF-16, trehalose was not capable of reversing survival and recovery phenotypes, suggesting the participation of additional signaling molecules. With all, here we describe a molecular mechanism underlying social communication that allows C. elegans to maintain arrested L1 larvae ready to quickly recover as soon as they encounter nutrient sources.
RESUMO
Cells can enter quiescence in adverse conditions and resume proliferation when the environment becomes favorable. Prolonged quiescence comes with a cost, reducing the subsequent speed and potential to return to proliferation. Here, we show that a similar process happens during Caenorhabditis elegans development, providing an in vivo model to study proliferative capacity after quiescence. Hatching under starvation provokes the arrest of blast cell divisions that normally take place during the first larval stage (L1). We have used a novel method to precisely quantify each stage of postembryonic development to analyze the consequences of prolonged L1 quiescence. We report that prolonged L1 quiescence delays the reactivation of blast cell divisions in C. elegans, leading to a delay in the initiation of postembryonic development. The transcription factor DAF-16/FOXO is necessary for rapid recovery after extended arrest, and this effect is independent from its role as a suppressor of cell proliferation. Instead, the activation of DAF-16 by decreased insulin signaling reduces the rate of L1 aging, increasing proliferative potential. We also show that yolk provisioning affects the proliferative potential after L1 arrest modulating the rate of L1 aging, providing a possible mechanistic link between insulin signaling and the maintenance of proliferative potential. Furthermore, variable yolk provisioning in embryos is one of the sources of interindividual variability in recovery after quiescence of genetically identical animals. Our results support the relevance of L1 arrest as an in vivo model to study stem cell-like aging and the mechanisms for maintenance of proliferation potential after quiescence.
Assuntos
Células-Tronco Adultas/metabolismo , Envelhecimento/metabolismo , Proteínas de Caenorhabditis elegans/metabolismo , Caenorhabditis elegans/genética , Fatores de Transcrição Forkhead/metabolismo , Insulina/metabolismo , Envelhecimento/genética , Animais , Caenorhabditis elegans/crescimento & desenvolvimento , Caenorhabditis elegans/metabolismo , Proteínas de Caenorhabditis elegans/genética , Divisão Celular , Proliferação de Células , Privação de Alimentos , Fatores de Transcrição Forkhead/genética , Regulação da Expressão Gênica no Desenvolvimento/genética , Regulação da Expressão Gênica no Desenvolvimento/fisiologia , Larva/crescimento & desenvolvimento , Larva/metabolismo , Mutação , Transdução de Sinais/genética , Fatores de TempoRESUMO
It has been more than 60 years since the discovery of kinetin, the first known member of a group of plant hormones called cytokinins. In this review we summarize the health-promoting activity of kinetin in animal systems, ranging from cells cultured in vitro through invertebrates to mammals. Kinetin has been shown to modulate aging, to delay age-related physiological decline and to protect against some neurodegenerative diseases. We also review studies on its mechanism of action, as well as point out gaps in our current knowledge.
Assuntos
Envelhecimento , Envelhecimento Saudável , Cinetina/uso terapêutico , Animais , Citocininas , Humanos , Cinetina/farmacologiaRESUMO
Purine homeostasis is ensured through a metabolic network widely conserved from prokaryotes to humans. Purines can either be synthesized de novo, reused, or produced by interconversion of extant metabolites using the so-called recycling pathway. Although thoroughly characterized in microorganisms, such as yeast or bacteria, little is known about regulation of the purine biosynthesis network in metazoans. In humans, several diseases are linked to purine metabolism through as yet poorly understood etiologies. Particularly, the deficiency in adenylosuccinate lyase (ADSL)-an enzyme involved both in the purine de novo and recycling pathways-causes severe muscular and neuronal symptoms. In order to address the mechanisms underlying this deficiency, we established Caenorhabditis elegans as a metazoan model organism to study purine metabolism, while focusing on ADSL. We show that the purine biosynthesis network is functionally conserved in C. elegans Moreover, adsl-1 (the gene encoding ADSL in C. elegans) is required for developmental timing, germline stem cell maintenance and muscle integrity. Importantly, these traits are not affected when solely the de novo pathway is abolished, and we present evidence that germline maintenance is linked specifically to ADSL activity in the recycling pathway. Hence, our results allow developmental and tissue specific phenotypes to be ascribed to separable steps of the purine metabolic network in an animal model.
Assuntos
Adenilossuccinato Liase/metabolismo , Proteínas de Caenorhabditis elegans/metabolismo , Células Germinativas/metabolismo , Homeostase , Músculo Esquelético/metabolismo , Purinas/metabolismo , Adenilossuccinato Liase/genética , Animais , Caenorhabditis elegans/genética , Caenorhabditis elegans/crescimento & desenvolvimento , Caenorhabditis elegans/metabolismo , Proteínas de Caenorhabditis elegans/genética , Células Germinativas/citologiaRESUMO
The eukaryotic nucleus controls most cellular processes. It is isolated from the cytoplasm by the nuclear envelope, which plays a prominent role in the structural organization of the cell, including nucleocytoplasmic communication, chromatin positioning, and gene expression. Alterations in nuclear composition and function are eminently pronounced upon stress and during premature and physiological aging. These alterations are often accompanied by epigenetic changes in histone modifications. We review, here, the role of nuclear envelope proteins and histone modifiers in the 3-dimensional organization of the genome and the implications for gene expression. In particular, we focus on the nuclear lamins and the chromatin-associated protein BAF, which are linked to Hutchinson-Gilford and Nestor-Guillermo progeria syndromes, respectively. We also discuss alterations in nuclear organization and the epigenetic landscapes during normal aging and various stress conditions, ranging from yeast to humans.
Assuntos
Envelhecimento/fisiologia , Epigênese Genética/fisiologia , Lâmina Nuclear/metabolismo , Estresse Fisiológico , Cromatina/metabolismo , Proteínas de Ligação a DNA/genética , Proteínas de Ligação a DNA/metabolismo , Código das Histonas/genética , Histonas/metabolismo , Humanos , Laminas/genética , Laminas/metabolismo , Lâmina Nuclear/patologia , Progéria/genética , Progéria/patologiaRESUMO
Iron is an essential micronutrient for all forms of life; low levels of iron cause human disease, while too much iron is toxic. Low iron levels induce reactive oxygen species (ROS) by disruption of the heme and iron-sulfur cluster-dependent electron transport chain (ETC). To identify bacterial metabolites that affect development, we screened the Keio Escherichia coli collection and uncovered 244 gene deletion mutants that slow Caenorhabditis elegans development. Several of these genes encode members of the ETC cytochrome bo oxidase complex, as well as iron importers. Surprisingly, either iron or anti-oxidant supplementation reversed the developmental delay. This suggests that low bacterial iron results in high bacterial ROS and vice versa, which causes oxidative stress in C. elegans that subsequently impairs mitochondrial function and delays development. Our data indicate that the bacterial diets of C. elegans provide precisely tailored amounts of iron to support proper development.
Assuntos
Caenorhabditis elegans/crescimento & desenvolvimento , Caenorhabditis elegans/microbiologia , Escherichia coli/fisiologia , Ferro/metabolismo , Espécies Reativas de Oxigênio/metabolismo , Animais , Antioxidantes/metabolismo , Caenorhabditis elegans/genética , Dieta , Complexo de Proteínas da Cadeia de Transporte de Elétrons/genética , Escherichia coli/genética , Deleção de Genes , Interações entre Hospedeiro e Microrganismos/fisiologia , Mitocôndrias , MutaçãoRESUMO
Mitochondrial functions are essential for life, critical for development, maintenance of stem cells, adaptation to physiological changes, responses to stress, and aging. The complexity of mitochondrial biogenesis requires coordinated nuclear and mitochondrial gene expression, owing to the need of stoichiometrically assemble the oxidative phosphorylation (OXPHOS) system for ATP production. It requires, in addition, the import of a large number of proteins from the cytosol to keep optimal mitochondrial function and metabolism. Moreover, mitochondria require lipid supply for membrane biogenesis, while it is itself essential for the synthesis of membrane lipids. To achieve mitochondrial homeostasis, multiple mechanisms of quality control have evolved to ensure that mitochondrial function meets cell, tissue, and organismal demands. Herein, we give an overview of mitochondrial mechanisms that are activated in response to stress, including mitochondrial dynamics, mitophagy and the mitochondrial unfolded protein response (UPRmt). We then discuss the role of these stress responses in aging, with particular focus on Caenorhabditis elegans. Finally, we review observations that point to the mitochondrial prohibitin (PHB) complex as a key player in mitochondrial homeostasis, being essential for mitochondrial biogenesis and degradation, and responding to mitochondrial stress. Understanding how mitochondria responds to stress and how such responses are regulated is pivotal to combat aging and disease.
RESUMO
The study of mechanisms that govern feeding behaviour and its related disorders is a matter of global health interest. The roundworm Caenorhabditis elegans is becoming a model organism of choice to study these conserved pathways. C. elegans feeding depends on the contraction of the pharynx (pumping). Thanks to the worm transparency, pumping can be directly observed under a stereoscope. Therefore, C. elegans feeding has been historically investigated by counting pharyngeal pumping or by other indirect approaches. However, those methods are short-term, time-consuming and unsuitable for independent measurements of sizable numbers of individuals. Although some particular devices and long-term methods have been lately reported, they fail in the automated, scalable and/or continuous aspects. Here we present an automated bioluminescence-based method for the analysis and continuous monitoring of worm feeding in a multi-well format. We validate the method using genetic, environmental and pharmacological modulators of pharyngeal pumping. This flexible methodology allows studying food intake at specific time-points or during longer periods of time, in single worms or in populations at any developmental stage. Additionally, changes in feeding rates in response to differential metabolic status or external environmental cues can be monitored in real time, allowing accurate kinetic measurements.