RESUMO
While a dramatic increase in obesity and related comorbidities is being witnessed, the underlying mechanisms of their spread remain unresolved. Epigenetic and other non-genetic mechanisms tend to be prominent candidates involved in the establishment and transmission of obesity and associated metabolic disorders to offspring. Here, we review recent findings addressing those candidates, in the context of maternal and paternal influences, and discuss the effectiveness of preventive measures.
RESUMO
DNA methylation plays essential roles in mammals. Of particular interest are parental methylation marks that originate from the oocyte or the sperm, and bring about mono-allelic gene expression at defined chromosomal regions. The remarkable somatic stability of these parental imprints in the pre-implantation embryo-where they resist global waves of DNA demethylation-is not fully understood despite the importance of this phenomenon. After implantation, some methylation imprints persist in the placenta only, a tissue in which many genes are imprinted. Again here, the underlying epigenetic mechanisms are not clear. Mouse studies have pinpointed the involvement of transcription factors, covalent histone modifications, and histone variants. These and other features linked to the stability of methylation imprints are instructive as concerns their conservation in humans, in which different congenital disorders are caused by perturbed parental imprints. Here, we discuss DNA and histone methylation imprints, and why unravelling maintenance mechanisms is important for understanding imprinting disorders in humans.
Assuntos
Alelos , Metilação de DNA , Regulação da Expressão Gênica no Desenvolvimento , Doenças Genéticas Inatas , Impressão Genômica , Animais , Doenças Genéticas Inatas/embriologia , Doenças Genéticas Inatas/genética , Histonas/genética , Histonas/metabolismo , Humanos , Camundongos , Processamento de Proteína Pós-TraducionalRESUMO
Alternative splicing (AS) generates remarkable regulatory and proteomic complexity in metazoans. However, the functions of most AS events are not known, and programs of regulated splicing remain to be identified. To address these challenges, we describe the Vertebrate Alternative Splicing and Transcription Database (VastDB), the largest resource of genome-wide, quantitative profiles of AS events assembled to date. VastDB provides readily accessible quantitative information on the inclusion levels and functional associations of AS events detected in RNA-seq data from diverse vertebrate cell and tissue types, as well as developmental stages. The VastDB profiles reveal extensive new intergenic and intragenic regulatory relationships among different classes of AS and previously unknown and conserved landscapes of tissue-regulated exons. Contrary to recent reports concluding that nearly all human genes express a single major isoform, VastDB provides evidence that at least 48% of multiexonic protein-coding genes express multiple splice variants that are highly regulated in a cell/tissue-specific manner, and that >18% of genes simultaneously express multiple major isoforms across diverse cell and tissue types. Isoforms encoded by the latter set of genes are generally coexpressed in the same cells and are often engaged by translating ribosomes. Moreover, they are encoded by genes that are significantly enriched in functions associated with transcriptional control, implying they may have an important and wide-ranging role in controlling cellular activities. VastDB thus provides an unprecedented resource for investigations of AS function and regulation.
Assuntos
Processamento Alternativo , Bases de Dados de Ácidos Nucleicos , Éxons , Redes Reguladoras de Genes , Isoformas de Proteínas , Animais , Galinhas , Humanos , Camundongos , Isoformas de Proteínas/biossíntese , Isoformas de Proteínas/genéticaRESUMO
Cardiac progenitors are specified early in development and progressively differentiate and mature into fully functional cardiomyocytes. This process is controlled by an extensively studied transcriptional program. However, the regulatory events coordinating the progression of such program from development to maturation are largely unknown. Here, we show that the genome organizer CTCF is essential for cardiogenesis and that it mediates genomic interactions to coordinate cardiomyocyte differentiation and maturation in the developing heart. Inactivation of Ctcf in cardiac progenitor cells and their derivatives in vivo during development caused severe cardiac defects and death at embryonic day 12.5. Genome wide expression analysis in Ctcf mutant hearts revealed that genes controlling mitochondrial function and protein production, required for cardiomyocyte maturation, were upregulated. However, mitochondria from mutant cardiomyocytes do not mature properly. In contrast, multiple development regulatory genes near predicted heart enhancers, including genes in the IrxA cluster, were downregulated in Ctcf mutants, suggesting that CTCF promotes cardiomyocyte differentiation by facilitating enhancer-promoter interactions. Accordingly, loss of CTCF disrupts gene expression and chromatin interactions as shown by chromatin conformation capture followed by deep sequencing. Furthermore, CRISPR-mediated deletion of an intergenic CTCF site within the IrxA cluster alters gene expression in the developing heart. Thus, CTCF mediates local regulatory interactions to coordinate transcriptional programs controlling transitions in morphology and function during heart development.
Assuntos
Cromatina/genética , Desenvolvimento Embrionário/genética , Ventrículos do Coração/crescimento & desenvolvimento , Coração/crescimento & desenvolvimento , Proteínas Repressoras/genética , Animais , Fator de Ligação a CCCTC , Diferenciação Celular/genética , Elementos Facilitadores Genéticos , Regulação da Expressão Gênica no Desenvolvimento , Coração/embriologia , Ventrículos do Coração/embriologia , Camundongos , Mitocôndrias/genética , Mitocôndrias/metabolismo , Organogênese/genética , Regiões Promotoras Genéticas , Ligação Proteica , Ativação Transcricional/genéticaRESUMO
RATIONALE: The evolutionary conserved Tbx3/Tbx5 gene cluster encodes T-box transcription factors that play crucial roles in the development and homeostasis of the cardiac conduction system in human and mouse. Both genes are expressed in overlapping patterns and function in strictly tissue-specific and dose-dependent manners, yet, their regulation is poorly understood. OBJECTIVE: To analyze the mechanism underlying the complex regulation of the Tbx3/Tbx5 cluster. METHODS AND RESULTS: By probing the 3-dimensional architecture of the Tbx3/Tbx5 cluster using high-resolution circular chromosome conformation capture sequencing in vivo, we found that its regulatory landscape is in a preformed conformation similar in embryonic heart, limbs, and brain. Tbx3 and its flanking gene desert form a 1 Mbp loop between CCCTC-binding factor (CTCF)-binding sites that is separated from the neighboring Tbx5 loop. However, Ctcf inactivation did not result in transcriptional regulatory interaction between Tbx3 and Tbx5. Multiple sites within the Tbx3 locus contact the promoter, including sites corresponding to regions known to contain variations in the human genome influencing conduction. We identified an atrioventricular-specific enhancer and a pan-cardiac enhancer that contact the promoter and each other and synergize to activate transcription in the atrioventricular conduction system. CONCLUSIONS: We provide a high-resolution model of the 3-dimensional structure and function of the Tbx3/Tbx5 locus and show that the locus is organized in a preformed, permissive structure. The Tbx3 locus forms a CTCF-independent autonomous regulatory domain with multiple combinatorial regulatory elements that control the precise pattern of Tbx3 in the cardiac conduction system.
Assuntos
Regulação da Expressão Gênica no Desenvolvimento , Sistema de Condução Cardíaco/metabolismo , Proteínas com Domínio T/genética , Proteínas com Domínio T/metabolismo , Região 3'-Flanqueadora , Animais , Sítios de Ligação , Proteínas Morfogenéticas Ósseas/genética , Proteínas Morfogenéticas Ósseas/metabolismo , Fator de Ligação a CCCTC , Cromossomos Artificiais Bacterianos , DNA Circular/genética , Genes Reporter , Proteínas de Fluorescência Verde/genética , Proteínas de Fluorescência Verde/metabolismo , Sistema de Condução Cardíaco/embriologia , Camundongos Endogâmicos C57BL , Camundongos Knockout , Camundongos Transgênicos , Morfogênese , Família Multigênica , Proteínas Repressoras/deficiência , Proteínas Repressoras/genética , Transdução de Sinais , Transcrição Gênica , Ativação TranscricionalRESUMO
Alzheimer's disease (AD) is an incurable neurodegenerative disorder clinically characterized by progressive cognitive impairment. A prominent pathologic hallmark in the AD brain is the abnormal accumulation of the amyloid-ß 1-42 peptide (Aß), but the exact pathways mediating Aß neurotoxicity remain enigmatic. Endoplasmic reticulum (ER) stress is induced during AD, and has been indirectly implicated as a mediator of Aß neurotoxicity. We report here that Aß activates the ER stress response factor X-box binding protein 1 (XBP1) in transgenic flies and in mammalian cultured neurons, yielding its active form, the transcription factor XBP1s. XBP1s shows neuroprotective activity in two different AD models, flies expressing Aß and mammalian cultured neurons treated with Aß oligomers. Trying to identify the mechanisms mediating XBP1s neuroprotection, we found that in PC12 cells treated with Aß oligomers, XBP1s prevents the accumulation of free calcium (Ca(2+)) in the cytosol. This protective activity can be mediated by the downregulation of a specific isoform of the ryanodine Ca(2+) channel, RyR3. In support of this observation, a mutation in the only ryanodine receptor (RyR) in flies also suppresses Aß neurotoxicity, indicating the conserved mechanisms between the two AD models. These results underscore the functional relevance of XBP1s in Aß toxicity, and uncover the potential of XBP1 and RyR as targets for AD therapeutics.
Assuntos
Peptídeos beta-Amiloides/genética , Proteínas de Ligação a DNA/genética , Proteínas de Drosophila/genética , Drosophila/genética , Retículo Endoplasmático/metabolismo , Fragmentos de Peptídeos/genética , Doença de Alzheimer/metabolismo , Peptídeos beta-Amiloides/metabolismo , Animais , Animais Geneticamente Modificados/genética , Cálcio/metabolismo , Olho/patologia , Feminino , Masculino , Neurônios/metabolismo , Neurônios/patologia , Células PC12 , Fragmentos de Peptídeos/metabolismo , Dobramento de Proteína , Splicing de RNA , RNA Interferente Pequeno/genética , RNA Interferente Pequeno/metabolismo , Ratos , Canal de Liberação de Cálcio do Receptor de Rianodina/metabolismo , TransfecçãoRESUMO
Prion diseases are incurable neurodegenerative disorders in which the normal cellular prion protein (PrP(C)) converts into a misfolded isoform (PrP(Sc)) with unique biochemical and structural properties that correlate with disease. In humans, prion disorders, such as Creutzfeldt-Jakob disease, present typically with a sporadic origin, where unknown mechanisms lead to the spontaneous misfolding and deposition of wild type PrP. To shed light on how wild-type PrP undergoes conformational changes and which are the cellular components involved in this process, we analyzed the dynamics of wild-type PrP from hamster in transgenic flies. In young flies, PrP demonstrates properties of the benign PrP(C); in older flies, PrP misfolds, acquires biochemical and structural properties of PrP(Sc), and induces spongiform degeneration of brain neurons. Aged flies accumulate insoluble PrP that resists high concentrations of denaturing agents and contains PrP(Sc)-specific conformational epitopes. In contrast to PrP(Sc) from mammals, PrP is proteinase-sensitive in flies. Thus, wild-type PrP rapidly converts in vivo into a neurotoxic, protease-sensitive isoform distinct from prototypical PrP(Sc). Next, we investigated the role of molecular chaperones in PrP misfolding in vivo. Remarkably, Hsp70 prevents the accumulation of PrP(Sc)-like conformers and protects against PrP-dependent neurodegeneration. This protective activity involves the direct interaction between Hsp70 and PrP, which may occur in active membrane microdomains such as lipid rafts, where we detected Hsp70. These results highlight the ability of wild-type PrP to spontaneously convert in vivo into a protease-sensitive isoform that is neurotoxic, supporting the idea that protease-resistant PrP(Sc) is not required for pathology. Moreover, we identify a new role for Hsp70 in the accumulation of misfolded PrP. Overall, we provide new insight into the mechanisms of spontaneous accumulation of neurotoxic PrP and uncover the potential therapeutic role of Hsp70 in treating these devastating disorders.
Assuntos
Proteínas de Choque Térmico HSP70/metabolismo , Proteínas PrPSc/química , Animais , Animais Geneticamente Modificados , Cricetinae , Drosophila/genética , Drosophila/metabolismo , Humanos , Proteínas PrPSc/metabolismo , Príons , Dobramento de Proteína , Isoformas de Proteínas/química , Isoformas de Proteínas/metabolismoRESUMO
Inductive signals across germ layers are important for the development of the endoderm in vertebrates and invertebrates (Tam, P.P., M. Kanai-Azuma, and Y. Kanai. 2003. Curr. Opin. Genet. Dev. 13:393-400; Nakagoshi, H. 2005. Dev. Growth Differ. 47:383-392). In flies, the visceral mesoderm secretes signaling molecules that diffuse into the underlying midgut endoderm, where conserved signaling cascades activate the Hox gene labial, which is important for the differentiation of copper cells (Bienz, M. 1997. Curr. Opin. Genet. Dev. 7:683-688). We present here a Drosophila melanogaster gene of the Fox family of transcription factors, FoxK, that mediates transforming growth factor beta (TGF-beta) signaling in the embryonic midgut endoderm. FoxK mutant embryos fail to generate midgut constrictions and lack Labial in the endoderm. Our observations suggest that TGF-beta signaling directly regulates FoxK through functional Smad/Mad-binding sites, whereas FoxK, in turn, regulates labial expression. We also describe a new cooperative activity of the transcription factors FoxK and Dfos/AP-1 that regulates labial expression in the midgut endoderm. This regulatory activity does not require direct labial activation by the TGF-beta effector Mad. Thus, we propose that the combined activity of the TGF-beta target genes FoxK and Dfos is critical for the direct activation of lab in the endoderm.