RESUMEN
SNPs affecting disease risk often reside in non-coding genomic regions. Here, we show that SNPs are highly enriched at mouse strain-selective adipose tissue binding sites for PPARγ, a nuclear receptor for anti-diabetic drugs. Many such SNPs alter binding motifs for PPARγ or cooperating factors and functionally regulate nearby genes whose expression is strain selective and imbalanced in heterozygous F1 mice. Moreover, genetically determined binding of PPARγ accounts for mouse strain-specific transcriptional effects of TZD drugs, providing proof of concept for personalized medicine related to nuclear receptor genomic occupancy. In human fat, motif-altering SNPs cause differential PPARγ binding, provide a molecular mechanism for some expression quantitative trait loci, and are risk factors for dysmetabolic traits in genome-wide association studies. One PPARγ motif-altering SNP is associated with HDL levels and other metabolic syndrome parameters. Thus, natural genetic variation in PPARγ genomic occupancy determines individual disease risk and drug response.
Asunto(s)
Hipoglucemiantes/metabolismo , PPAR gamma/genética , PPAR gamma/metabolismo , Polimorfismo de Nucleótido Simple , Tejido Adiposo , Animales , Expresión Génica , Humanos , Ratones , Ratones de la Cepa 129 , Ratones Endogámicos C57BL , Factores de Transcripción/metabolismoRESUMEN
Peroxisome proliferator-activated receptor γ (PPARγ) is a nuclear receptor that is a vital regulator of adipogenesis, insulin sensitivity, and lipid metabolism. Activation of PPARγ by antidiabetic thiazolidinediones (TZD) reverses insulin resistance but also leads to weight gain that limits the use of these drugs. There are two main PPARγ isoforms, but the specific functions of each are not established. Here we generated mouse lines in which endogenous PPARγ1 and PPARγ2 were epitope-tagged to interrogate isoform-specific genomic binding, and mice deficient in either PPARγ1 or PPARγ2 to assess isoform-specific gene regulation. Strikingly, although PPARγ1 and PPARγ2 contain identical DNA binding domains, we uncovered isoform-specific genomic binding sites in addition to shared sites. Moreover, PPARγ1 and PPARγ2 regulated a different set of genes in adipose tissue depots, suggesting distinct roles in adipocyte biology. Indeed, mice with selective deficiency of PPARγ1 maintained body temperature better than wild-type or PPARγ2-deficient mice. Most remarkably, although TZD treatment improved glucose tolerance in mice lacking either PPARγ1 or PPARγ2, the PPARγ1-deficient mice were protected from TZD-induced body weight gain compared with PPARγ2-deficient mice. Thus, PPARγ isoforms have specific and separable metabolic functions that may be targeted to improve therapy for insulin resistance and diabetes.
Asunto(s)
Resistencia a la Insulina , Tiazolidinedionas , Adipocitos/metabolismo , Animales , Regulación de la Expresión Génica , Resistencia a la Insulina/genética , Ratones , PPAR gamma/genética , Isoformas de Proteínas/genética , Isoformas de Proteínas/metabolismoRESUMEN
Brown adipose tissue is a thermogenic organ that dissipates chemical energy as heat to protect animals against hypothermia and to counteract metabolic disease. However, the transcriptional mechanisms that determine the thermogenic capacity of brown adipose tissue before environmental cold are unknown. Here we show that histone deacetylase 3 (HDAC3) is required to activate brown adipose tissue enhancers to ensure thermogenic aptitude. Mice with brown adipose tissue-specific genetic ablation of HDAC3 become severely hypothermic and succumb to acute cold exposure. Uncoupling protein 1 (UCP1) is nearly absent in brown adipose tissue lacking HDAC3, and there is also marked downregulation of mitochondrial oxidative phosphorylation genes resulting in diminished mitochondrial respiration. Remarkably, although HDAC3 acts canonically as a transcriptional corepressor, it functions as a coactivator of oestrogen-related receptor α (ERRα) in brown adipose tissue. HDAC3 coactivation of ERRα is mediated by deacetylation of PGC-1α and is required for the transcription of Ucp1, Ppargc1a (encoding PGC-1α), and oxidative phosphorylation genes. Importantly, HDAC3 promotes the basal transcription of these genes independently of adrenergic stimulation. Thus, HDAC3 uniquely primes Ucp1 and the thermogenic transcriptional program to maintain a critical capacity for thermogenesis in brown adipose tissue that can be rapidly engaged upon exposure to dangerously cold temperature.
Asunto(s)
Tejido Adiposo Pardo/metabolismo , Regulación de la Expresión Génica , Histona Desacetilasas/metabolismo , Termogénesis , Animales , Respiración de la Célula , Frío , Elementos de Facilitación Genéticos/genética , Calor , Humanos , Masculino , Ratones , Mitocondrias/metabolismo , Fosforilación Oxidativa , Coactivador 1-alfa del Receptor Activado por Proliferadores de Peroxisomas gamma/genética , Coactivador 1-alfa del Receptor Activado por Proliferadores de Peroxisomas gamma/metabolismo , Receptores de Estrógenos/metabolismo , Termogénesis/genética , Proteína Desacopladora 1/genética , Proteína Desacopladora 1/metabolismo , Receptor Relacionado con Estrógeno ERRalfaRESUMEN
PR (PRD1-BF1-RIZ1 homologous) domain-containing 16 (PRDM16) drives a brown fat differentiation program, but the mechanisms by which PRDM16 activates brown fat-selective genes have been unclear. Through chromatin immunoprecipitation (ChIP) followed by deep sequencing (ChIP-seq) analyses in brown adipose tissue (BAT), we reveal that PRDM16 binding is highly enriched at a broad set of brown fat-selective genes. Importantly, we found that PRDM16 physically binds to MED1, a component of the Mediator complex, and recruits it to superenhancers at brown fat-selective genes. PRDM16 deficiency in BAT reduces MED1 binding at PRDM16 target sites and causes a fundamental change in chromatin architecture at key brown fat-selective genes. Together, these data indicate that PRDM16 controls chromatin architecture and superenhancer activity in BAT.
Asunto(s)
Tejido Adiposo Pardo/metabolismo , Cromatina/metabolismo , Proteínas de Unión al ADN/metabolismo , Subunidad 1 del Complejo Mediador/metabolismo , Factores de Transcripción/metabolismo , Transcripción Genética/fisiología , Animales , Cromatina/química , Cromatina/genética , Elementos de Facilitación Genéticos , RatonesRESUMEN
Rosiglitazone (rosi) is a powerful insulin sensitizer, but serious toxicities have curtailed its widespread clinical use. Rosi functions as a high-affinity ligand for peroxisome proliferator-activated receptor γ (PPARγ), the adipocyte-predominant nuclear receptor (NR). The classic model, involving binding of ligand to the NR on DNA, explains positive regulation of gene expression, but ligand-dependent repression is not well understood. We addressed this issue by studying the direct effects of rosi on gene transcription using global run-on sequencing (GRO-seq). Rosi-induced changes in gene body transcription were pronounced after 10 min and correlated with steady-state mRNA levels as well as with transcription at nearby enhancers (enhancer RNAs [eRNAs]). Up-regulated eRNAs occurred almost exclusively at PPARγ-binding sites, to which rosi treatment recruited coactivators, including MED1, p300, and CBP. In contrast, transcriptional repression by rosi involved a loss of coactivators from eRNA sites devoid of PPARγ and enriched for other transcription factors, including AP-1 factors and C/EBPs. Thus, rosi activates and represses transcription by fundamentally different mechanisms that could inform the future development of anti-diabetic drugs.
Asunto(s)
Adipocitos/efectos de los fármacos , Regulación de la Expresión Génica/efectos de los fármacos , Hipoglucemiantes/farmacología , PPAR gamma/metabolismo , Tiazolidinedionas/farmacología , Células 3T3-L1 , Animales , Humanos , Subunidad 1 del Complejo Mediador/metabolismo , Ratones , Unión Proteica , Rosiglitazona , TranscriptomaRESUMEN
Sequence-specific DNA binding recruits transcription factors (TFs) to the genome to regulate gene expression. Here, we perform high resolution mapping of CEBP proteins to determine how sequence dictates genomic occupancy. We demonstrate a fundamental difference between the sequence repertoire utilized by CEBPs in vivo versus the palindromic sequence preference reported by classical in vitro models, by identifying a palindromic motif at <1% of the genomic binding sites. On the native genome, CEBPs bind a diversity of related 10 bp sequences resulting from the fusion of degenerate and canonical half-sites. Altered DNA specificity of CEBPs in cells occurs through heterodimerization with other bZip TFs, and approximately 40% of CEBP-binding sites in primary human cells harbor motifs characteristic of CEBP heterodimers. In addition, we uncover an important role for sequence bias at core-motif-flanking bases for CEBPs and demonstrate that flanking bases regulate motif function across mammalian bZip TFs. Favorable flanking bases confer efficient TF occupancy and transcriptional activity, and DNA shape may explain how the flanks alter TF binding. Importantly, motif optimization within the 10-mer is strongly correlated with cell-type-independent recruitment of CEBPß, providing key insight into how sequence sub-optimization affects genomic occupancy of widely expressed CEBPs across cell types.
Asunto(s)
Proteínas Potenciadoras de Unión a CCAAT/química , Motivos de Nucleótidos , Transcripción Genética , Animales , Secuencia de Bases , Factores de Transcripción con Cremalleras de Leucina de Carácter Básico/química , Factores de Transcripción con Cremalleras de Leucina de Carácter Básico/metabolismo , Sitios de Unión , Proteínas Potenciadoras de Unión a CCAAT/metabolismo , Inmunoprecipitación de Cromatina , Dimerización , Secuenciación de Nucleótidos de Alto Rendimiento , Humanos , Ratones , Polimorfismo de Nucleótido Simple , Unión Proteica , Organismos Libres de Patógenos Específicos , Relación Estructura-Actividad , Especificidad por SustratoRESUMEN
Macrophages, a key cellular component of inflammation, become functionally polarized in a signal- and context-specific manner. Th2 cytokines such as interleukin 4 (IL-4) polarize macrophages to a state of alternative activation that limits inflammation and promotes wound healing. Alternative activation is mediated by a transcriptional program that is influenced by epigenomic modifications, including histone acetylation. Here we report that macrophages lacking histone deacetylase 3 (HDAC3) display a polarization phenotype similar to IL-4-induced alternative activation and, furthermore, are hyperresponsive to IL-4 stimulation. Throughout the macrophage genome, HDAC3 deacetylates histone tails at regulatory regions, leading to repression of many IL-4-regulated genes characteristic of alternative activation. Following exposure to Schistosoma mansoni eggs, a model of Th2 cytokine-mediated disease that is limited by alternative activation, pulmonary inflammation was ameliorated in mice lacking HDAC3 in macrophages. Thus, HDAC3 functions in alternative activation as a brake whose release could be of benefit in the treatment of multiple inflammatory diseases.
Asunto(s)
Epigénesis Genética , Histona Desacetilasas/genética , Activación de Macrófagos/genética , Macrófagos/metabolismo , Animales , Histona Desacetilasas/metabolismo , Interleucina-4/genética , Interleucina-4/metabolismo , Macrófagos/inmunología , Ratones , Ratones Endogámicos , Neumonía/enzimología , Neumonía/inmunología , Neumonía/parasitología , Schistosoma mansoni , Células Th2/inmunología , Células Th2/metabolismoRESUMEN
Glucocorticoids (GCs) are commonly prescribed drugs, but their anti-inflammatory benefits are mitigated by metabolic side effects. Their transcriptional effects, including tissue-specific gene activation and repression, are mediated by the glucocorticoid receptor (GR), which is known to bind as a homodimer to a palindromic DNA sequence. Using ChIP-exo in mouse liver under endogenous corticosterone exposure, we report here that monomeric GR interaction with a half-site motif is more prevalent than homodimer binding. Monomers colocalize with lineage-determining transcription factors in both liver and primary macrophages, and the GR half-site motif drives transcription, suggesting that monomeric binding is fundamental to GR's tissue-specific functions. In response to exogenous GC in vivo, GR dimers assemble on chromatin near ligand-activated genes, concomitant with monomer evacuation of sites near repressed genes. Thus, pharmacological GCs mediate gene expression by favoring GR homodimer occupancy at classic palindromic sites at the expense of monomeric binding. The findings have important implications for improving therapies that target GR.
Asunto(s)
Genómica/métodos , Glucocorticoides/farmacología , Receptores de Glucocorticoides/genética , Activación Transcripcional , Animales , Células Cultivadas , Cromatina/genética , Cromatina/metabolismo , Inmunoprecipitación de Cromatina , Clonación Molecular , Expresión Génica , Terapia Genética , Secuenciación de Nucleótidos de Alto Rendimiento , Hígado/efectos de los fármacos , Hígado/metabolismo , Macrófagos/efectos de los fármacos , Macrófagos/metabolismo , Masculino , Ratones , Ratones Endogámicos BALB C , Receptores de Glucocorticoides/metabolismo , Análisis de Secuencia de ARN , Factores de Transcripción/genética , Factores de Transcripción/metabolismoRESUMEN
The transcriptional mechanisms by which temporary exposure to developmental signals instigates adipocyte differentiation are unknown. During early adipogenesis, we find transient enrichment of the glucocorticoid receptor (GR), CCAAT/enhancer-binding protein beta (CEBPbeta), p300, mediator subunit 1, and histone H3 acetylation near genes involved in cell proliferation, development, and differentiation, including the gene encoding the master regulator of adipocyte differentiation, peroxisome proliferator-activated receptor gamma2 (PPARgamma2). Occupancy and enhancer function are triggered by adipogenic signals, and diminish upon their removal. GR, which is important for adipogenesis but need not be active in the mature adipocyte, functions transiently with other enhancer proteins to propagate a new program of gene expression that includes induction of PPARgamma2, thereby providing a memory of the earlier adipogenic signal. Thus, the conversion of preadipocyte to adipocyte involves the formation of an epigenomic transition state that is not observed in cells at the beginning or end of the differentiation process.
Asunto(s)
Adipogénesis/fisiología , Epigénesis Genética , Transducción de Señal , Acetilación , Animales , Proteína beta Potenciadora de Unión a CCAAT/metabolismo , Línea Celular , Histonas/metabolismo , Ratones , Receptores Activados del Proliferador del Peroxisoma/metabolismo , Receptores de Glucocorticoides/metabolismoRESUMEN
FGF21 is an atypical member of the FGF family that functions as a hormone to regulate carbohydrate and lipid metabolism. Here we demonstrate that the actions of FGF21 in mouse adipose tissue, but not in liver, are modulated by the nuclear receptor Rev-erbα, a potent transcriptional repressor. Interrogation of genes induced in the absence of Rev-erbα for Rev-erbα-binding sites identified ßKlotho, an essential coreceptor for FGF21, as a direct target gene of Rev-erbα in white adipose tissue but not liver. Rev-erbα ablation led to the robust elevated expression of ßKlotho. Consequently, the effects of FGF21 were markedly enhanced in the white adipose tissue of mice lacking Rev-erbα. A major Rev-erbα-controlled enhancer at the Klb locus was also bound by the adipocytic transcription factor peroxisome proliferator-activated receptor (PPAR) γ, which regulates its activity in the opposite direction. These findings establish Rev-erbα as a specific modulator of FGF21 signaling in adipose tissue.
Asunto(s)
Adipocitos/metabolismo , Tejido Adiposo/metabolismo , Factores de Crecimiento de Fibroblastos/metabolismo , Miembro 1 del Grupo D de la Subfamilia 1 de Receptores Nucleares/metabolismo , Transducción de Señal/fisiología , Adipocitos/citología , Tejido Adiposo/citología , Animales , Femenino , Factores de Crecimiento de Fibroblastos/genética , Proteínas Klotho , Masculino , Proteínas de la Membrana/genética , Proteínas de la Membrana/metabolismo , Ratones , Ratones Noqueados , Miembro 1 del Grupo D de la Subfamilia 1 de Receptores Nucleares/genética , PPAR gamma/genética , PPAR gamma/metabolismoRESUMEN
Chromatin influences gene expression by restricting access of DNA binding proteins to their cognate sites in the genome. Large-scale characterization of nucleosome positioning in Saccharomyces cerevisiae has revealed a stereotyped promoter organization in which a nucleosome-free region (NFR) is present within several hundred base pairs upstream of the translation start site. Many transcription factors bind within NFRs and nucleate chromatin remodelling events which then expose other cis-regulatory elements. However, it is not clear how transcription-factor binding and chromatin influence quantitative attributes of gene expression. Here we show that nucleosomes function largely to decouple the threshold of induction from dynamic range. With a series of variants of one promoter, we establish that the affinity of exposed binding sites is a primary determinant of the level of physiological stimulus necessary for substantial gene activation, and sites located within nucleosomal regions serve to scale expression once chromatin is remodelled. Furthermore, we find that the S. cerevisiae phosphate response (PHO) pathway exploits these promoter designs to tailor gene expression to different environmental phosphate levels. Our results suggest that the interplay of chromatin and binding-site affinity provides a mechanism for fine-tuning responses to the same cellular state. Moreover, these findings may be a starting point for more detailed models of eukaryotic transcriptional control.
Asunto(s)
Cromatina/genética , Cromatina/metabolismo , Regulación Fúngica de la Expresión Génica , Regiones Promotoras Genéticas/genética , Saccharomyces cerevisiae/genética , Proteínas de Unión al ADN/genética , Genes Fúngicos/genética , Genes Reporteros/genética , Modelos Genéticos , Nucleosomas/genética , Nucleosomas/metabolismo , Iniciación de la Cadena Peptídica Traduccional , Fosfatos/farmacología , Proteínas de Saccharomyces cerevisiae/genética , Factores de Transcripción/genética , Activación TranscripcionalRESUMEN
The identification of factors that define adipocyte precursor potential has important implications for obesity. Preadipocytes are fibroblastoid cells committed to becoming round lipid-laden adipocytes. In vitro, this differentiation process is facilitated by confluency, followed by adipogenic stimuli. During adipogenesis, a large number of cytostructural genes are repressed before adipocyte gene induction. Here we report that the transcriptional repressor transcription factor 7-like 1 (TCF7L1) binds and directly regulates the expression of cell structure genes. Depletion of TCF7L1 inhibits differentiation, because TCF7L1 indirectly induces the adipogenic transcription factor peroxisome proliferator-activated receptor γ in a manner that can be replaced by inhibition of myosin II activity. TCF7L1 is induced by cell contact in adipogenic cell lines, and ectopic expression of TCF7L1 alleviates the confluency requirement for adipocytic differentiation of precursor cells. In contrast, TCF7L1 is not induced during confluency of non-adipogenic fibroblasts, and, remarkably, forced expression of TCF7L1 is sufficient to commit non-adipogenic fibroblasts to an adipogenic fate. These results establish TCF7L1 as a transcriptional hub coordinating cell-cell contact with the transcriptional repression required for adipogenic competency.
Asunto(s)
Tejido Adiposo/citología , Proteína 1 Similar al Factor de Transcripción 7/fisiología , Animales , Diferenciación Celular/fisiología , Linaje de la Célula , Ratones , PPAR gamma/genéticaRESUMEN
BACKGROUND: Metabolic homeostasis in mammals critically depends on the regulation of fasting-induced genes by CREB in the liver. Previous genome-wide analysis has shown that only a small percentage of CREB target genes are induced in response to fasting-associated signaling pathways. The precise molecular mechanisms by which CREB specifically targets these genes in response to alternating hormonal cues remain to be elucidated. RESULTS: We performed chromatin immunoprecipitation coupled to high-throughput sequencing of CREB in livers from both fasted and re-fed mice. In order to quantitatively compare the extent of CREB-DNA interactions genome-wide between these two physiological conditions we developed a novel, robust analysis method, termed the 'single sample independence' (SSI) test that greatly reduced the number of false-positive peaks. We found that CREB remains constitutively bound to its target genes in the liver regardless of the metabolic state. Integration of the CREB cistrome with expression microarrays of fasted and re-fed mouse livers and ChIP-seq data for additional transcription factors revealed that the gene expression switches between the two metabolic states are associated with co-localization of additional transcription factors at CREB sites. CONCLUSIONS: Our results support a model in which CREB is constitutively bound to thousands of target genes, and combinatorial interactions between DNA-binding factors are necessary to achieve the specific transcriptional response of the liver to fasting. Furthermore, our genome-wide analysis identifies thousands of novel CREB target genes in liver, and suggests a previously unknown role for CREB in regulating ER stress genes in response to nutrient influx.
Asunto(s)
Proteína de Unión a Elemento de Respuesta al AMP Cíclico/metabolismo , Ingestión de Alimentos , Ayuno/metabolismo , Genómica , Hígado/metabolismo , Animales , Secuencia de Bases , Inmunoprecipitación de Cromatina , ADN/metabolismo , Regulación de la Expresión Génica , Masculino , Ratones , Ratones Endogámicos C57BL , Análisis de Secuencia por Matrices de Oligonucleótidos , Transcripción GenéticaRESUMEN
Adipocyte differentiation is controlled by many transcription factors, but few known downstream targets of these factors are necessary for adipogenesis. Here we report that retinol saturase (RetSat), which is an enzyme implicated in the generation of dihydroretinoid metabolites, is induced during adipogenesis and is directly regulated by the transcription factor peroxisome proliferator activated receptor gamma (PPARgamma). Ablation of RetSat dramatically inhibited adipogenesis but, surprisingly, this block was not overcome by the putative product of RetSat enzymatic activity. On the other hand, ectopic RetSat with an intact, but not a mutated, FAD/NAD dinucleotide-binding motif increased endogenous PPARgamma transcriptional activity and promoted adipogenesis. Indeed, RetSat was not required for adipogenesis when cells were provided with exogenous PPARgamma ligands. In adipose tissue, RetSat is expressed in adipocytes but is unexpectedly downregulated in obesity, most likely owing to infiltration of macrophages that we demonstrate to repress RetSat expression. Thiazolidinedione treatment reversed low RetSat expression in adipose tissue of obese mice. Thus, RetSat plays an important role in the biology of adipocytes, where it favors normal differentiation, yet is reduced in the obese state. RetSat is thus a novel target for therapeutic intervention in metabolic disease.
Asunto(s)
Adipogénesis , Regulación hacia Abajo/genética , Obesidad/enzimología , Obesidad/patología , Oxidorreductasas actuantes sobre Donantes de Grupo CH-CH/metabolismo , Células 3T3-L1 , Adipocitos/citología , Adipocitos/enzimología , Animales , Secuencia de Bases , Sitios de Unión , Proteína beta Potenciadora de Unión a CCAAT/metabolismo , Activación Enzimática , Inducción Enzimática , Femenino , Humanos , Intrones/genética , Ratones , Datos de Secuencia Molecular , Nucleótidos/metabolismo , Obesidad/genética , Oxidorreductasas actuantes sobre Donantes de Grupo CH-CH/genética , PPAR gamma/metabolismo , Elementos de Respuesta/genética , Transcripción Genética , Vitamina A/análogos & derivados , Vitamina A/metabolismoRESUMEN
The role of PPAR gamma (PPARγ) has been well characterized in the developmental process of adipogenesis, yet its aberrant expression patterns and functions in cancer subtypes are less understood. Although PPARγ has been recently demonstrated to play non-cell-autonomous roles in promoting bladder urothelial carcinoma (UC) progression, underlying mechanisms of the cell-intrinsic oncogenic activity remain unknown. Here, we report robust expression and nuclear accumulation of PPARγ in 47% of samples of patients with UC, exceeding mRNA expression patterns published by The Cancer Genome Atlas. In vitro assays revealed for the first time that treatment of UC cells with PPARγ inverse agonist or PPARG knockout by CRISPR-Cas9 reduces proliferation, migration, and invasion of multiple established UC cell lines, most strongly in those characterized by PPARG genomic amplification or activating mutations of RXRA, the obligate heterodimer of PPARγ. Through genome-wide approaches including chromatin immunoprecipitation sequencing and RNA sequencing, we define a novel set of PPARγ-regulated genes in UC, including Sonic Hedgehog (SHH). Similar to PPARγ, genetic inhibition of SHH reduces proliferation and motility. Finally, we demonstrate the PPARγ dependency of UC tumors in vivo by genetic and pharmacologic PPARγ inhibition in subcutaneous xenografts. Collectively, our data indicate that PPARγ promotes UC progression in a subset of patients, at least in part, through cell-autonomous mechanisms linked to SHH signaling. IMPLICATIONS: Genome-wide analysis of DNA-binding sites for oncogenic factor PPARγ revealed SHH as a novel downstream target involved in UC progression, providing important insight into the tumorigenic nature and molecular mechanism of PPARγ signaling in UC.
Asunto(s)
Carcinoma de Células Transicionales/metabolismo , PPAR gamma/metabolismo , Neoplasias de la Vejiga Urinaria/metabolismo , Animales , Benzamidas/farmacología , Carcinoma de Células Transicionales/tratamiento farmacológico , Carcinoma de Células Transicionales/genética , Carcinoma de Células Transicionales/patología , Línea Celular Tumoral , Xenoinjertos , Humanos , Masculino , Ratones , Ratones Desnudos , Mutación , PPAR gamma/antagonistas & inhibidores , PPAR gamma/biosíntesis , PPAR gamma/genética , Piridinas/farmacología , Transducción de Señal , Neoplasias de la Vejiga Urinaria/tratamiento farmacológico , Neoplasias de la Vejiga Urinaria/genética , Neoplasias de la Vejiga Urinaria/patologíaRESUMEN
Brown adipose tissue (BAT) generates heat to maintain body temperature and suppress obesity. Agonists for nuclear receptors PPARα and PPARγ both affect brown adipocyte function, yet the interplay between these factors in BAT is uncertain. Here, we report that PPARα shares most genomic binding sites with PPARγ, and these common binding sites are more related to BAT function than PPARγ-selective sites without PPARα. Integrating PPARα and PPARγ genomic occupancy with cold-responsive BAT transcriptomes identifies a subset of 16 genes with potential relevance to BAT function. Among these, we focused on the lysosomal protease cathepsin Z (CTSZ) and showed it is necessary for mitochondrial respiration in both mouse and human brown adipocytes. Thus, CTSZ is a shared PPARα/γ target gene in BAT and a regulator of brown adipocyte thermogenic function.
Asunto(s)
Adipocitos Marrones/metabolismo , PPAR alfa/metabolismo , PPAR gamma/metabolismo , Termogénesis/genética , Tejido Adiposo Pardo/metabolismo , Animales , Secuencia de Bases , Sitios de Unión , Catepsina Z/genética , Catepsina Z/metabolismo , Frío , Genoma , Humanos , Masculino , Ratones Endogámicos C57BL , PPAR alfa/agonistas , PPAR gamma/agonistas , Unión ProteicaRESUMEN
The identification of small molecule ligands for the peroxisome proliferator-activated receptors (PPARs) has been instrumental in elucidating their biological roles. In particular, agonists have been the focus of much of the research in the field with relatively few antagonists being described and all of those being selective for PPARalpha or PPARgamma. The comparison of these agonist and antagonist ligands in cellular and animal systems has often led to surprising results and new insights into the biology of the PPARs. The PPARbeta/delta receptor is emerging as an important regulator of energy metabolism, inflammation, and cell growth and differentiation; however, only agonist ligands have been described for this receptor thus far. Here we describe the first report of a PPARbeta/delta small molecule antagonist ligand. This antagonist ligand will be a useful tool for elucidating the biological roles of PPARbeta/delta.
Asunto(s)
Ligandos , PPAR delta/antagonistas & inhibidores , PPAR-beta/antagonistas & inhibidores , Células Cultivadas , Inmunoprecipitación de Cromatina , Relación Dosis-Respuesta a Droga , Ácidos Grasos/metabolismo , Transferencia Resonante de Energía de Fluorescencia , Regulación de la Expresión Génica/efectos de los fármacos , Humanos , Estructura Molecular , Músculo Esquelético/citología , Músculo Esquelético/efectos de los fármacos , Músculo Esquelético/metabolismo , Oxidación-Reducción/efectos de los fármacos , Sulfonas/química , Sulfonas/farmacología , Tiazoles/química , Tiazoles/farmacología , Tiofenos/química , Tiofenos/farmacologíaRESUMEN
Classically, activated transcription by nuclear receptors (NRs) is due to a ligand-induced switch from corepressor- to coactivator-bound states. However, coactivators and corepressors recognize overlapping surfaces of liganded and unliganded NRs, respectively. Here we show that, at sufficiently high concentration, the NR corepressor (NCoR) influences the activity of the liver X receptor (LXR) even in the presence of a potent full agonist that destabilizes NCoR binding. Partial agonist ligands that less effectively dissociate NCoR from LXR are even more sensitive to NCoR levels, in a target gene-selective manner. Thus, differential recruitment of NCoR is a major determinant of partial agonism and selective LXR modulation of target genes.
Asunto(s)
Proteínas de Unión al ADN/agonistas , Proteínas de Unión al ADN/metabolismo , Proteínas Nucleares/metabolismo , Receptores Citoplasmáticos y Nucleares/agonistas , Receptores Citoplasmáticos y Nucleares/metabolismo , Proteínas Represoras/metabolismo , Activación Transcripcional , Benzoatos/química , Benzoatos/metabolismo , Bencilaminas/química , Bencilaminas/metabolismo , Línea Celular , Proteínas de Unión al ADN/química , Proteínas de Unión al ADN/genética , Dimerización , Humanos , Hidrocarburos Fluorados , Ligandos , Receptores X del Hígado , Estructura Molecular , Proteínas Nucleares/genética , Co-Represor 1 de Receptor Nuclear , Receptores Nucleares Huérfanos , Interferencia de ARN , Receptores Citoplasmáticos y Nucleares/química , Receptores Citoplasmáticos y Nucleares/genética , Proteínas Represoras/genética , Receptores X Retinoide/química , Receptores X Retinoide/metabolismo , Sulfonamidas/química , Sulfonamidas/metabolismoRESUMEN
Thiazolidinedione drugs (TZDs) target the transcriptional activity of peroxisome proliferator activated receptor γ (PPARγ) to reverse insulin resistance in type 2 diabetes, but side effects limit their clinical use. Here, using human adipose stem cell-derived adipocytes, we demonstrate that SNPs were enriched at sites of patient-specific PPARγ binding, which correlated with the individual-specific effects of the TZD rosiglitazone (rosi) on gene expression. Rosi induction of ABCA1, which regulates cholesterol metabolism, was dependent upon SNP rs4743771, which modulated PPARγ binding by influencing the genomic occupancy of its cooperating factor, NFIA. Conversion of rs4743771 from the inactive SNP allele to the active one by clustered regularly interspaced short palindromic repeats (CRISPR)/Cas9-mediated editing rescued PPARγ binding and rosi induction of ABCA1 expression. Moreover, rs4743771 is a major determinant of undesired serum cholesterol increases in rosi-treated diabetics. These data highlight human genetic variation that impacts PPARγ genomic occupancy and patient responses to antidiabetic drugs, with implications for developing personalized therapies for metabolic disorders.