RESUMEN
The transcription factor MYC is overexpressed in most cancers, where it drives multiple hallmarks of cancer progression. MYC is known to promote oncogenic transcription by binding to active promoters. In addition, MYC has also been shown to invade distal enhancers when expressed at oncogenic levels, but this enhancer binding has been proposed to have low gene-regulatory potential. Here, we demonstrate that MYC directly regulates enhancer activity to promote cancer type-specific gene programs predictive of poor patient prognosis. MYC induces transcription of enhancer RNA through recruitment of RNA polymerase II (RNAPII), rather than regulating RNAPII pause-release, as is the case at promoters. This process is mediated by MYC-induced H3K9 demethylation and acetylation by GCN5, leading to enhancer-specific BRD4 recruitment through its bromodomains, which facilitates RNAPII recruitment. We propose that MYC drives prognostic cancer type-specific gene programs through induction of an enhancer-specific epigenetic switch, which can be targeted by BET and GCN5 inhibitors.
Asunto(s)
Neoplasias , Factores de Transcripción , Humanos , Factores de Transcripción/genética , Proteínas Nucleares/genética , Pronóstico , Elementos de Facilitación Genéticos/genética , ARN Polimerasa II/genética , ARN Polimerasa II/metabolismo , Epigénesis Genética , Neoplasias/genética , Proteínas Proto-Oncogénicas c-myc/genética , Proteínas Proto-Oncogénicas c-myc/metabolismo , Proteínas que Contienen Bromodominio , Proteínas de Ciclo Celular/genéticaRESUMEN
Uncoupling Proteins (UCPs) are integral ion channels residing in the inner mitochondrial membrane. UCP2 is ubiquitously expressed, while UCP3 is found primarily in muscles and adipose tissue. Although the exact molecular mechanism of action is controversial, it is generally agreed that both homologues function to facilitate mitochondrial fatty acid oxidation. UCP2 and -3 expression is activated by the peroxisome proliferator-activated receptors (PPARs), but so far no PPAR response element has been reported in the vicinity of the Ucp2 and Ucp3 genes. Using genome-wide profiling of PPARgamma occupancy in 3T3-L1 adipocytes we demonstrate that PPARgamma associates with three chromosomal regions in the vicinity of the Ucp3 locus and weakly with a site in intron 1 of the Ucp2 gene. These sites are isolated from the nearest neighboring sites by >900 kb. The most prominent PPARgamma binding site in the Ucp2 and Ucp3 loci is located in intron 1 of the Ucp3 gene and is the only site that facilitates PPARgamma transactivation of a heterologous promoter. This site furthermore transactivates the endogenous Ucp3 promoter, and using chromatin conformation capture we show that it loops out to specifically interact with the Ucp2 promoter and intron 1. Our data indicate that PPARgamma transactivation of both UCP2 and -3 is mediated through this novel enhancer in Ucp3 intron 1.
Asunto(s)
Adipocitos/metabolismo , Elementos de Facilitación Genéticos , Intrones , Canales Iónicos/metabolismo , Proteínas Mitocondriales/metabolismo , PPAR gamma/genética , Activación Transcripcional , Células 3T3-L1 , Adipocitos/citología , Animales , Cromatina/química , Humanos , Ratones , PPAR gamma/metabolismo , Regiones Promotoras Genéticas , Unión Proteica , Retroviridae/genética , Proteína Desacopladora 2 , Proteína Desacopladora 3RESUMEN
Adipocyte differentiation is driven by waves of transcriptional regulators that reprogram the enhancer landscape and change the wiring of the promoter interactome. Here, we use high-throughput chromosome conformation enhancer capture to interrogate the role of enhancer-to-enhancer interactions during differentiation of human mesenchymal stem cells. We find that enhancers form an elaborate network that is dynamic during differentiation and coupled with changes in enhancer activity. Transcription factors (TFs) at baited enhancers amplify TF binding at target enhancers, a phenomenon we term cross-interaction stabilization of TFs. Moreover, highly interconnected enhancers (HICE) act as integration hubs orchestrating differentiation by the formation of three-dimensional enhancer communities, inside which, HICE, and other enhancers, converge on phenotypically important gene promoters. Collectively, these results indicate that enhancer interactions play a key role in the regulation of enhancer function, and that HICE are important for both signal integration and compartmentalization of the genome.
Asunto(s)
Linaje de la Célula/genética , Elementos de Facilitación Genéticos , Células Madre Mesenquimatosas/citología , Adipocitos/citología , Adipogénesis/genética , Células Cultivadas , Redes Reguladoras de Genes , Humanos , Osteoblastos/citología , Osteogénesis/genética , Factores de Transcripción/metabolismoRESUMEN
The Mediator subunit MED1/TRAP220/DRIP205/PBP interacts directly with many nuclear receptors and was long thought to be responsible for tethering Mediator to peroxisome proliferator-activated receptor (PPAR)-responsive promoters. However, it was demonstrated recently that PPARgamma can recruit Mediator by MED1-independent mechanisms. Here, we show that target gene activation by ectopically expressed PPARgamma and PPARalpha is independent of MED1. Consistent with this finding, recruitment of PPARgamma, MED6, MED8, TATA box-binding protein (TBP), and RNA polymerase II (RNAPII) to the enhancer and proximal promoter of the PPARgamma target gene Fabp4 is also independent of MED1. Using a small interfering RNA (siRNA)-based approach, we identify MED14 as a novel critical Mediator component for PPARgamma-dependent transactivation, and we demonstrate that MED14 interacts directly with the N terminus of PPARgamma in a ligand-independent manner. Interestingly, MED14 knockdown does not affect the recruitment of PPARgamma, MED6, and MED8 to the Fabp4 enhancer but does reduce their occupancy of the Fabp4 proximal promoter. In agreement with the necessity of MED14 for PPARgamma transcriptional activity, we show that knockdown of MED14 impairs adipogenesis of 3T3-L1 cells. Thus, MED14 constitutes a novel anchoring point between Mediator and the N-terminal domain of PPARgamma that is necessary for functional PPARgamma-mediated recruitment of Mediator and transactivation of PPARgamma subtype-specific target genes.