RESUMEN
The eukaryotic epigenome has an instrumental role in determining and maintaining cell identity and function. Epigenetic components such as DNA methylation, histone tail modifications, chromatin accessibility, and DNA architecture are tightly correlated with central cellular processes, while their dysregulation manifests in aberrant gene expression and disease. The ability to specifically edit the epigenome holds the promise of enhancing understanding of how epigenetic modifications function and enabling manipulation of cell phenotype for research or therapeutic purposes. Genome engineering technologies use highly specific DNA-targeting tools to precisely deposit epigenetic changes in a locus-specific manner, creating diverse epigenome editing platforms. This review summarizes these technologies and insights from recent studies, describes the complex relationship between epigenetic components and gene regulation, and highlights caveats and promises of the emerging field of epigenome editing, including applications for translational purposes, such as epigenetic therapy and regenerative medicine.
Asunto(s)
Epigenómica , Genoma , Sistemas CRISPR-Cas , Metilación de ADN , Edición Génica , Regulación de la Expresión Génica , Predisposición Genética a la Enfermedad , Código de Histonas , Humanos , Procesamiento Proteico-PostraduccionalRESUMEN
Induced pluripotent stem cells (iPSCs) can be derived from somatic cells by ectopic expression of different transcription factors, classically Oct4 (also known as Pou5f1), Sox2, Klf4 and Myc (abbreviated as OSKM). This process is accompanied by genome-wide epigenetic changes, but how these chromatin modifications are biochemically determined requires further investigation. Here we show in mice and humans that the histone H3 methylated Lys 27 (H3K27) demethylase Utx (also known as Kdm6a) regulates the efficient induction, rather than maintenance, of pluripotency. Murine embryonic stem cells lacking Utx can execute lineage commitment and contribute to adult chimaeric animals; however, somatic cells lacking Utx fail to robustly reprogram back to the ground state of pluripotency. Utx directly partners with OSK reprogramming factors and uses its histone demethylase catalytic activity to facilitate iPSC formation. Genomic analysis indicates that Utx depletion results in aberrant dynamics of H3K27me3 repressive chromatin demethylation in somatic cells undergoing reprogramming. The latter directly hampers the derepression of potent pluripotency promoting gene modules (including Sall1, Sall4 and Utf1), which can cooperatively substitute for exogenous OSK supplementation in iPSC formation. Remarkably, Utx safeguards the timely execution of H3K27me3 demethylation observed in embryonic day 10.5-11 primordial germ cells (PGCs), and Utx-deficient PGCs show cell-autonomous aberrant epigenetic reprogramming dynamics during their embryonic maturation in vivo. Subsequently, this disrupts PGC development by embryonic day 12.5, and leads to diminished germline transmission in mouse chimaeras generated from Utx-knockout pluripotent cells. Thus, we identify Utx as a novel mediator with distinct functions during the re-establishment of pluripotency and germ cell development. Furthermore, our findings highlight the principle that molecular regulators mediating loss of repressive chromatin during in vivo germ cell reprogramming can be co-opted during in vitro reprogramming towards ground state pluripotency.
Asunto(s)
Reprogramación Celular/genética , Reprogramación Celular/fisiología , Células Madre Embrionarias/metabolismo , Epigénesis Genética , Células Germinativas/metabolismo , Histona Demetilasas/metabolismo , Proteínas Nucleares/metabolismo , Alelos , Animales , Biocatálisis , Linaje de la Célula , Quimera , Células Madre Embrionarias/citología , Células Madre Embrionarias/enzimología , Femenino , Fibroblastos , Técnicas de Silenciamiento del Gen , Células Germinativas/enzimología , Células HEK293 , Histona Demetilasas/deficiencia , Histona Demetilasas/genética , Humanos , Células Madre Pluripotentes Inducidas/citología , Células Madre Pluripotentes Inducidas/enzimología , Células Madre Pluripotentes Inducidas/metabolismo , Factor 4 Similar a Kruppel , Masculino , Ratones , Ratones Noqueados , Proteínas Nucleares/deficiencia , Proteínas Nucleares/genética , Transgenes/genéticaRESUMEN
Neuronal activity-inducible gene transcription correlates with rapid and transient increases in histone acetylation at promoters and enhancers of activity-regulated genes. Exactly how histone acetylation modulates transcription of these genes has remained unknown. We used single-cell in situ transcriptional analysis to show that Fos and Npas4 are transcribed in stochastic bursts in mouse neurons and that membrane depolarization increases mRNA expression by increasing burst frequency. We then expressed dCas9-p300 or dCas9-HDAC8 fusion proteins to mimic or block activity-induced histone acetylation locally at enhancers. Adding histone acetylation increased Fos transcription by prolonging burst duration and resulted in higher Fos protein levels and an elevation of resting membrane potential. Inhibiting histone acetylation reduced Fos transcription by reducing burst frequency and impaired experience-dependent Fos protein induction in the hippocampus in vivo. Thus, activity-inducible histone acetylation tunes the transcriptional dynamics of experience-regulated genes to affect selective changes in neuronal gene expression and cellular function.