RESUMEN
DNA methylation is a key layer of epigenetic regulation. The deposition of methylation marks relies on the catalytic activity of DNA methyltransferases (DNMTs), and their active removal relies on the activity of ten-eleven translocation (TET) enzymes. Paradoxically, in important biological contexts these antagonistic factors are co-expressed and target overlapping genomic regions. The ensuing cyclic biochemistry of cytosine modifications gives rise to a continuous, out-of-thermal equilibrium transition through different methylation states. But what is the purpose of this intriguing turnover of DNA methylation? Recent evidence demonstrates that methylation turnover is enriched at gene distal regulatory elements, including enhancers, and can give rise to large-scale oscillatory dynamics. We discuss this phenomenon and propose that DNA methylation turnover might facilitate key lineage decisions.
Asunto(s)
ADN (Citosina-5-)-Metiltransferasas/metabolismo , Metilación de ADN , Proteínas de Unión al ADN/metabolismo , Epigénesis Genética , Oxigenasas de Función Mixta/metabolismo , Proteínas Proto-Oncogénicas/metabolismo , Linaje de la Célula , Citosina/metabolismo , ADN (Citosina-5-)-Metiltransferasas/genética , Proteínas de Unión al ADN/genética , Dioxigenasas , Humanos , Oxigenasas de Función Mixta/genética , Proteínas Proto-Oncogénicas/genéticaRESUMEN
Developmental programming links growth in early life with health status in adulthood. Although environmental factors such as maternal diet can influence the growth and adult health status of offspring, the genetic influences on this process are poorly understood. Using the mouse as a model, we identify the imprinted gene Grb10 as a mediator of nutrient supply and demand in the postnatal period. The combined actions of Grb10 expressed in the mother, controlling supply, and Grb10 expressed in the offspring, controlling demand, jointly regulate offspring growth. Furthermore, Grb10 determines the proportions of lean and fat tissue during development, thereby influencing energy homeostasis in the adult. Most strikingly, we show that the development of normal lean/fat proportions depends on the combined effects of Grb10 expressed in the mother, which has the greater effect on offspring adiposity, and Grb10 expressed in the offspring, which influences lean mass. These distinct functions of Grb10 in mother and pup act complementarily, which is consistent with a coadaptation model of imprinting evolution, a model predicted but for which there is limited experimental evidence. In addition, our findings identify Grb10 as a key genetic component of developmental programming, and highlight the need for a better understanding of mother-offspring interactions at the genetic level in predicting adult disease risk.
Asunto(s)
Tamaño Corporal/genética , Proteína Adaptadora GRB10/genética , Animales , Femenino , Proteína Adaptadora GRB10/metabolismo , Regulación del Desarrollo de la Expresión Génica , Impresión Genómica , Carioferinas/fisiología , Lactancia/genética , Ratones , Ratones Noqueados , Receptores Citoplasmáticos y Nucleares/fisiología , Factor de Transcripción STAT5/fisiología , Proteína Exportina 1RESUMEN
Cellular senescence is a stable form of cell cycle arrest with roles in many pathophysiological processes including development, tissue repair, cancer, and aging. Senescence does not represent a single entity but rather a heterogeneous phenotype that depends on the trigger and cell type of origin. Such heterogeneous features include alterations to chromatin structure and epigenetic states. New technologies are beginning to unravel the distinct mechanisms regulating chromatin structure during senescence. Here, we describe the multiple levels of chromatin organization associated with senescence: global and focal, linear, and higher order.
Asunto(s)
Envejecimiento/genética , Senescencia Celular/genética , Cromatina/genética , Heterocromatina/genética , Envejecimiento/patología , Animales , Epigénesis Genética/genética , Genoma Humano , HumanosRESUMEN
HMGA1 is an abundant non-histone chromatin protein that has been implicated in embryonic development, cancer, and cellular senescence, but its specific role remains elusive. Here, we combine functional genomics approaches with graph theory to investigate how HMGA1 genomic deposition controls high-order chromatin networks in an oncogene-induced senescence model. While the direct role of HMGA1 in gene activation has been described previously, we find little evidence to support this. Instead, we show that the heterogeneous linear distribution of HMGA1 drives a specific 3D chromatin organization. HMGA1-dense loci form highly interactive networks, similar to, but independent of, constitutive heterochromatic loci. This, coupled with the exclusion of HMGA1-poor chromatin regions, leads to coordinated gene regulation through the repositioning of genes. In the absence of HMGA1, the whole process is largely reversed, but many regulatory interactions also emerge, amplifying the inflammatory senescence-associated secretory phenotype. Such HMGA1-mediated fine-tuning of gene expression contributes to the heterogeneous nature of senescence at the single-cell level. A similar 'buffer' effect of HMGA1 on inflammatory signalling is also detected in lung cancer cells. Our study reveals a mechanism through which HMGA1 modulates chromatin compartmentalization and gene regulation in senescence and beyond.
Asunto(s)
Senescencia Celular , Cromatina , Proteína HMGA1a , Humanos , Proteína HMGA1a/metabolismo , Proteína HMGA1a/genética , Senescencia Celular/genética , Cromatina/metabolismo , Cromatina/genética , Redes Reguladoras de Genes , Regulación de la Expresión Génica , Línea Celular Tumoral , Neoplasias Pulmonares/genética , Neoplasias Pulmonares/metabolismo , Neoplasias Pulmonares/patologíaRESUMEN
Ageing is the gradual decline in organismal fitness that occurs over time leading to tissue dysfunction and disease. At the cellular level, ageing is associated with reduced function, altered gene expression and a perturbed epigenome. Recent work has demonstrated that the epigenome is already rejuvenated by the maturation phase of somatic cell reprogramming, which suggests full reprogramming is not required to reverse ageing of somatic cells. Here we have developed the first "maturation phase transient reprogramming" (MPTR) method, where reprogramming factors are selectively expressed until this rejuvenation point then withdrawn. Applying MPTR to dermal fibroblasts from middle-aged donors, we found that cells temporarily lose and then reacquire their fibroblast identity, possibly as a result of epigenetic memory at enhancers and/or persistent expression of some fibroblast genes. Excitingly, our method substantially rejuvenated multiple cellular attributes including the transcriptome, which was rejuvenated by around 30 years as measured by a novel transcriptome clock. The epigenome was rejuvenated to a similar extent, including H3K9me3 levels and the DNA methylation ageing clock. The magnitude of rejuvenation instigated by MPTR appears substantially greater than that achieved in previous transient reprogramming protocols. In addition, MPTR fibroblasts produced youthful levels of collagen proteins, and showed partial functional rejuvenation of their migration speed. Finally, our work suggests that optimal time windows exist for rejuvenating the transcriptome and the epigenome. Overall, we demonstrate that it is possible to separate rejuvenation from complete pluripotency reprogramming, which should facilitate the discovery of novel anti-ageing genes and therapies.
Asunto(s)
Células Madre Pluripotentes Inducidas , Rejuvenecimiento , Reprogramación Celular/genética , Metilación de ADN , Epigenoma , Epigenómica/métodos , Fibroblastos , Humanos , Persona de Mediana EdadRESUMEN
Senescence is a fate-determined state, accompanied by reorganization of heterochromatin. Although lineage-appropriate genes can be temporarily repressed through facultative heterochromatin, stable silencing of lineage-inappropriate genes often involves the constitutive heterochromatic mark, histone H3 lysine 9 trimethylation (H3K9me3). The fate of these heterochromatic genes during senescence is unclear. In the present study, we show that a small number of lineage-inappropriate genes, exemplified by the LCE2 skin genes, are derepressed during senescence from H3K9me3 regions in fibroblasts. DNA FISH experiments reveal that these gene loci, which are condensed at the nuclear periphery in proliferative cells, are decompacted during senescence. Decompaction of the locus is not sufficient for LCE2 expression, which requires p53 and C/EBPß signaling. NLRP3, which is predominantly expressed in macrophages from an open topologically associated domain (TAD), is also derepressed in senescent fibroblasts due to the local disruption of the H3K9me3-rich TAD that contains it. NLRP3 has been implicated in the amplification of inflammatory cytokine signaling in senescence and aging, highlighting the functional relevance of gene induction from 'permissive' H3K9me3 regions in senescent cells.
Asunto(s)
Heterocromatina , Histonas , Heterocromatina/genética , Histonas/genética , Proteína con Dominio Pirina 3 de la Familia NLR/genética , Senescencia Celular/genética , Expresión GénicaRESUMEN
Lamins are crucial proteins for nuclear functionality. Here, we provide new evidence showing that increased lamin B1 levels contribute to the pathophysiology of Huntington's disease (HD), a CAG repeat-associated neurodegenerative disorder. Through fluorescence-activated nuclear suspension imaging, we show that nucleus from striatal medium-sized spiny and CA1 hippocampal neurons display increased lamin B1 levels, in correlation with altered nuclear morphology and nucleocytoplasmic transport disruption. Moreover, ChIP-sequencing analysis shows an alteration of lamin-associated chromatin domains in hippocampal nuclei, accompanied by changes in chromatin accessibility and transcriptional dysregulation. Supporting lamin B1 alterations as a causal role in mutant huntingtin-mediated neurodegeneration, pharmacological normalization of lamin B1 levels in the hippocampus of the R6/1 mouse model of HD by betulinic acid administration restored nuclear homeostasis and prevented motor and cognitive dysfunction. Collectively, our work points increased lamin B1 levels as a new pathogenic mechanism in HD and provides a novel target for its intervention.
Asunto(s)
Enfermedad de Huntington , Animales , Cuerpo Estriado , Enfermedad de Huntington/genética , Lamina Tipo B/genética , Ratones , NeuronasRESUMEN
How the epigenetic landscape is established in development is still being elucidated. Here, we uncover developmental pluripotency associated 2 and 4 (DPPA2/4) as epigenetic priming factors that establish a permissive epigenetic landscape at a subset of developmentally important bivalent promoters characterized by low expression and poised RNA-polymerase. Differentiation assays reveal that Dppa2/4 double knockout mouse embryonic stem cells fail to exit pluripotency and differentiate efficiently. DPPA2/4 bind both H3K4me3-marked and bivalent gene promoters and associate with COMPASS- and Polycomb-bound chromatin. Comparing knockout and inducible knockdown systems, we find that acute depletion of DPPA2/4 results in rapid loss of H3K4me3 from key bivalent genes, while H3K27me3 is initially more stable but lost following extended culture. Consequently, upon DPPA2/4 depletion, these promoters gain DNA methylation and are unable to be activated upon differentiation. Our findings uncover a novel epigenetic priming mechanism at developmental promoters, poising them for future lineage-specific activation.
Asunto(s)
Dipeptidil Peptidasa 4/genética , Epigénesis Genética , Células Madre Embrionarias de Ratones/citología , Factores de Transcripción/genética , Animales , Diferenciación Celular , Línea Celular , Cromatina/genética , Cromatina/metabolismo , Metilación de ADN , Dipeptidil Peptidasa 4/metabolismo , Regulación del Desarrollo de la Expresión Génica , Técnicas de Inactivación de Genes , Histonas/genética , Histonas/metabolismo , Ratones , Células Madre Embrionarias de Ratones/metabolismo , Factores de Transcripción/metabolismoRESUMEN
Senescence is a state of stable proliferative arrest, generally accompanied by the senescence-associated secretory phenotype, which modulates tissue homeostasis. Enhancer-promoter interactions, facilitated by chromatin loops, play a key role in gene regulation but their relevance in senescence remains elusive. Here, we use Hi-C to show that oncogenic RAS-induced senescence in human diploid fibroblasts is accompanied by extensive enhancer-promoter rewiring, which is closely connected with dynamic cohesin binding to the genome. We find de novo cohesin peaks often at the 3' end of a subset of active genes. RAS-induced de novo cohesin peaks are transcription-dependent and enriched for senescence-associated genes, exemplified by IL1B, where de novo cohesin binding is involved in new loop formation. Similar IL1B induction with de novo cohesin appearance and new loop formation are observed in terminally differentiated macrophages, but not TNFα-treated cells. These results suggest that RAS-induced senescence represents a cell fate determination-like process characterised by a unique gene expression profile and 3D genome folding signature, mediated in part through cohesin redistribution on chromatin.
Asunto(s)
Proteínas de Ciclo Celular/metabolismo , Senescencia Celular/genética , Cromatina/metabolismo , Proteínas Cromosómicas no Histona/metabolismo , Transcripción Genética , Factor de Unión a CCCTC/metabolismo , Diferenciación Celular/genética , Línea Celular , Elementos de Facilitación Genéticos/genética , Regulación de la Expresión Génica , Sitios Genéticos , Genoma , Humanos , Interleucina-1/genética , Macrófagos/citología , Regiones Promotoras Genéticas , Unión Proteica/efectos de los fármacos , Factor de Necrosis Tumoral alfa/farmacología , Proteínas ras/metabolismo , CohesinasRESUMEN
The architectural protein CTCF is a mediator of chromatin conformation, but how CTCF binding to DNA is orchestrated to maintain long-range gene expression is poorly understood. Here we perform RNAi knockdown to reduce CTCF levels and reveal a shared subset of CTCF-bound sites are robustly resistant to protein depletion. The 'persistent' CTCF sites are enriched at domain boundaries and chromatin loops constitutive to all cell types. CRISPR-Cas9 deletion of 2 persistent CTCF sites at the boundary between a long-range epigenetically active (LREA) and silenced (LRES) region, within the Kallikrein (KLK) locus, results in concordant activation of all 8 KLK genes within the LRES region. CTCF genome-wide depletion results in alteration in Topologically Associating Domain (TAD) structure, including the merging of TADs, whereas TAD boundaries are not altered where persistent sites are maintained. We propose that the subset of essential CTCF sites are involved in cell-type constitutive, higher order chromatin architecture.
Asunto(s)
Factor de Unión a CCCTC/metabolismo , Cromatina/metabolismo , Epigénesis Genética , Sitios de Unión , Factor de Unión a CCCTC/genética , Cromatina/química , Cromatina/genética , ADN/genética , ADN/metabolismo , Humanos , Regiones Promotoras Genéticas , Unión Proteica , Dominios ProteicosRESUMEN
Interleukin-1 alpha (IL-1α) is a powerful cytokine that modulates immunity, and requires canonical cleavage by calpain for full activity. Mature IL-1α is produced after inflammasome activation and during cell senescence, but the protease cleaving IL-1α in these contexts is unknown. We show IL-1α is activated by caspase-5 or caspase-11 cleavage at a conserved site. Caspase-5 drives cleaved IL-1α release after human macrophage inflammasome activation, while IL-1α secretion from murine macrophages only requires caspase-11, with IL-1ß release needing caspase-11 and caspase-1. Importantly, senescent human cells require caspase-5 for the IL-1α-dependent senescence-associated secretory phenotype (SASP) in vitro, while senescent mouse hepatocytes need caspase-11 for the SASP-driven immune surveillance of senescent cells in vivo. Together, we identify IL-1α as a novel substrate of noncanonical inflammatory caspases and finally provide a mechanism for how IL-1α is activated during senescence. Thus, targeting caspase-5 may reduce inflammation and limit the deleterious effects of accumulated senescent cells during disease and Aging.
Asunto(s)
Caspasas/metabolismo , Senescencia Celular , Inflamasomas/metabolismo , Inflamación/metabolismo , Interleucina-1alfa/metabolismo , Animales , Células Cultivadas , Femenino , Células HeLa , Humanos , Interleucina-1alfa/análisis , Ratones , Ratones Endogámicos C57BLRESUMEN
Senescent cells interact with the surrounding microenvironment achieving diverse functional outcomes. We have recently identified that NOTCH1 can drive 'lateral induction' of a unique senescence phenotype in adjacent cells by specifically upregulating the NOTCH ligand JAG1. Here we show that NOTCH signalling can modulate chromatin structure autonomously and non-autonomously. In addition to senescence-associated heterochromatic foci (SAHF), oncogenic RAS-induced senescent (RIS) cells exhibit a massive increase in chromatin accessibility. NOTCH signalling suppresses SAHF and increased chromatin accessibility in this context. Strikingly, NOTCH-induced senescent cells, or cancer cells with high JAG1 expression, drive similar chromatin architectural changes in adjacent cells through cell-cell contact. Mechanistically, we show that NOTCH signalling represses the chromatin architectural protein HMGA1, an association found in multiple human cancers. Thus, HMGA1 is involved not only in SAHFs but also in RIS-driven chromatin accessibility. In conclusion, this study identifies that the JAG1-NOTCH-HMGA1 axis mediates the juxtacrine regulation of chromatin architecture.
Asunto(s)
Senescencia Celular , Receptor Notch1/metabolismo , Cromatina/genética , Cromatina/metabolismo , Proteína HMGA1a/genética , Proteína HMGA1a/metabolismo , Heterocromatina/genética , Heterocromatina/metabolismo , Humanos , Proteína Jagged-1 , Receptor Notch1/genética , Transducción de SeñalRESUMEN
G-quadruplex (G4) structural motifs have been linked to transcription, replication and genome instability and are implicated in cancer and other diseases. However, it is crucial to demonstrate the bona fide formation of G4 structures within an endogenous chromatin context. Herein we address this through the development of G4 ChIP-seq, an antibody-based G4 chromatin immunoprecipitation and high-throughput sequencing approach. We find â¼10,000 G4 structures in human chromatin, predominantly in regulatory, nucleosome-depleted regions. G4 structures are enriched in the promoters and 5' UTRs of highly transcribed genes, particularly in genes related to cancer and in somatic copy number amplifications, such as MYC. Strikingly, de novo and enhanced G4 formation are associated with increased transcriptional activity, as shown by HDAC inhibitor-induced chromatin relaxation and observed in immortalized as compared to normal cellular states. Our findings show that regulatory, nucleosome-depleted chromatin and elevated transcription shape the endogenous human G4 DNA landscape.
Asunto(s)
Cromatina/fisiología , G-Cuádruplex , Secuencias Reguladoras de Ácidos Nucleicos , Línea Celular , Cromatina/química , Inmunoprecipitación de Cromatina , Secuenciación de Nucleótidos de Alto Rendimiento , Humanos , Reacción en Cadena en Tiempo Real de la Polimerasa , Transcripción GenéticaRESUMEN
Senescence, a persistent form of cell-cycle arrest, is often associated with a diverse secretome, which provides complex functionality for senescent cells within the tissue microenvironment. We show that oncogene-induced senescence is accompanied by a dynamic fluctuation of NOTCH1 activity, which drives a TGF-ß-rich secretome, while suppressing the senescence-associated pro-inflammatory secretome through inhibition of C/EBPß. NOTCH1 and NOTCH1-driven TGF-ß contribute to 'lateral induction of senescence' through a juxtacrine NOTCH-JAG1 pathway. In addition, NOTCH1 inhibition during senescence facilitates upregulation of pro-inflammatory cytokines, promoting lymphocyte recruitment and senescence surveillance in vivo. As enforced activation of NOTCH1 signalling confers a near mutually exclusive secretory profile compared with typical senescence, our data collectively indicate that the dynamic alteration of NOTCH1 activity during senescence dictates a functional balance between these two distinct secretomes: one representing TGF-ß and the other pro-inflammatory cytokines, highlighting that NOTCH1 is a temporospatial controller of secretome composition.