Epigenetic Mechanisms of Longevity and Aging.

Aging is an inevitable outcome of life, characterized by progressive decline in tissue and organ function and increased risk of mortality. Accumulating evidence links aging to genetic and epigenetic alterations. Given the reversible nature of epigenetic mechanisms, these pathways provide promising avenues for therapeutics against age-related decline and disease. In this review, we provide a comprehensive overview of epigenetic studies from invertebrate organisms, vertebrate models, tissues, and in vitro systems. We establish links between common operative aging pathways and hallmark chromatin signatures that can be used to identify "druggable" targets to counter human aging and age-related disease.

Induced DNA demethylation can reshape chromatin topology at the IGF2-H19 locus.

Choriocarcinomas are embryonal tumours with loss of imprinting and hypermethylation at the insulin-like growth factor 2 (IGF2)-H19 locus. The DNA methyltransferase inhibitor, 5-Aza-2'deoxycytidine (5-AzaCdR) is an approved epigenetic cancer therapy. However, it is not known to what extent 5-AzaCdR influences other epigenetic marks. In this study, we set out to determine whether 5-AzaCdR treatment can reprogram the epigenomic organization of the IGF2-H19 locus in a choriocarcinoma cancer cell line (JEG3). We found that localized DNA demethylation at the H19 imprinting control region (ICR) induced by 5-AzaCdR, reduced IGF2, increased H19 expression, increased CTCF and cohesin recruitment and changed histone modifications. Furthermore chromatin accessibility was increased locus-wide and chromatin looping topography was altered such that a CTCF site downstream of the H19 enhancers switched its association with the CTCF site upstream of the IGF2 promoters to associate with the ICR. We identified a stable chromatin looping domain, which forms independently of DNA methylation. This domain contains the IGF2 gene and is marked by a histone H3 lysine 27 trimethylation block between CTCF site upstream of the IGF2 promoters and the Centrally Conserved Domain upstream of the ICR. Together, these data provide new insights into the responsiveness of chromatin topography to DNA methylation changes.

Disruption of genomic neighbourhood at the imprinted IGF2-H19 locus in Beckwith-Wiedemann syndrome and Silver-Russell syndrome.

Hyper- and hypomethylation at the IGF2-H19 imprinting control region (ICR) result in reciprocal changes in IGF2-H19 expression and the two contrasting growth disorders, Beckwith-Wiedemann syndrome (BWS) and Silver-Russell syndrome (SRS). DNA methylation of the ICR controls the reciprocal imprinting of IGF2 and H19 by preventing the binding of the insulator protein, CTCF. We here show that local changes in histone modifications and CTCF--cohesin binding at the ICR in BWS and SRS together with DNA methylation correlate with the higher order chromatin structure at the locus. In lymphoblastoid cells from control individuals, we found the repressive histone H3K9me3 and H4K20me3 marks associated with the methylated paternal ICR allele and the bivalent H3K4me2/H3K27me3 mark together with H3K9ac and CTCF--cohesin associated with the non-methylated maternal allele. In patient-derived cell lines, the mat/pat asymmetric distribution of these epigenetic marks was lost with H3K9me3 and H4K20me3 becoming biallelic in the BWS and H3K4me2, H3K27me3 and H3K9ac together with CTCF-cohesin becoming biallelic in the SRS. We further show that in BWS and SRS cells, there is opposing chromatin looping conformation mediated by CTCF--cohesin binding sites surrounding the locus. In normal cells, lack of CTCF--cohesin binding at the paternal ICR is associated with monoallelic interaction between two CTCF sites flanking the locus. CTCF--cohesin binding at the maternal ICR blocks this interaction by associating with the CTCF site downstream of the enhancers. The two alternative chromatin conformations are differently favoured in BWS and SRS likely predisposing the locus to the activation of IGF2 or H19, respectively.

Cohesin is required for higher-order chromatin conformation at the imprinted IGF2-H19 locus.

Cohesin is a chromatin-associated protein complex that mediates sister chromatid cohesion by connecting replicated DNA molecules. Cohesin also has important roles in gene regulation, but the mechanistic basis of this function is poorly understood. In mammalian genomes, cohesin co-localizes with CCCTC binding factor (CTCF), a zinc finger protein implicated in multiple gene regulatory events. At the imprinted IGF2-H19 locus, CTCF plays an important role in organizing allele-specific higher-order chromatin conformation and functions as an enhancer blocking transcriptional insulator. Here we have used chromosome conformation capture (3C) assays and RNAi-mediated depletion of cohesin to address whether cohesin affects higher order chromatin conformation at the IGF2-H19 locus in human cells. Our data show that cohesin has a critical role in maintaining CTCF-mediated chromatin conformation at the locus and that disruption of this conformation coincides with changes in IGF2 expression. We show that the cohesin-dependent, higher-order chromatin conformation of the locus exists in both G1 and G2 phases of the cell cycle and is therefore independent of cohesin's function in sister chromatid cohesion. We propose that cohesin can mediate interactions between DNA molecules in cis to insulate genes through the formation of chromatin loops, analogous to the cohesin mediated interaction with sister chromatids in trans to establish cohesion.

Somatically acquired hypomethylation of IGF2 in breast and colorectal cancer.

The imprinted insulin-like growth factor 2 (IGF2) gene is expressed predominantly from the paternal allele. Loss of imprinting (LOI) associated with hypomethylation at the promoter proximal sequence (DMR0) of the IGF2 gene was proposed as a predisposing constitutive risk biomarker for colorectal cancer. We used pyrosequencing to assess whether IGF2 DMR0 methylation is either present constitutively prior to cancer or whether it is acquired tissue-specifically after the onset of cancer. DNA samples from tumour tissues and matched non-tumour tissues from 22 breast and 42 colorectal cancer patients as well as peripheral blood samples obtained from colorectal cancer patients [SEARCH (n=case 192, controls 96)], breast cancer patients [ABC (n=case 364, controls 96)] and the European Prospective Investigation of Cancer [EPIC-Norfolk (n=breast 228, colorectal 225, controls 895)] were analysed. The EPIC samples were collected 2-5 years prior to diagnosis of breast or colorectal cancer. IGF2 DMR0 methylation levels in tumours were lower than matched non-tumour tissue. Hypomethylation of DMR0 was detected in breast (33%) and colorectal (80%) tumour tissues with a higher frequency than LOI indicating that methylation levels are a better indicator of cancer than LOI. In the EPIC population, the prevalence of IGF2 DMR0 hypomethylation was 9.5% and this correlated with increased age not cancer risk. Thus, IGF2 DMR0 hypomethylation occurs as an acquired tissue-specific somatic event rather than a constitutive innate epimutation. These results indicate that IGF2 DMR0 hypomethylation has diagnostic potential for colon cancer rather than value as a surrogate biomarker for constitutive LOI.

An integrated multi-omics approach identifies epigenetic alterations associated with Alzheimer's disease.

Protein aggregation is the hallmark of neurodegeneration, but the molecular mechanisms underlying late-onset Alzheimer's disease (AD) are unclear. Here we integrated transcriptomic, proteomic and epigenomic analyses of postmortem human brains to identify molecular pathways involved in AD. RNA sequencing analysis revealed upregulation of transcription- and chromatin-related genes, including the histone acetyltransferases for H3K27ac and H3K9ac. An unbiased proteomic screening singled out H3K27ac and H3K9ac as the main enrichments specific to AD. In turn, epigenomic profiling revealed gains in the histone H3 modifications H3K27ac and H3K9ac linked to transcription, chromatin and disease pathways in AD. Increasing genome-wide H3K27ac and H3K9ac in a fly model of AD exacerbated amyloid-ß42-driven neurodegeneration. Together, these findings suggest that AD involves a reconfiguration of the epigenome, wherein H3K27ac and H3K9ac affect disease pathways by dysregulating transcription- and chromatin-gene feedback loops. The identification of this process highlights potential epigenetic strategies for early-stage disease treatment.

Author Correction: An integrated multi-omics approach identifies epigenetic alterations associated with Alzheimer's disease.

An amendment to this paper has been published and can be accessed via a link at the top of the paper.

Epigenetic Regulation in Neurodegenerative Diseases.

Mechanisms of epigenetic regulation, including DNA methylation, chromatin remodeling, and histone post-translational modifications, are involved in multiple aspects of neuronal function and development. Recent discoveries have shed light on critical functions of chromatin in the aging brain, with an emerging realization that the maintenance of a healthy brain relies heavily on epigenetic mechanisms. Here, we present recent advances, with a focus on histone modifications and the implications for several neurodegenerative diseases including Alzheimer's disease (AD), Huntington's disease (HD), and amyotrophic lateral sclerosis (ALS). We highlight common and unique epigenetic mechanisms among these situations and point to emerging therapeutic approaches.

Publisher Correction: Dysregulation of the epigenetic landscape of normal aging in Alzheimer's disease.

In the version of this article initially published online, the fifth author's name was given as Alexander Amlie-Wolf. The correct name is Alexandre Amlie-Wolf. The error has been corrected in the print, PDF and HTML versions of this article.

Dysregulation of the epigenetic landscape of normal aging in Alzheimer's disease.

Aging is the strongest risk factor for Alzheimer's disease (AD), although the underlying mechanisms remain unclear. The chromatin state, in particular through the mark H4K16ac, has been implicated in aging and thus may play a pivotal role in age-associated neurodegeneration. Here we compare the genome-wide enrichment of H4K16ac in the lateral temporal lobe of AD individuals against both younger and elderly cognitively normal controls. We found that while normal aging leads to H4K16ac enrichment, AD entails dramatic losses of H4K16ac in the proximity of genes linked to aging and AD. Our analysis highlights the presence of three classes of AD-related changes with distinctive functional roles. Furthermore, we discovered an association between the genomic locations of significant H4K16ac changes with genetic variants identified in prior AD genome-wide association studies and with expression quantitative trait loci. Our results establish the basis for an epigenetic link between aging and AD.

TDP-43 Promotes Neurodegeneration by Impairing Chromatin Remodeling.

Regulation of chromatin structure is critical for brain development and function. However, the involvement of chromatin dynamics in neurodegeneration is less well understood. Here we find, launching from Drosophila models of amyotrophic lateral sclerosis and frontotemporal dementia, that TDP-43 impairs the induction of multiple key stress genes required to protect from disease by reducing the recruitment of the chromatin remodeler Chd1 to chromatin. Chd1 depletion robustly enhances TDP-43-mediated neurodegeneration and promotes the formation of stress granules. Conversely, upregulation of Chd1 restores nucleosomal dynamics, promotes normal induction of protective stress genes, and rescues stress sensitivity of TDP-43-expressing animals. TDP-43-mediated impairments are conserved in mammalian cells, and, importantly, the human ortholog CHD2 physically interacts with TDP-43 and is strikingly reduced in level in temporal cortex of human patient tissue. These findings indicate that TDP-43-mediated neurodegeneration causes impaired chromatin dynamics that prevents appropriate expression of protective genes through compromised function of the chromatin remodeler Chd1/CHD2. Enhancing chromatin dynamics may be a treatment approach to amyotrophic lateral scleorosis (ALS)/frontotemporal dementia (FTD).

Changes in the Transcriptome of Human Astrocytes Accompanying Oxidative Stress-Induced Senescence.

Aging is a major risk factor for many neurodegenerative disorders. A key feature of aging biology that may underlie these diseases is cellular senescence. Senescent cells accumulate in tissues with age, undergo widespread changes in gene expression, and typically demonstrate altered, pro-inflammatory profiles. Astrocyte senescence has been implicated in neurodegenerative disease, and to better understand senescence-associated changes in astrocytes, we investigated changes in their transcriptome using RNA sequencing. Senescence was induced in human fetal astrocytes by transient oxidative stress. Brain-expressed genes, including those involved in neuronal development and differentiation, were downregulated in senescent astrocytes. Remarkably, several genes indicative of astrocytic responses to injury were also downregulated, including glial fibrillary acidic protein and genes involved in the processing and presentation of antigens by major histocompatibility complex class II proteins, while pro-inflammatory genes were upregulated. Overall, our findings suggest that senescence-related changes in the function of astrocytes may impact the pathogenesis of age-related brain disorders.

Quantitative chromosome conformation capture.

It is becoming increasingly apparent that chromatin is not randomly folded into the nucleus, but instead is highly organized into specific conformations within the nucleus. One consequence of such higher order structure is that chromatin looping can bring together genomic elements which are separated by several hundreds of kilobases, such as enhancers and promoters, and functionally facilitate their interaction. The Chromosome Conformation Capture (3C) assay is a powerful technique to detect looping structures and assess the probability of interaction between distant genomic elements (1-3). Here we describe the 3C methodology, its power, and limitations, together with the controls and normalization steps required for an accurate analysis.