Poly(ADP-Ribosyl)ation affects stabilization of CHE-1 protein in response to DNA damage

Post-translation modifications play a crucial role in coordinating the cellular response to DNA damage. Double strand DNA breaks (DSBs) trigger the activation of ATM and Chk2 kinases, which represent the primary transducers in the signalling cascade. Among the high number of phosphorylated proteins, our attention was focused on Che-1, a novel ATM and Chk2 substrate whose role in DNA damage response has been recently shown. Phosphorylated Che-1 accumulates and promotes transcription of p53 and p53-responsive genes, which are critical for the maintenance of G2 arrest and for DNA repair processes . Poly(ADP-ribosyl)ation is a post-translational modification that shows an emerging role in the signal transduction to the DDR machinery. Poly(ADP-ribose) polymerase 1 (PARP-1), the main enzyme involved in this modification, is recruited on DNA lesions and catalyzes the synthesis of poly(ADP-ribose) polymers (PAR) on itself and on target proteins. In particular, a recent work demonstrated that PAR synthesis at DSBs sites is necessary to recruit ATM kinase, which can interact non-covalently with PAR. In this study we showed that poly(ADP-ribosyl)ation, beyond phosphorylation, is involved in the regulation of Che-1 stabilization following DNA damage. We demonstrated that Che-1 accumulation upon doxorubicin treatment is reduced after inhibition of PARP activity in HCT116 cells and in PARP-1 knock-out or silenced cells. In accordance, impairment in Che-1 accumulation by PARP inhibition reduced Che-1 occupancy at p21 promoter and affected the expression of the corresponding gene. Epistasis experiments showed that the effect of poly(ADP-ribosyl)ation on Che-1 stabilization is independent from ATM kinase activity. Indeed we demonstrated that Che-1 protein co-immunoprecipitates with ADP-ribose polymers and that PARP-1 directly interacts with Che-1, promoting its modification in vitro and in vivo. Altogether, these findings suggest that poly(ADP-ribosyl)ation of Che-1 represents a mechanism enabling the precise control over the level of Che-1 protein in response to DNA damage

A Meta-Analysis of Brain DNA Methylation Across Sex, Age, and Alzheimer\u27s Disease Points for Accelerated Epigenetic Aging in Neurodegeneration

Alzheimer\u27s disease (AD) is characterized by specific alterations of brain DNA methylation (DNAm) patterns. Age and sex, two major risk factors for AD, are also known to largely affect the epigenetic profiles in brain, but their contribution to AD-associated DNAm changes has been poorly investigated. In this study we considered publicly available DNAm datasets of four brain regions (temporal, frontal, entorhinal cortex, and cerebellum) from healthy adult subjects and AD patients, and performed a meta-analysis to identify sex-, age-, and AD-associated epigenetic profiles. In one of these datasets it was also possible to distinguish 5-methylcytosine (5mC) and 5-hydroxymethylcytosine (5hmC) profiles. We showed that DNAm differences between males and females tend to be shared between the four brain regions, while aging differently affects cortical regions compared to cerebellum. We found that the proportion of sex-dependent probes whose methylation is modified also during aging is higher than expected, but that differences between males and females tend to be maintained, with only a few probes showing age-by-sex interaction. We did not find significant overlaps between AD- and sex-associated probes, nor disease-by-sex interaction effects. On the contrary, we found that AD-related epigenetic modifications are significantly enriched in probes whose DNAm varies with age and that there is a high concordance between the direction of changes (hyper or hypo-methylation) in aging and AD, supporting accelerated epigenetic aging in the disease. In summary, our results suggest that age-associated DNAm patterns concur to the epigenetic deregulation observed in AD, providing new insights on how advanced age enables neurodegeneration

A meta-analysis of brain DNA methylation across sex, age and Alzheimer’s disease points for accelerated epigenetic aging in neurodegeneration [preprint]

Alzheimer’s disease (AD) is characterized by specific alterations of brain DNA methylation (DNAm) patterns. Age and sex, two major risk factors for AD, are also known to largely affect the epigenetic profiles in the brain, but their contribution to AD-associated DNAm changes has been poorly investigated. In this study we considered publicly available DNAm datasets of 4 brain regions (temporal, frontal, entorhinal cortex and cerebellum) from healthy adult subjects and AD patients, and performed a meta-analysis to identify sex-, age- and AD-associated epigenetic profiles. We showed that DNAm differences between males and females tend to be shared between the 4 brain regions, while aging differently affects cortical regions compared to cerebellum. We found that the proportion of sex-dependent probes whose methylation changes also during aging is higher than expected, but that differences between males and females tend to be maintained, with only few probes showing sex-by-age interaction. We did not find significant overlaps between AD- and sex-associated probes, nor disease-by-sex interaction effects. On the contrary, we found that AD-related epigenetic modifications are significantly enriched in probes whose DNAm changes with age and that there is a high concordance between the direction of changes (hyper or hypo-methylation) in aging and AD, supporting accelerated epigenetic aging in the disease. In conclusion, we demonstrated that age-associated, but not sex-associated DNAm concurs to the epigenetic deregulation observed in AD, providing new insight on how advanced age enables neurodegeneration

Down syndrome, accelerated aging and immunosenescence

Down syndrome is the most common chromosomal disorder, associated with moderate to severe intellectual disability. While life expectancy of Down syndrome population has greatly increased over the last decades, mortality rates are still high and subjects are facing prematurely a phenomenon of atypical and accelerated aging. The presence of an immune impairment in Down syndrome subjects is suggested for a long time by the existence of an increased incidence of infections, the incomplete efficacy of vaccinations, and a high prevalence of autoimmunity. Immunologic abnormalities have been described since many years in this population, both from a numerical and a functional points of view, and these abnormalities can mirror the ones observed during normal aging. In this review, we summarize our knowledge on immunologic disturbances commonly observed in subjects with Down syndrome, and in innate and adaptive immunity, as well as regarding chronic inflammation. We then discuss the role of accelerated aging in these observed abnormalities and finally review the potential age-associated molecular and cellular mechanisms involved

Poly(ADP-ribosyl)ation is involved in the epigenetic control of TET1 gene transcription

TET enzymes are the epigenetic factors involved in the formation of the Sixth DNA base 5-hydroxymethylcytosine, whose deregulation has been associated with tumorigenesis. In particular, TET1 acts as tumor suppressor preventing cell proliferation and tumor metastasis and it has frequently been found down-regulated in cancer. Thus, considering the importance of a tight control of TET1 expression, the epigenetic mechanisms involved in the transcriptional regulation of TET1 gene are here investigated. The involvement of poly(ADP-ribosyl)ation in the control of DNA and histone methylation on TET1 gene was examined. PARP activity is able to positively regulate TET1 expression maintaining a permissive chromatin state characterized by DNA hypomethylation of TET1 CpG island as well as high levels of H3K4 trimethylation. These epigenetic modifications were affected by PAR depletion causing TET1 downregulation and in turn reduced recruitment of TET1 protein on HOXA9 target gene. In conclusion, this work shows that PARP activity is a transcriptional regulator of TET1 gene through the control of epigenetic events and it suggests that deregulation of these mechanisms could account for TET1 repression in cancer

Age-Related Epigenetic Derangement upon Reprogramming and Differentiation of Cells from the Elderly

Aging is a complex multi-layered phenomenon. The study of aging in humans is based on the use of biological material from hard-to-gather tissues and highly specific cohorts. The introduction of cell reprogramming techniques posed promising features for medical practice and basic research. Recently, a growing number of studies have been describing the generation of induced pluripotent stem cells (iPSCs) from old or centenarian biologic material. Nonetheless, Reprogramming techniques determine a profound remodelling on cell epigenetic architecture whose extent is still largely debated. Given that cell epigenetic profile changes with age, the study of cell-fate manipulation approaches on cells deriving from old donors or centenarians may provide new insights not only on regenerative features and physiology of these cells, but also on reprogramming-associated and age-related epigenetic derangement

Disentangling age-dependent DNA methylation: deterministic, stochastic, and nonlinear

DNA methylation variability arises due to concurrent genetic and environmental influences. Each of them is a mixture of regular and noisy sources, whose relative contribution has not been satisfactorily understood yet. We conduct a systematic assessment of the age-dependent methylation by the signal-to-noise ratio and identify a wealth of "deterministic" CpG probes (about 90%), whose methylation variability likely originates due to genetic and general environmental factors. The remaining 10% of "stochastic" CpG probes are arguably governed by the biological noise or incidental environmental factors. Investigating the mathematical functional relationship between methylation levels and variability, we find that in about 90% of the age-associated differentially methylated positions, the variability changes as the square of the methylation level, whereas in the most of the remaining cases the dependence is linear. Furthermore, we demonstrate that the methylation level itself in more than 15% cases varies nonlinearly with age (according to the power law), in contrast to the previously assumed linear changes. Our findings present ample evidence of the ubiquity of strong DNA methylation regulation, resulting in the individual age-dependent and nonlinear methylation trajectories, whose divergence explains the cross-sectional variability. It may also serve a basis for constructing novel nonlinear epigenetic clocks

Validation of suitable internal control genes for expression studies in aging.

Quantitative data from experiments of gene expression are often normalized through levels of housekeeping genes transcription by assuming that expression of these genes is highly uniform. This practice is being questioned as it becomes increasingly clear that the level of housekeeping genes expression may vary considerably in certain biological samples. To date, the validation of reference genes in aging has received little attention and suitable reference genes have not yet been defined. Our aim was to evaluate the expression stability of frequently used reference genes in human peripheral blood mononuclear cells with respect to aging. Using quantitative RT-PCR, we carried out an extensive evaluation of five housekeeping genes, i.e. 18s rRNA, ACTB, GAPDH, HPRT1 and GUSB, for stability of expression in samples from donors in the age range 35-74 years. The consistency in the expression stability was quantified on the basis of the coefficient of variation and two algorithms termed geNorm and NormFinder. Our results indicated GUSB be the most suitable transcript and 18s the least for accurate normalization in PBMCs. We also demonstrated that aging is a confounding factor with respect to stability of 18s, HPRT1 and ACTB expression, which were particularly prone to variability in aged donors

Age-related DNA methylation changes: Potential impact on skeletal muscle aging in humans

Human aging is accompanied by a decline in muscle mass and muscle function, which is commonly referred to as sarcopenia. Sarcopenia is associated with detrimental clinical outcomes, such as a reduced quality of life, frailty, an increased risk of falls, fractures, hospitalization, and mortality. The exact underlying mechanisms of sarcopenia are poorly delineated and the molecular mechanisms driving the development and progression of this disorder remain to be uncovered. Previous studies have described age-related differences in gene expression, with one study identifying an age-specific expression signature of sarcopenia, but little is known about the influence of epigenetics, and specially of DNA methylation, in its pathogenesis. In this review, we will focus on the available knowledge in literature on the characterization of DNA methylation profiles during skeletal muscle aging and the possible impact of physical activity and nutrition. We will consider the possible use of the recently developed DNA methylation-based biomarkers of aging called epigenetic clocks in the assessment of physical performance in older individuals. Finally, we will discuss limitations and future directions of this field

Evolutionary Implications of Environmental Toxicant Exposure

Homo sapiens have been exposed to various toxins and harmful compounds that change according to various phases of human evolution. Population genetics studies showed that such exposures lead to adaptive genetic changes; while observing present exposures to different toxicants, the first molecular mechanism that confers plasticity is epigenetic remodeling and, in particular, DNA methylation variation, a molecular mechanism proposed for medium-term adaptation. A large amount of scientific literature from clinical and medical studies revealed the high impact of such exposure on human biology; thus, in this review, we examine and infer the impact that different environmental toxicants may have in shaping human evolution. We first describe how environmental toxicants shape natural human variation in terms of genetic and epigenetic diversity, and then we describe how DNA methylation may influence mutation rate and, thus, genetic variability. We describe the impact of these substances on biological fitness in terms of reproduction and survival, and in conclusion, we focus on their effect on brain evolution and physiology