Distinct DNA methylomes of newborns and centenarians Heyn, Holger; Li, Ning; Ferreira, Humberto J. ...
Proceedings of the National Academy of Sciences - PNAS,
06/2012, Letnik:
109, Številka:
26
Journal Article
Recenzirano
Odprti dostop
Human aging cannot be fully understood in terms of the constrained genetic setting. Epigenetic drift is an alternative means of explaining age-associated alterations. To address this issue, we ...performed whole-genome bisulfite sequencing (WGBS) of newborn and centenarian genomes. The centenarian DNA had a lower DNA methylation content and a reduced correlation in the methylation status of neighboring cytosine—phosphate—guanine (CpGs) throughout the genome in comparison with the more homogeneously methylated newborn DNA. The more hypomethylated CpGs observed in the centenarian DNA compared with the neonate covered all genomic compartments, such as promoters, exonic, intronic, and intergenic regions. For regulatory regions, the most hypomethylated sequences in the centenarian DNA were present mainly at CpG-poor promoters and in tissue-specific genes, whereas a greater level of DNA methylation was observed in CpG island promoters. We extended the study to a larger cohort of newborn and nonagenarian samples using a 450,000 CpG-site DNA methylation microarray that reinforced the observation of more hypomethylated DNA sequences in the advanced age group. WGBS and 450,000 analyses of middle-age individuals demonstrated DNA methylomes in the crossroad between the newborn and the nonagenarian/centenarian groups. Our study constitutes a unique DNA methylation analysis of the extreme points of human life at a single-nucleotide resolution level.
Abstract
Motivation
DNA methylation is essential for normal embryogenesis and development in mammals and can be captured at single base pair resolution by whole genome bisulfite sequencing (WGBS). ...Current available analysis tools are becoming rapidly outdated as they lack sensible functionality and efficiency to handle large amounts of data now commonly created.
Results
We developed gemBS, a fast high-throughput bioinformatics pipeline specifically designed for large scale BS-Seq analysis that combines a high performance BS-mapper (GEM3) and a variant caller specifically for BS-Seq data (BScall). gemBS provides genotype information and methylation estimates for all genomic cytosines in different contexts (CpG and non-CpG) and a set of quality reports for comprehensive and reproducible analysis. gemBS is highly modular and can be easily automated, while producing robust and accurate results.
Availability and implementation
gemBS is released under the GNU GPLv3+ license. Source code and documentation are freely available from www.statgen.cat/gemBS.
Supplementary information
Supplementary data are available at Bioinformatics online.
Mammalian Y chromosomes are often neglected from genomic analysis. Due to their inherent assembly difficulties, high repeat content, and large ampliconic regions, only a handful of species have their ...Y chromosome properly characterized. To date, just a single human reference quality Y chromosome, of European ancestry, is available due to a lack of accessible methodology. To facilitate the assembly of such complicated genomic territory, we developed a novel strategy to sequence native, unamplified flow sorted DNA on a MinION nanopore sequencing device. Our approach yields a highly continuous assembly of the first human Y chromosome of African origin. It constitutes a significant improvement over comparable previous methods, increasing continuity by more than 800%. Sequencing native DNA also allows to take advantage of the nanopore signal data to detect epigenetic modifications in situ. This approach is in theory generalizable to any species simplifying the assembly of extremely large and repetitive genomes.
Here, we report DNA methylation and hydroxymethylation dynamics at nucleotide resolution using C/EBPα-enhanced reprogramming of B cells into induced pluripotent cells (iPSCs). We observed successive ...waves of hydroxymethylation at enhancers, concomitant with a decrease in DNA methylation, suggesting active demethylation. Consistent with this finding, ablation of the DNA demethylase Tet2 almost completely abolishes reprogramming. C/EBPα, Klf4, and Tfcp2l1 each interact with Tet2 and recruit the enzyme to specific DNA sites. During reprogramming, some of these sites maintain high levels of 5hmC, and enhancers and promoters of key pluripotency factors become demethylated as early as 1 day after Yamanaka factor induction. Surprisingly, methylation changes precede chromatin opening in distinct chromatin regions, including Klf4 bound sites, revealing a pioneer factor activity associated with alteration in DNA methylation. Rapid changes in hydroxymethylation similar to those in B cells were also observed during compound-accelerated reprogramming of fibroblasts into iPSCs, highlighting the generality of our observations.
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•Base-resolution profiling of DNA (hydroxy)methylation during iPSC reprogramming•Major contribution of Tet2-mediated demethylation throughout reprogramming•C/EBPα, Klf4, and Tfcp2l1 drive Tet2-mediated enhancer demethylation and activation•Klf4 induces enhancer demethylation in the absence of nucleosome repositioning
Using a highly efficient reprogramming system, Sardina et al. examined the dynamics of DNA methylation and hydroxymethylation. They found that throughout the process several transcription factors can recruit Tet2 to specific sites, leading to demethylation. Some of these sites became demethylated before chromatin opening.
An increased level of Lp(a) lipoprotein has been identified as a risk factor for coronary artery disease that is highly heritable. The genetic determinants of the Lp(a) lipoprotein level and their ...relevance for the risk of coronary disease are incompletely understood.
We used a novel gene chip containing 48,742 single-nucleotide polymorphisms (SNPs) in 2100 candidate genes to test for associations in 3145 case subjects with coronary disease and 3352 control subjects. Replication was tested in three independent populations involving 4846 additional case subjects with coronary disease and 4594 control subjects.
Three chromosomal regions (6q26-27, 9p21, and 1p13) were strongly associated with the risk of coronary disease. The LPA locus on 6q26-27 encoding Lp(a) lipoprotein had the strongest association. We identified a common variant (rs10455872) at the LPA locus with an odds ratio for coronary disease of 1.70 (95% confidence interval CI, 1.49 to 1.95) and another independent variant (rs3798220) with an odds ratio of 1.92 (95% CI, 1.48 to 2.49). Both variants were strongly associated with an increased level of Lp(a) lipoprotein, a reduced copy number in LPA (which determines the number of kringle IV-type 2 repeats), and a small Lp(a) lipoprotein size. Replication studies confirmed the effects of both variants on the Lp(a) lipoprotein level and the risk of coronary disease. A meta-analysis showed that with a genotype score involving both LPA SNPs, the odds ratios for coronary disease were 1.51 (95% CI, 1.38 to 1.66) for one variant and 2.57 (95% CI, 1.80 to 3.67) for two or more variants. After adjustment for the Lp(a) lipoprotein level, the association between the LPA genotype score and the risk of coronary disease was abolished.
We identified two LPA variants that were strongly associated with both an increased level of Lp(a) lipoprotein and an increased risk of coronary disease. Our findings provide support for a causal role of Lp(a) lipoprotein in coronary disease.
Natural killer (NK) cells are lymphocytes involved in antimicrobial and antitumoral immune responses. Using N-ethyl-N-nitrosourea mutagenesis in mice, we identified a mutant with increased resistance ...to viral infections because of the presence of hyperresponsive NK cells. Whole-genome sequencing and functional analysis revealed a loss-of-function mutation in the Ncr1 gene encoding the activating receptor NKp46. The down-regulation of NK cell activity by NKp46 was associated with the silencing of the Helios transcription factor in NK cells. NKp46 was critical for the subsequent development of antiviral and antibacterial T cell responses, which suggests that the regulation of NK cell function by NKp46 allows for the optimal development of adaptive immune responses. NKp46 blockade enhanced NK cell reactivity in vivo, which could enable the design of immunostimulation strategies in humans.
F cells measure the presence of fetal hemoglobin, a heritable quantitative trait in adults that accounts for substantial phenotypic diversity of sickle cell disease and β thalassemia. We applied a ...genome-wide association mapping strategy to individuals with contrasting extreme trait values and mapped a new F cell quantitative trait locus to BCL11A, which encodes a zinc-finger protein, on chromosome 2p15. The 2p15 BCL11A quantitative trait locus accounts for 15.1% of the trait variance.
The cost of whole-genome bisulfite sequencing (WGBS) remains a bottleneck for many studies and it is therefore imperative to extract as much information as possible from a given dataset. This is ...particularly important because even at the recommend 30X coverage for reference methylomes, up to 50% of high-resolution features such as differentially methylated positions (DMPs) cannot be called with current methods as determined by saturation analysis. To address this limitation, we have developed a tool that dynamically segments WGBS methylomes into blocks of comethylation (COMETs) from which lost information can be recovered in the form of differentially methylated COMETs (DMCs). Using this tool, we demonstrate recovery of ∼30% of the lost DMP information content as DMCs even at very low (5X) coverage. This constitutes twice the amount that can be recovered using an existing method based on differentially methylated regions (DMRs). In addition, we explored the relationship between COMETs and haplotypes in lymphoblastoid cell lines of African and European origin. Using best fit analysis, we show COMETs to be correlated in a population-specific manner, suggesting that this type of dynamic segmentation may be useful for integrated (epi)genome-wide association studies in the future.
The molecular classification of glioblastoma (GBM) based on gene expression might better explain outcome and response to treatment than clinical factors. Whole transcriptome sequencing using ...next-generation sequencing platforms is rapidly becoming accepted as a tool for measuring gene expression for both research and clinical use. Fresh frozen (FF) tissue specimens of GBM are difficult to obtain since tumor tissue obtained at surgery is often scarce and necrotic and diagnosis is prioritized over freezing. After diagnosis, leftover tissue is usually stored as formalin-fixed paraffin-embedded (FFPE) tissue. However, RNA from FFPE tissues is usually degraded, which could hamper gene expression analysis. We compared RNA-Seq data obtained from matched pairs of FF and FFPE GBM specimens. Only three FFPE out of eleven FFPE-FF matched samples yielded informative results. Several quality-control measurements showed that RNA from FFPE samples was highly degraded but maintained transcriptomic similarities to RNA from FF samples. Certain issues regarding mutation analysis and subtype prediction were detected. Nevertheless, our results suggest that RNA-Seq of FFPE GBM specimens provides reliable gene expression data that can be used in molecular studies of GBM if the RNA is sufficiently preserved.
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