The use of two kinase inhibitors (2i) enables derivation of mouse embryonic stem cells (ESCs) in the pluripotent ground state. Using whole-genome bisulfite sequencing (WGBS), we show that male 2i ...ESCs are globally hypomethylated compared to conventional ESCs maintained in serum. In serum, female ESCs are hypomethyated similarly to male ESCs in 2i, and DNA methylation is further reduced in 2i. Regions with elevated DNA methylation in 2i strongly correlate with the presence of H3K9me3 on endogenous retroviruses (ERVs) and imprinted loci. The methylome of male ESCs in serum parallels postimplantation blastocyst cells, while 2i stalls ESCs in a hypomethylated, ICM-like state. WGBS analysis during adaptation of 2i ESCs to serum suggests that deposition of DNA methylation is largely random, while loss of DNA methylation during reversion to 2i occurs passively, initiating at TET1 binding sites. Together, our analysis provides insight into DNA methylation dynamics in cultured ESCs paralleling early developmental processes.
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•WGBS reveals dynamics of conversion between hypo- and hypermethylated mESC states•Maintained DNA methylation in 2i ESCs correlates with the presence of H3K9me3•The methylome of ground state 2i ESCs resembles preimplantation blastocyst cells•Hypomethylation in 2i is dependent on both inhibitors and LIF
Genome-wide DNA methylation analysis highlights changes that occur during pluripotent state transitions and identifies regions of methylation stability.
Six extant species of non-human great apes are currently recognized: Sumatran and Bornean orangutans, eastern and western gorillas, and chimpanzees and bonobos 1. However, large gaps remain in our ...knowledge of fine-scale variation in hominoid morphology, behavior, and genetics, and aspects of great ape taxonomy remain in flux. This is particularly true for orangutans (genus: Pongo), the only Asian great apes and phylogenetically our most distant relatives among extant hominids 1. Designation of Bornean and Sumatran orangutans, P. pygmaeus (Linnaeus 1760) and P. abelii (Lesson 1827), as distinct species occurred in 2001 1, 2. Here, we show that an isolated population from Batang Toru, at the southernmost range limit of extant Sumatran orangutans south of Lake Toba, is distinct from other northern Sumatran and Bornean populations. By comparing cranio-mandibular and dental characters of an orangutan killed in a human-animal conflict to those of 33 adult male orangutans of a similar developmental stage, we found consistent differences between the Batang Toru individual and other extant Ponginae. Our analyses of 37 orangutan genomes provided a second line of evidence. Model-based approaches revealed that the deepest split in the evolutionary history of extant orangutans occurred ∼3.38 mya between the Batang Toru population and those to the north of Lake Toba, whereas both currently recognized species separated much later, about 674 kya. Our combined analyses support a new classification of orangutans into three extant species. The new species, Pongo tapanuliensis, encompasses the Batang Toru population, of which fewer than 800 individuals survive.
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•We describe a new species of great apes, the Tapanuli orangutan Pongo tapanuliensis•Genomic analyses corroborate morphological distinctiveness of P. tapanuliensis•P. tapanuliensis comprises the oldest evolutionary lineage in the genus Pongo•With fewer than 800 individuals, P. tapanuliensis is among the most endangered great apes
Nater et al. describe a new great ape species, the Tapanuli orangutan Pongo tapanuliensis. An isolated population from Batang Toru is highly distinct from the northern Sumatran and Bornean species, based on morphological variation, corroborated by population genomic analyses. Fewer than 800 individuals of P. tapanuliensis survive in the wild.
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.
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.
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Efficient sharing and integration of phenotypic data is crucial for advancing biomedical research and enhancing patient outcomes in precision medicine and public health. To achieve ...this, the health data community has developed standards to promote the harmonization of variable names and values. However, the use of diverse standards across different research centers can hinder progress. Here we present Convert-Pheno, an open-source software toolkit that enables the interconversion of common data models for phenotypic data such as Beacon v2 Models, CDISC-ODM, OMOP-CDM, Phenopackets v2, and REDCap. Along with the software, we have created a detailed documentation that includes information on deployment and installation.
Pyrosequencing is a sequencing-by-synthesis method that quantitatively monitors the real-time incorporation of nucleotides through the enzymatic conversion of released pyrophosphate into a ...proportional light signal. Quantitative measures are of special importance for DNA methylation analysis in various developmental and pathological situations. Analysis of DNA methylation patterns by pyrosequencing combines a simple reaction protocol with reproducible and accurate measures of the degree of methylation at several CpGs in close proximity with high quantitative resolution. After bisulfite treatment and PCR, the degree of each methylation at each CpG position in a sequence is determined from the ratio of T and C. The process of purification and sequencing can be repeated for the same template to analyze other CpGs in the same amplification product. Quantitative epigenotypes are obtained using this protocol in approximately 4 h for up to 96 DNA samples when bisulfite-treated DNA is already available as the starting material.
Chronic lymphocytic leukemia is a complex and heterogeneous hematological malignancy. The advance of high-throughput multi-omics technologies has significantly influenced chronic lymphocytic leukemia ...research and paved the way for precision medicine approaches. In this review, we explore the role of machine learning in the analysis of multi-omics data in this hematological malignancy. We discuss recent literature on different machine learning models applied to single omic studies in chronic lymphocytic leukemia, with a special focus on the potential contributions to precision medicine. Finally, we highlight the recently published machine learning applications in multi-omics data in this area of research as well as their potential and limitations.
Genome sequencing projects are discovering millions of genetic variants in humans, and interpretation of their functional effects is essential for understanding the genetic basis of variation in ...human traits. Here we report sequencing and deep analysis of messenger RNA and microRNA from lymphoblastoid cell lines of 462 individuals from the 1000 Genomes Project--the first uniformly processed high-throughput RNA-sequencing data from multiple human populations with high-quality genome sequences. We discover extremely widespread genetic variation affecting the regulation of most genes, with transcript structure and expression level variation being equally common but genetically largely independent. Our characterization of causal regulatory variation sheds light on the cellular mechanisms of regulatory and loss-of-function variation, and allows us to infer putative causal variants for dozens of disease-associated loci. Altogether, this study provides a deep understanding of the cellular mechanisms of transcriptome variation and of the landscape of functional variants in the human genome.
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.
Abstract
In the late 19th century, formalin fixation with paraffin-embedding (FFPE) of tissues was developed as a fixation and conservation method and is still used to this day in routine clinical ...and pathological practice. The implementation of state-of-the-art nucleic acid sequencing technologies has sparked much interest for using historical FFPE samples stored in biobanks as they hold promise in extracting new information from these valuable samples. However, formalin fixation chemically modifies DNA, which potentially leads to incorrect sequences or misinterpretations in downstream processing and data analysis. Many publications have concentrated on one type of DNA damage, but few have addressed the complete spectrum of FFPE-DNA damage. Here, we review mitigation strategies in (I) pre-analytical sample quality control, (II) DNA repair treatments, (III) analytical sample preparation and (IV) bioinformatic analysis of FFPE-DNA. We then provide recommendations that are tested and illustrated with DNA from 13-year-old liver specimens, one FFPE preserved and one fresh frozen, applying target-enriched sequencing. Thus, we show how DNA damage can be compensated, even when using low quantities (50 ng) of fragmented FFPE-DNA (DNA integrity number 2.0) that cannot be amplified well (Q129 bp/Q41 bp = 5%). Finally, we provide a checklist called ‘ERROR-FFPE-DNA’ that summarises recommendations for the minimal information in publications required for assessing fitness-for-purpose and inter-study comparison when using FFPE samples.
Graphical Abstract
Graphical Abstract
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