scholarly journals GENE-40. CHARACTERIZING EPIGENETIC INTRATUMORAL HETEROGENEITY IN GLIOMA USING SINGLE-CELL BISULFITE SEQUENCING

2019 ◽  
Vol 21 (Supplement_6) ◽  
pp. vi106-vi106
Author(s):  
Kevin C Johnson ◽  
Kevin Anderson ◽  
Elise Courtois ◽  
Floris Barthel ◽  
Michael Samuels ◽  
...  
Keyword(s):  
Dna Methylation ◽  
Single Cell ◽  
Bisulfite Sequencing ◽  
Cpg Island ◽  
Regulatory Element ◽  
Cell Types ◽  
Intratumoral Heterogeneity ◽  
Gene Promoters ◽  
Cpg Island Methylator Phenotype ◽  
Reduced Representation

Abstract Genetic and epigenetic alterations contribute to the observed intratumoral heterogeneity in adult glioma. Current glioma classification, based on genotype (e.g., IDH1 mutations) and DNA methylation profiles (e.g., glioma CpG Island Methylator Phenotype), can provide clinically relevant tumor subgroups. However, traditional bulk sampling fails to adequately capture the full complement of epigenomic heterogeneity, and may mask deadly features present in less abundant glioma cells. To more precisely characterize the glioma epigenome, we separately profiled single-cell DNA methylation (Reduced Representation Bisulfite Sequencing, RRBS), single-cell RNA expression (10X genomics), and bulk whole genome sequencing in nine gliomas. The genomic regions profiled by scRRBS were primarily gene promoters, but adequate coverage was also reached for glioma-specific enhancer elements and binding sites of chromatin remodelers. Unsupervised clustering of single-cell DNA methylation data revealed intratumoral variability in epigenetic classification and these cell types were distinguished by regulatory element DNA methylation. We further integrated single-cell epigenetic, single-cell transcriptomic, and genomic features to better understand gene regulation and reconstruct each tumor’s lineage history. Together, our study aims to generate a glioma cellular hierarchy shaped by the epigenetic programs that drive tumor growth.

scholarly journals Development of Enhanced Reduced Representation Bisulfite Sequencing Method for Single-cell Methylome Analysis

2017 ◽  
Vol 3 (2) ◽  
pp. 49 ◽  
Author(s):  
Junko Yamane ◽  
Tomoya Mori ◽  
Nobuko Taniyama ◽  
Kenta Kobayashi ◽  
Wataru Fujibuchi
Keyword(s):  
Dna Methylation ◽  
Single Cell ◽  
Bisulfite Sequencing ◽  
Single Cell Analysis ◽  
Cell Types ◽  
Cell Identity ◽  
Reduced Representation ◽  
Reduced Representation Bisulfite Sequencing ◽  
Methylome Analysis ◽  
Methylation Patterns

Single-cell analysis provides molecular signatures to define cell identity. To characterize cell types, DNA methylation patterns are often used as a flag of internal molecular status. There are a few reports of single-cell methylome techniques that involves bisulfite conversion. However, the step often causes DNA fragmentation, which leads to severe PCR substrate reduction. Here we developed a new version of single-cell reduced representation bisulfite sequencing (scRRBS) method to recover more CpG sites to be analysed. Our method succeeded to increase of 4.1 times in sample yield and 1.6 times in CpG site coverage. Importantly, our results indicate that the obtained single-cell DNA methylation sites are homogeneous among 12 cells, thus it may provide molecular barcodes for cell types. In summary, we succeed to develop enhanced scRRBS method and it will be more useful tool to define cell identity in the near future.

scholarly journals DNA methylation and copy number variation profiling of T-cell lymphoblastic leukemia and lymphoma

2020 ◽  
Vol 10 (4) ◽  
Author(s):  
Zahra Haider ◽  
Mattias Landfors ◽  
Irina Golovleva ◽  
Martin Erlanson ◽  
Kjeld Schmiegelow ◽  
...  
Keyword(s):  
Dna Methylation ◽  
T Cell ◽  
Copy Number Variation ◽  
Copy Number ◽  
Cpg Island ◽  
Lymphoblastic Leukemia ◽  
Clinical Manifestations ◽  
Cpg Island Methylator Phenotype ◽  
Genetic Landscape ◽  
Number Variation

AbstractDespite having common overlapping immunophenotypic and morphological features, T-cell lymphoblastic leukemia (T-ALL) and lymphoma (T-LBL) have distinct clinical manifestations, which may represent separate diseases. We investigated and compared the epigenetic and genetic landscape of adult and pediatric T-ALL (n = 77) and T-LBL (n = 15) patient samples by high-resolution genome-wide DNA methylation and Copy Number Variation (CNV) BeadChip arrays. DNA methylation profiling identified the presence of CpG island methylator phenotype (CIMP) subgroups within both pediatric and adult T-LBL and T-ALL. An epigenetic signature of 128 differentially methylated CpG sites was identified, that clustered T-LBL and T-ALL separately. The most significant differentially methylated gene loci included the SGCE/PEG10 shared promoter region, previously implicated in lymphoid malignancies. CNV analysis confirmed overlapping recurrent aberrations between T-ALL and T-LBL, including 9p21.3 (CDKN2A/CDKN2B) deletions. A significantly higher frequency of chromosome 13q14.2 deletions was identified in T-LBL samples (36% in T-LBL vs. 0% in T-ALL). This deletion, encompassing the RB1, MIR15A and MIR16-1 gene loci, has been reported as a recurrent deletion in B-cell malignancies. Our study reveals epigenetic and genetic markers that can distinguish between T-LBL and T-ALL, and deepen the understanding of the biology underlying the diverse disease localization.

scholarly journals Characterization of Global DNA Methylation in Different Gene Regions Reveals Candidate Biomarkers in Pigs with High and Low Levels of Boar Taint

2020 ◽  
Vol 7 (2) ◽  
pp. 77 ◽  
Author(s):  
Xiao Wang ◽  
Haja N. Kadarmideen
Keyword(s):  
Dna Methylation ◽  
Cpg Island ◽  
Cpg Islands ◽  
Gene Interaction ◽  
Boar Taint ◽  
Reduced Representation ◽  
Gene Interaction Networks ◽  
Reduced Representation Bisulfite Sequencing ◽  
Pathways Analysis ◽  
Porcine Gene

DNA methylation of different gene components, including different exons and introns, or different lengths of exons and introns is associated with differences in gene expression. To investigate the methylation of porcine gene components associated with the boar taint (BT) trait, this study used reduced representation bisulfite sequencing (RRBS) data from nine porcine testis samples in three BT groups (low, medium and high BT). The results showed that the methylation levels of the first exons and first introns were lower than those of the other exons and introns. The first exons/introns of CpG island regions had even lower levels of methylation. A total of 123 differentially methylated promoters (DMPs), 194 differentially methylated exons (DMEs) and 402 differentially methylated introns (DMIs) were identified, of which 80 DMPs (DMP-CpGis), 112 DMEs (DME-CpGis) and 166 DMIs (DMI-CpGis) were discovered in CpG islands. Importantly, GPX1 contained one each of DMP, DME, DMI, DMP-CpGi, DME-CpGi and DMI-CpGi. Gene-GO term relationships and pathways analysis showed DMP-CpGi-related genes are mainly involved in methylation-related biological functions. In addition, gene–gene interaction networks consisted of nodes that were hypo-methylated GPX1, hypo-methylated APP, hypo-methylated ATOX1, hyper-methylated ADRB2, hyper-methylated RPS6KA1 and hyper-methylated PNMT. They could be used as candidate biomarkers for reducing boar taint in pigs, after further validation in large cohorts.

scholarly journals How to Design a Whole-Genome Bisulfite Sequencing Experiment

Epigenomes ◽  
2018 ◽  
Vol 2 (4) ◽  
pp. 21 ◽  
Author(s):  
Claudius Grehl ◽  
Markus Kuhlmann ◽  
Claude Becker ◽  
Bruno Glaser ◽  
Ivo Grosse
Keyword(s):  
Dna Methylation ◽  
Bisulfite Sequencing ◽  
Alternative Methods ◽  
Transcriptional Gene Silencing ◽  
Whole Genome ◽  
Reduced Representation ◽  
Genome Region ◽  
Whole Genome Bisulfite Sequencing ◽  
Sequencing Experiment ◽  
Genome Bisulfite Sequencing

Aside from post-translational histone modifications and small RNA populations, the epigenome of an organism is defined by the level and spectrum of DNA methylation. Methyl groups can be covalently bound to the carbon-5 of cytosines or the carbon-6 of adenine bases. DNA methylation can be found in both prokaryotes and eukaryotes. In the latter, dynamic variation is shown across species, along development, and by cell type. DNA methylation usually leads to a lower binding affinity of DNA-interacting proteins and often results in a lower expression rate of the subsequent genome region, a process also referred to as transcriptional gene silencing. We give an overview of the current state of research facilitating the planning and implementation of whole-genome bisulfite-sequencing (WGBS) experiments. We refrain from discussing alternative methods for DNA methylation analysis, such as reduced representation bisulfite sequencing (rrBS) and methylated DNA immunoprecipitation sequencing (MeDIPSeq), which have value in specific experimental contexts but are generally disadvantageous compared to WGBS.

scholarly journals Ultradeep Bisulfite Sequencing Analysis of DNA Methylation Patterns in Multiple Gene Promoters by 454 Sequencing

2007 ◽  
Vol 67 (18) ◽  
pp. 8511-8518 ◽  
Author(s):  
Kristen H. Taylor ◽  
Robin S. Kramer ◽  
J. Wade Davis ◽  
Juyuan Guo ◽  
Deiter J. Duff ◽  
...  
Keyword(s):  
Dna Methylation ◽  
Bisulfite Sequencing ◽  
454 Sequencing ◽  
Sequencing Analysis ◽  
Gene Promoters ◽  
Multiple Gene ◽  
Methylation Patterns ◽  
Bisulfite Sequencing Analysis

Profiling of Aberrant DNA Methylation In AML Reveals Subclasses of CpG Islands with Epigenetic or Genetic Association

Blood ◽  
2010 ◽  
Vol 116 (21) ◽  
pp. 2498-2498
Author(s):  
Claudia Gebhard ◽  
Mohammed Sadeh ◽  
Dagmar Glatz ◽  
Lucia Schwarzfischer ◽  
Rainer Spang ◽  
...  
Keyword(s):  
Dna Methylation ◽  
De Novo ◽  
Conflicts Of Interest ◽  
Cpg Island ◽  
Cpg Islands ◽  
Sequencing Data ◽  
Cpg Island Methylator Phenotype ◽  
Chip Sequencing ◽  
Aberrant Dna Methylation ◽  
Methylation Patterns

Abstract Abstract 2498 CpG islands show frequent and often disease-specific epigenetic alterations during malignant transformation, however, the underlying mechanisms are poorly understood. We used methyl-CpG immunoprecipitation (MCIp) to generate comparative DNA methylation profiles of 30 patients with acute myeloid leukemia for human CpG islands across the genome. DNA methylation profiles across 23.000 CpG islands revealed highly heterogeneous methylation patterns in AML with over 6000 CpG islands showing aberrant de novo methylation in AML. Based on these profiles we selected a subset of 380 CpG islands (covering 15.000 individual CpGs) for detailed fine-mapping analyses of aberrant DNA methylation in 185 patients with AML (50% normal karyotype). We found that a proportion of patients (5/185) displayed a concerted hypermethylation at almost all studied loci, representing the rare CpG island methylator phenotype (CIMP) in AML. Meta analysis of methylation profiling and published ChIP sequencing data separated CpG islands in two groups. A highly correlated subgroup of CpG island regions was strongly associated with histone H3 lysine 27 trimethylation in human hematopoietic progenitor cells, suggesting that disease-related de novo DNA methylation at these CpG islands is linked with polycomb group protein (PcG)-mediated repression. The group of mainly non-PcG target CpG islands showed heterogeneous methylation patterns across patients and unsupervised hierarchical clustering revealed a correlation of methylation profiles with genetic disease markers, including oncofusion proteins as well as CEBPA- and NPM1-mutations. Our study suggests that both epigenetic as well as genetic aberrations may underlay AML-related changes in CpG island DNA methylation states. Disclosures: No relevant conflicts of interest to declare.

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