Genome-wide DNA Methylation Patterns Research Articles

Mining Novel Candidate Imprinted Genes Using Genome-Wide Methylation Screening and Literature Review

Large-scale transcriptome and methylome data analyses obtained by high-throughput technologies have been enabling the identification of novel imprinted genes. We investigated genome-wide DNA methylation patterns in multiple human tissues, using a high-resolution microarray to uncover hemimethylated CpGs located in promoters overlapping CpG islands, aiming to identify novel candidate imprinted genes. Using our approach, we recovered ~30% of the known human imprinted genes, and a further 168 candidates were identified, 61 of which with at least three hemimethylated CpGs shared by more than two tissue types. Thirty-four of these candidate genes are members of the protocadherin cluster on 5q31.3; in mice, protocadherin genes have non-imprinted random monoallelic expression, which might also be the case in humans. Among the remaining 27 genes, ZNF331 was recently validated as an imprinted gene, and six of them have been reported as candidates, supporting our prediction. Five candidates (CCDC166, ARC, PLEC, TONSL, and VPS28) map to 8q24.3, and might constitute a novel imprinted cluster. Additionally, we performed a comprehensive compilation of known human and mice imprinted genes from literature and databases, and a comparison among high-throughput imprinting studies in humans. The screening for hemimethylated CpGs shared by multiple human tissues, together with the extensive review, appears to be a useful approach to reveal candidate imprinted genes.

Genome-wide DNA methylation patterns in coronary heart disease.

To better understand the molecular mechanisms of atherosclerosis, we conducted acomparative analysis of DNA methylation patterns in right coronary arteries in the area of advanced atherosclerotic plaques (CAP), great saphenous vein (GSV), and internal mammary artery (IMA) of patients affected by coronary heart disease. DNA methylation data (accession number E‑GEOD-62867) were divided into three paired groups: CAP vs. IMA, CAP vs. GSV, and IMA vs. GSV. Differentially methylated genes (DMGs) were extracted to analyze the changes in the DMGs in the three different tissues. The gplots package was used for the clustering and heatmap analysis of DMGs. Subsequently, DMG-related pathways were identified using DAVID (Database for Annotation, Visualization and Integrated Discovery) and transcription factors (TFs) were predicted. Based on the filtering criterion of p< 0.05, and a mean beta value difference of ≥0.2, there were 252, 373, and 259 DMGs, respectively, in the CAP vs. IMA, CAP vs. GSV, and IMA vs. GSV groups. Interestingly, the S100A10 gene was hypomethylated in CAP compared with IMA and GSV. Clustering and heatmap analyses suggested that DMGs were segregated into two distinct clusters. Hypermethylated genes in CAP as compared with GSV were only involved in the pathway of fat digestion and absorption, while hypomethylated genes in CAP compared with GSV mainly participated in immune response-associated pathways (cytokine-cytokine receptor interaction, MAPK signaling pathway). The DNA methylation differences in vascular tissues of patients with coronary artery disease may provide new insights into the mechanisms underlying the development of atherosclerosis. The functions identified here-cytokine-cytokine receptor interaction, MAPK signaling pathway, DMG (S100A10), and TF (NF-kB)-may serve as potential targets in the treatment of atherosclerosis.

A streamlined method for analysing genome-wide DNA methylation patterns from low amounts of FFPE DNA

BackgroundFormalin fixed paraffin embedded (FFPE) tumor samples are a major source of DNA from patients in cancer research. However, FFPE is a challenging material to work with due to macromolecular fragmentation and nucleic acid crosslinking. FFPE tissue particularly possesses challenges for methylation analysis and for preparing sequencing-based libraries relying on bisulfite conversion. Successful bisulfite conversion is a key requirement for sequencing-based methylation analysis.MethodsHere we describe a complete and streamlined workflow for preparing next generation sequencing libraries for methylation analysis from FFPE tissues. This includes, counting cells from FFPE blocks and extracting DNA from FFPE slides, testing bisulfite conversion efficiency with a polymerase chain reaction (PCR) based test, preparing reduced representation bisulfite sequencing libraries and massively parallel sequencing.ResultsThe main features and advantages of this protocol are:An optimized method for extracting good quality DNA from FFPE tissues.An efficient bisulfite conversion and next generation sequencing library preparation protocol that uses 50 ng DNA from FFPE tissue.Incorporation of a PCR-based test to assess bisulfite conversion efficiency prior to sequencing.ConclusionsWe provide a complete workflow and an integrated protocol for performing DNA methylation analysis at the genome-scale and we believe this will facilitate clinical epigenetic research that involves the use of FFPE tissue.

Trends in DNA Methylation with Age Replicate Across Diverse Human Populations.

Aging is associated with widespread changes in genome-wide patterns of DNA methylation. Thousands of CpG sites whose tissue-specific methylation levels are strongly correlated with chronological age have been previously identified. However, the majority of these studies have focused primarily on cosmopolitan populations living in the developed world; it is not known if age-related patterns of DNA methylation at these loci are similar across a broad range of human genetic and ecological diversity. We investigated genome-wide methylation patterns using saliva- and whole blood-derived DNA from two traditionally hunting and gathering African populations: the Baka of the western Central African rain forest and the ≠Khomani San of the South African Kalahari Desert. We identified hundreds of CpG sites whose methylation levels are significantly associated with age, thousands that are significant in a meta-analysis, and replicate trends previously reported in populations of non-African descent. We confirmed that an age-associated site in the promoter of the gene ELOVL2 shows a remarkably congruent relationship with aging in humans, despite extensive genetic and environmental variation across populations. We also demonstrate that genotype state at methylation quantitative trait loci (meQTLs) can affect methylation trends at some age-associated CpG sites. Our study explores the relationship between CpG methylation and chronological age in populations of African hunter-gatherers, who rely on different diets across diverse ecologies. While many age-related CpG sites replicate across populations, we show that considering common genetic variation at meQTLs further improves our ability to detect previously identified age associations.

Abstract LB-097: DNA methylation at enhancers distinguishes distinct breast cancer lineages

Abstract Breast cancers exhibit genome-wide aberrant DNA methylation patterns. To investigate how these affect the transcriptome and which changes are linked to transformation or progression we applied genome-wide expression-methylation quantitative trait loci (emQTL) analysis between 5meCpG and gene expression. We find that on a whole genome scale, DNA methylation and gene expression have remarkably and reproducibly conserved patterns of association, both in cis and in trans, which lead to identify distinct transcriptional networks and specific disease cell lineages deviating from the normal tissue. In three breast cancer cohorts (n=104, n=253 and n=330), we invariably identify enhancers whose DNA methylation tethers the binding of three key transcription factors in breast cancer in a lineage specific manner. Our emQTL analysis also identifies tumor infiltrating immune cell signatures. Using ChromHMM segmentation and ChIP-seq associated TF binding regions, we identify enhancers and TF binding sites around emQTLs-CpGs. We functionally validate the genes associated with these CpGs using knock-down by siRNA and applying GRO-seq analysis after transcriptional stimulation with estrogen. DNA methylation at TF binding regions is considered an early event during normal breast cell transformation into estrogen dependent breast cancer. Two distinct key gene regulatory networks reported here are prominently altered by DNA methylation. Citation Format: Thomas Fleischer, Xavier Tekpli, Anthony Mathelier, Shixiong Wang, Daniel Nebdal, Hari P. Dhakal, Kristine Kleivi Sandberg, Ellen Schliting, Oslo Breast Cancer Research Consortium (OSBREAC), Anne-Lise Børresen-Dale, Elin Borgen, Bjørn Naume, Ragnhild Eskeland, Arnoldo Frigessi, Jörg Tost, Antoni Hurtado, Vessela N. Kristensen. DNA methylation at enhancers distinguishes distinct breast cancer lineages [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2017; 2017 Apr 1-5; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2017;77(13 Suppl):Abstract nr LB-097. doi:10.1158/1538-7445.AM2017-LB-097

EXERCISE TRAINING-INDUCED CHANGES ON DNA METHYLATION IN MILD COGNITIVELY IMPAIRED OLDER AFRICAN AMERICANS: GEMS STUDY

DNA methylation at CpG sites is an important epigenetic modification of the human genome that can affect gene expression and potentially influence health outcomes. Notably, aging and physical exercise have been shown to influence epigenetic modification with during minimal exercise, inducing hypomethylation of the entire genome within muscle cells. However, there is paucity of data on exercise-induced training adaptation on DNA methylation in mild cognitively impaired (MCI) older African American (AA) undergoing well-characterized and supervised exercise intervention. Objective of Study: We explored the effects of a six-month aerobic and stretch exercise intervention on the epigenetic modifications and genome-wide DNA methylation of enrolled participants. Twenty-nine older AAs volunteers confirmed to be MCI, were randomly, assigned into a six-month program of either aerobic or stretch exercise. Participants underwent 40 minutes of supervised training 3 times/week. The maximal oxygen consumption (VO2Max) was determined at baseline and repeated after subjects completed the 6-month exercise program. Genome-wide methylation levels at CpG sites from 19 participants at baseline and a 6-month exercise training were profiled by using the Infinium HumanMethylation450 BeadChip assay. We examined the association between the change in CpG sites intensities and VO2 Max after 6 months of exercise adjusting for Age, APOE4 carrier status and randomization group. The participants averaged ∼71 years old in age, were 67.86% female and similar in mean relative VO2 Max (Stretch=24.72 (8.68) vs. Aerobic=22.74 (2.69); p = 0.715) at baseline. In total, we obtained DNA methylation data or 485,513 CpG sites from blood of 19 participants at baseline and after the exercise adaptation. Overall, six CpG sites demonstrated significant training-induced changes in methylation profile: p-values below 5 x 10-5 (cg06648780, cg06120404, cg08733824, cg14318197, cg26580761, cg24583624) with the first two CpG sites located on the ALCAM (leukocyte adhesion molecule) and C1orf88 (primary cilia formation) genes. We provide a genome-wide DNA methylation patterns associated with training-induced changes after a 6-month exercise adaptation. Whereas, efforts to understand the implications of these changes are ongoing, future studies should inform the relationships of changes in DNA methylation to mRNA expression in a predominantly AA sample, and correlate such changes with the MCI clinical phenotype.

Genome-wide DNA methylation analysis of the porcine hypothalamus-pituitary-ovary axis

Previous studies have suggested that DNA methylation in both CpG and CpH (where H = C, T or A) contexts plays a critical role in biological functions of different tissues. However, the genome-wide DNA methylation patterns of porcine hypothalamus-pituitary-ovary (HPO) tissues remain virtually unexplored. In this study, methylomes of HPO tissues were profiled to investigate their differences and similarities. We found that HPO methylomes displayed tissue-specific methylation patterns in both CpG and CpH contexts. At gene locations, the methylation and density of CpGs was negatively linked at transcription start sites but positively linked at transcription end sites. The densities of CpGs and CpHs at CpG island (CGI) locations were negatively correlated with their methylation. Moreover, the methylation interactions between CGIs and genes showed similar pattern in the CpG context but tissue-specific pattern in the CpH context. CpGs located in CGIs, upstream regions and exons were protected from methylation dynamics, whereas CGI shores, CGI shelves and intergenic regions were more likely to be targets of methylation changes. The methylation dynamics enriching in a tissue-specific manner appeared to maintain and establish the biological functions of HPO tissues. Our analyses provided valuable insights into the tissue-specific methylomes of porcine HPO tissues.

The mutational signatures and molecular alterations of bladder cancer

All cancers are transformed from normal cells as a result of somatic alterations in the genomes. During the last few years, many research groups, including the International Cancer Genome Consortium (ICGC) and The Cancer Genome Atlas (TCGA), have conducted large-scale cancer genome studies worldwide. These efforts identified millions of somatic mutations accumulated in cancer cells, including point mutations, copy number variations and chromosomal rearrangements, as well as gene expression profiles and genome-wide DNA methylation patterns. These datasets accelerated our understanding of many biological processes operative in cancers. Genomes of bladder cancer, one of the most common cancers worldwide, were recently analyzed by several groups, which provide a comprehensive landscape of molecular alterations. This review article discusses the patterns of somatic mutations, mutational signatures, frequently altered genes and molecular pathways in bladder cancers.

The mutational signatures and molecular alterations of bladder cancer

: All cancers are transformed from normal cells as a result of somatic alterations in the genomes. During the last few years, many research groups, including the International Cancer Genome Consortium (ICGC) and The Cancer Genome Atlas (TCGA), have conducted large-scale cancer genome studies worldwide. These efforts identified millions of somatic mutations accumulated in cancer cells, including point mutations, copy number variations and chromosomal rearrangements, as well as gene expression profiles and genome-wide DNA methylation patterns. These datasets accelerated our understanding of many biological processes operative in cancers. Genomes of bladder cancer, one of the most common cancers worldwide, were recently analyzed by several groups, which provide a comprehensive landscape of molecular alterations. This review article discusses the patterns of somatic mutations, mutational signatures, frequently altered genes and molecular pathways in bladder cancers.

Histone 3.3 hotspot mutations in conventional osteosarcomas: a comprehensive clinical and molecular characterization of six H3F3A mutated cases

BackgroundHistone 3.3 (H3.3) hotspot mutations in bone tumors occur in the vast majority of giant cell tumors of bone (GCTBs; 96%), chondroblastomas (95%) and in a few cases of osteosarcomas. However, clinical presentation, histopathological features, and additional molecular characteristics of H3.3 mutant osteosarcomas are largely unknown.MethodsIn this multicentre, retrospective study, a total of 106 conventional high-grade osteosarcomas, across all age groups were re-examined for hotspot mutations in the H3.3 coding genes H3F3A and H3F3B. H3.3 mutant osteosarcomas were re-evaluated in a multidisciplinary manner and analyzed for genome-wide DNA-methylation patterns and DNA copy number aberrations alongside H3.3 wild-type osteosarcomas and H3F3A G34W/L mutant GCTBs.ResultsSix osteosarcomas (6/106) carried H3F3A hotspot mutations. No mutations were found in H3F3B. All patients with H3F3A mutant osteosarcoma were older than 30 years with a median age of 65 years. Copy number aberrations that are commonly encountered in high-grade osteosarcomas also occurred in H3F3A mutant osteosarcomas. Unlike a single osteosarcoma with a H3F3A K27M mutation, the DNA methylation profiles of H3F3A G34W/R mutant osteosarcomas were clearly different from H3.3 wild-type osteosarcomas, but more closely related to GCTBs. The most differentially methylated promoters between H3F3A G34W/R mutant and H3.3 wild-type osteosarcomas were in KLLN/PTEN (p < 0.00005) and HIST1H2BB (p < 0.0005).ConclusionsH3.3 mutations in osteosarcomas may occur in H3F3A at mutational hotspots. They are overall rare, but become more frequent in osteosarcoma patients older than 30 years. Osteosarcomas carrying H3F3A G34W/R mutations are associated with epigenetic dysregulation of KLLN/PTEN and HIST1H2BB.

DNA methylation in lung tissues of mouse offspring exposed in utero to polycyclic aromatic hydrocarbons

Polycyclic aromatic hydrocarbons (PAHs) comprise an important class of environmental pollutants that are known to cause lung cancer in animals and are suspected lung carcinogens in humans. Moreover, evidence from cell-based studies points to PAHs as modulators of the epigenome. The objective of this work was to assess patterns of genome-wide DNA methylation in lung tissues of adult offspring initiated in utero with the transplacental PAH carcinogens dibenzo [def,p]chrysene (DBC) or benzo [a]pyrene (BaP). Genome-wide methylation patterns for normal (not exposed), normal adjacent and lung tumor tissues obtained from adult offspring were determined using methylated DNA immunoprecipitation (MeDIP) with the NimbleGen mouse DNA methylation CpG island array. Lung tumor incidence in 45-week old mice initiated with BaP was 32%, much lower than that of the DBC-exposed offspring at 96%. Also, male offspring appeared more susceptible to BaP as compared to females. Distinct patterns of DNA methylation were associated with non-exposed, normal adjacent and adenocarcinoma lung tissues, as determined by principal components, hierarchical clustering and gene ontology analyses. From these methylation profiles, a set of genes of interest was identified that includes potential important targets for epigenetic modification during the process of lung tumorigenesis in animals exposed to environmental PAHs.

Genome-wide DNA methylation measurements in prostate tissues uncovers novel prostate cancer diagnostic biomarkers and transcription factor binding patterns

BackgroundCurrent diagnostic tools for prostate cancer lack specificity and sensitivity for detecting very early lesions. DNA methylation is a stable genomic modification that is detectable in peripheral patient fluids such as urine and blood plasma that could serve as a non-invasive diagnostic biomarker for prostate cancer.MethodsWe measured genome-wide DNA methylation patterns in 73 clinically annotated fresh-frozen prostate cancers and 63 benign-adjacent prostate tissues using the Illumina Infinium HumanMethylation450 BeadChip array. We overlaid the most significantly differentially methylated sites in the genome with transcription factor binding sites measured by the Encyclopedia of DNA Elements consortium. We used logistic regression and receiver operating characteristic curves to assess the performance of candidate diagnostic models.ResultsWe identified methylation patterns that have a high predictive power for distinguishing malignant prostate tissue from benign-adjacent prostate tissue, and these methylation signatures were validated using data from The Cancer Genome Atlas Project. Furthermore, by overlaying ENCODE transcription factor binding data, we observed an enrichment of enhancer of zeste homolog 2 binding in gene regulatory regions with higher DNA methylation in malignant prostate tissues.ConclusionsDNA methylation patterns are greatly altered in prostate cancer tissue in comparison to benign-adjacent tissue. We have discovered patterns of DNA methylation marks that can distinguish prostate cancers with high specificity and sensitivity in multiple patient tissue cohorts, and we have identified transcription factors binding in these differentially methylated regions that may play important roles in prostate cancer development.

Human liver epigenetic alterations in non-alcoholic steatohepatitis are related to insulin action

ABSTRACTBoth genetic and lifestyle factors contribute to the risk of non-alcoholic steatohepatitis (NASH). Additionally, epigenetic modifications may also play a key role in the pathogenesis of NASH. We therefore investigated liver DNA methylation, as a marker for epigenetic alterations, in individuals with simple steatosis and NASH, and further tested if these alterations were associated with clinical phenotypes. Liver biopsies obtained from 95 obese individuals (age: 49.5 ± 7.7 years, BMI: 43 ± 5.7 kg/m2, type 2 diabetes [T2D]: 35) as a wedge biopsy during a Roux-en-Y gastric bypass operation were investigated. Thirty-four individuals had a normal liver phenotype, 35 had simple steatosis, and 26 had NASH. Genome-wide DNA methylation pattern was analyzed using the Infinium HumanMethylation450 BeadChip. mRNA expression was analyzed from 42 individuals using the HumanHT-12 Expression BeadChip. We identified 1,292 CpG sites representing 677 unique genes differentially methylated in liver of individuals with NASH (q < 0.001), independently of T2D, age, sex, and BMI. Focusing on the top-ranking 30 and another 37 CpG sites mapped to genes enriched in pathways of metabolism (q = 0.0036) and cancer (q = 0.0001) all together, 59 NASH-associated CpG sites correlated with fasting insulin levels independently of age, fasting glucose, or T2D. From these, we identified 30 correlations between DNA methylation and mRNA expression, for example LDHB (r = −0.45, P = 0.003). We demonstrated that NASH, more than simple steatosis, associates with differential DNA methylation in the human liver. These epigenetic alterations in NASH are linked with insulin metabolism.

Dietary restriction protects from age-associated DNA methylation and induces epigenetic reprogramming of lipid metabolism

BackgroundDietary restriction (DR), a reduction in food intake without malnutrition, increases most aspects of health during aging and extends lifespan in diverse species, including rodents. However, the mechanisms by which DR interacts with the aging process to improve health in old age are poorly understood. DNA methylation could play an important role in mediating the effects of DR because it is sensitive to the effects of nutrition and can affect gene expression memory over time.ResultsHere, we profile genome-wide changes in DNA methylation, gene expression and lipidomics in response to DR and aging in female mouse liver. DR is generally strongly protective against age-related changes in DNA methylation. During aging with DR, DNA methylation becomes targeted to gene bodies and is associated with reduced gene expression, particularly of genes involved in lipid metabolism. The lipid profile of the livers of DR mice is correspondingly shifted towards lowered triglyceride content and shorter chain length of triglyceride-associated fatty acids, and these effects become more pronounced with age.ConclusionsOur results indicate that DR remodels genome-wide patterns of DNA methylation so that age-related changes are profoundly delayed, while changes at loci involved in lipid metabolism affect gene expression and the resulting lipid profile.

DNA methylation-based classification and grading system for meningioma: a multicentre, retrospective analysis

The WHO classification of brain tumours describes 15 subtypes of meningioma. Nine of these subtypes are allotted to WHO grade I, and three each to grade II and grade III. Grading is based solely on histology, with an absence of molecular markers. Although the existing classification and grading approach is of prognostic value, it harbours shortcomings such as ill-defined parameters for subtypes and grading criteria prone to arbitrary judgment. In this study, we aimed for a comprehensive characterisation of the entire molecular genetic landscape of meningioma to identify biologically and clinically relevant subgroups. In this multicentre, retrospective analysis, we investigated genome-wide DNA methylation patterns of meningiomas from ten European academic neuro-oncology centres to identify distinct methylation classes of meningiomas. The methylation classes were further characterised by DNA copy number analysis, mutational profiling, and RNA sequencing. Methylation classes were analysed for progression-free survival outcomes by the Kaplan-Meier method. The DNA methylation-based and WHO classification schema were compared using the Brier prediction score, analysed in an independent cohort with WHO grading, progression-free survival, and disease-specific survival data available, collected at the Medical University Vienna (Vienna, Austria), assessing methylation patterns with an alternative methylation chip. We retrospectively collected 497 meningiomas along with 309 samples of other extra-axial skull tumours that might histologically mimic meningioma variants. Unsupervised clustering of DNA methylation data clearly segregated all meningiomas from other skull tumours. We generated genome-wide DNA methylation profiles from all 497 meningioma samples. DNA methylation profiling distinguished six distinct clinically relevant methylation classes associated with typical mutational, cytogenetic, and gene expression patterns. Compared with WHO grading, classification by individual and combined methylation classes more accurately identifies patients at high risk of disease progression in tumours with WHO grade I histology, and patients at lower risk of recurrence among WHO grade II tumours (p=0·0096) from the Brier prediction test). We validated this finding in our independent cohort of 140 patients with meningioma. DNA methylation-based meningioma classification captures clinically more homogenous groups and has a higher power for predicting tumour recurrence and prognosis than the WHO classification. The approach presented here is potentially very useful for stratifying meningioma patients to observation-only or adjuvant treatment groups. We consider methylation-based tumour classification highly relevant for the future diagnosis and treatment of meningioma. German Cancer Aid, Else Kröner-Fresenius Foundation, and DKFZ/Heidelberg Institute of Personalized Oncology/Precision Oncology Program.

DNA methylation profiling reveals the presence of population-specific signatures correlating with phenotypic characteristics.

Phenotypic characteristics are known to vary substantially among different ethnicities around the globe. These variations are mediated by number of stochastic events and cannot be attributed to genetic architecture alone. DNA methylation is a well-established mechanism that sculpts our epigenome influencing phenotypic variation including disease manifestation. Since DNA methylation is an important determinant for health issues of a population, it demands a thorough investigation of the natural differences in genome wide DNA methylation patterns across different ethnic groups. This study is based on comparative analyses of methylome from five different ethnicities with major focus on Indian subjects. The current study uses hierarchical clustering approaches, principal component analysis and locus specific differential methylation analysis on Illumina 450K methylation data to compare methylome of different ethnic subjects. Our data indicates that the variations in DNA methylation patterns of Indians are less among themselves compared to other global population. It empirically correlated with dietary, cultural and demographical divergences across different ethnic groups. Our work further suggests that Indians included in this study, despite their genetic similarity with the Caucasian population, are in close proximity with Japanese in terms of their methylation signatures.

Identification of a Pseudomonas aeruginosa PAO1 DNA Methyltransferase, Its Targets, and Physiological Roles.

ABSTRACTDNA methylation is widespread among prokaryotes, and most DNA methylation reactions are catalyzed by adenine DNA methyltransferases, which are part of restriction-modification (R-M) systems. R-M systems are known for their role in the defense against foreign DNA; however, DNA methyltransferases also play functional roles in gene regulation. In this study, we used single-molecule real-time (SMRT) sequencing to uncover the genome-wide DNA methylation pattern in the opportunistic pathogen Pseudomonas aeruginosa PAO1. We identified a conserved sequence motif targeted by an adenine methyltransferase of a type I R-M system and quantified the presence of N6-methyladenine using liquid chromatography-tandem mass spectrometry (LC-MS/MS). Changes in the PAO1 methylation status were dependent on growth conditions and affected P. aeruginosa pathogenicity in a Galleria mellonella infection model. Furthermore, we found that methylated motifs in promoter regions led to shifts in sense and antisense gene expression, emphasizing the role of enzymatic DNA methylation as an epigenetic control of phenotypic traits in P. aeruginosa. Since the DNA methylation enzymes are not encoded in the core genome, our findings illustrate how the acquisition of accessory genes can shape the global P. aeruginosa transcriptome and thus may facilitate adaptation to new and challenging habitats.

Genome-wide DNA methylation analysis in renal ischemia reperfusion injury

Renal ischemia reperfusion injury (IRI) is frequently encountered after kidney transplantation and is a leading cause of acute renal failure. Aberrant gene expression and epigenetic regulation occur during the pathophysiology of IRI. In this study, we used reduced representation bisulfite sequencing to identify the DNA methylome of renal tissues during IRI and the sham-operated tissues in C57BL/6. The methylation status of approximately 1.29 million CpGs located in an average of 11554 CpG islands and 17113 promoters in genome was determined. Compared with sham-operated kidney, both acute and chronic IRI significantly decreased the genome-wide methylation level (1.1–1.8%) and the CpG methylation level in the promoter (0.4–0.5%), CpG island (0.5–1.3%), exon (1.3–1.9%), and intron (0.8–1.1%; all P<10−153). The promoters of 200, 191, and 79 genes were differentially methylated in the renal tissues at 24h, 7days, and at both the time points after IRI, respectively. Among the 79 genes, which were consistently epigenetically regulated at two time points, 18 genes (22.8%) showed differential expression after IRI in a previous study of renal expression. We validated the promoter methylation status and expression of five out of the 18 genes, including 2700049A03Rik, Ccr9, Fgd2, Pfkfb3, and Sdc4 in an independent renal tissue cohort. We found that all the five genes exhibited altered methylation of promoter (P=0.009–0.0001) following renal injury. The promoter methylation of 2700049A03Rik and Ccr9 was negatively correlated with their mRNA expression in renal tissues (P<0.001 and P<0.0001, respectively). Our study not only demonstrated a genome-wide DNA methylation pattern in the IR-injured renal tissue for the first time, but also indicated that the regulation of promoter methylation is an important mechanism underlying persistent alteration of gene expression.

Methylomic profiling of cortex samples from completed suicide cases implicates a role for PSORS1C3 in major depression and suicide

Major depressive disorder (MDD) represents a major social and economic health issue and constitutes a major risk factor for suicide. The molecular pathology of suicidal depression remains poorly understood, although it has been hypothesised that regulatory genomic processes are involved in the pathology of both MDD and suicidality. In this study, genome-wide patterns of DNA methylation were assessed in depressed suicide completers (n=20) and compared with non-psychiatric, sudden-death controls (n=20) using tissue from two cortical brain regions (Brodmann Area 11 (BA11) and Brodmann Area 25 (BA25)). Analyses focused on identifying differentially methylated regions (DMRs) associated with suicidal depression and epigenetic variation were explored in the context of polygenic risk scores for major depression and suicide. Weighted gene co-methylation network analysis was used to identify modules of co-methylated loci associated with depressed suicide completers and polygenic burden for MDD and suicide attempt. We identified a DMR upstream of the PSORS1C3 gene, subsequently validated using bisulfite pyrosequencing and replicated in a second set of suicide samples, which is characterised by significant hypomethylation in both cortical brain regions in MDD suicide cases. We also identified discrete modules of co-methylated loci associated with polygenic risk burden for suicide attempt, but not major depression. Suicide-associated co-methylation modules were enriched among gene networks implicating biological processes relevant to depression and suicidality, including nervous system development and mitochondria function. Our data suggest that there are coordinated changes in DNA methylation associated with suicide that may offer novel insights into the molecular pathology associated with depressed suicide completers.

DNA Methylation Profiles of Blood Cells Are Distinct between Early-Onset Obese and Control Individuals.

Obesity is a highly prevalent, chronic disorder that has been increasing in incidence in young patients. Both epigenetic and genetic aberrations may play a role in the pathogenesis of obesity. Therefore, in-depth epigenomic and genomic analyses will advance our understanding of the detailed molecular mechanisms underlying obesity and aid in the selection of potential biomarkers for obesity in youth. Here, we performed microarray-based DNA methylation and gene expression profiling of peripheral white blood cells obtained from six young, obese individuals and six healthy controls. We observed that the hierarchical clustering of DNA methylation, but not gene expression, clearly segregates the obese individuals from the controls, suggesting that the metabolic disturbance that occurs as a result of obesity at a young age may affect the DNA methylation of peripheral blood cells without accompanying transcriptional changes. To examine the genome-wide differences in the DNA methylation profiles of young obese and control individuals, we identified differentially methylated CpG sites and investigated their genomic and epigenomic contexts. The aberrant DNA methylation patterns in obese individuals can be summarized as relative gains and losses of DNA methylation in gene promoters and gene bodies, respectively. We also observed that the CpG islands of obese individuals are more susceptible to DNA methylation compared to controls. Our pilot study suggests that the genome-wide aberrant DNA methylation patterns of obese individuals may advance not only our understanding of the epigenomic pathogenesis but also early screening of obesity in youth.