Genomic DNA Methylation Patterns Research Articles

Two approaches reveal a new paradigm of \u2018switchable or genetics-influenced allele-specific DNA methylation\u2019 with potential in human disease

Imprinted genes are vulnerable to environmental influences during early embryonic development, thereby contributing to the onset of disease in adulthood. Monoallelic methylation at several germline imprints has been reported as DNMT1-dependent. However, which of these two epigenetic attributes, DNMT1-dependence or allelic methylation, renders imprinted genes susceptible to environmental stressors has not been determined. Herein, we developed a new approach, referred to as NORED, to identify 2468 DNMT1-dependent DNA methylation patterns in the mouse genome. We further developed an algorithm based on a genetic variation-independent approach (referred to as MethylMosaic) to detect 2487 regions with bimodal methylation patterns. Two approaches identified 207 regions, including known imprinted germline allele-specific methylation patterns (ASMs), that were both NORED and MethylMosaic regions. Examination of methylation in four independent mouse embryonic stem cell lines shows that two regions identified by both NORED and MethylMosaic (Hcn2 and Park7) did not display parent-of-origin-dependent allelic methylation. In these four F1 hybrid cell lines, genetic variation in Cast allele at Hcn2 locus introduces a transcription factor binding site for MTF-1 that may predispose Cast allelic hypomethylation in a reciprocal cross with either C57 or 129 strains. In contrast, each allele of Hcn2 ASM in J1 inbred cell line and Park7 ASM in four F1 hybrid cell lines seems to exhibit similar propensity to be either hypo- or hypermethylated, suggesting a ‘random, switchable’ ASM. Together with published results, our data on ASMs prompted us to propose a hypothesis of regional ‘autosomal chromosome inactivation (ACI)’ that may control a subset of autosomal genes. Therefore, our results open a new avenue to understand monoallelic methylation and provide a rich resource of candidate genes to examine in environmental and nutritional exposure models.

Blood DNA methylation pattern is altered in mesial temporal lobe epilepsy

Mesial temporal lobe epilepsy (MTLE) is a common epileptic disorder; little is known whether it is associated with peripheral epigenetic changes. Here we compared blood whole genomic DNA methylation pattern in MTLE patients (n = 30) relative to controls (n = 30) with the Human Methylation 450 K BeadChip assay, and explored genes and pathways that were differentially methylated using bioinformatics profiling. The MTLE and control groups showed significantly different (P < 1.03e-07) DNA methylation at 216 sites, with 164 sites involved hyper- and 52 sites hypo- methylation. Two hyper- and 32 hypo-methylated sites were associated with promoters, while 87 hyper- and 43 hypo-methylated sites corresponded to coding regions. The differentially methylated genes were largely related to pathways predicted to participate in anion binding, oxidoreductant activity, growth regulation, skeletal development and drug metabolism, with the most distinct ones included SLC34A2, CLCN6, CLCA4, CYP3A43, CYP3A4 and CYP2C9. Among the MTLE patients, panels of genes also appeared to be differentially methylated relative to disease duration, resistance to anti-epileptics and MRI alterations of hippocampal sclerosis. The peripheral epigenetic changes observed in MTLE could be involved in certain disease-related modulations and warrant further translational investigations.

Genome-wide DNA methylation reprogramming in response to inorganic arsenic links inhibition of CTCF binding, DNMT expression and cellular transformation

Chronic low dose inorganic arsenic (iAs) exposure leads to changes in gene expression and epithelial-to-mesenchymal transformation. During this transformation, cells adopt a fibroblast-like phenotype accompanied by profound gene expression changes. While many mechanisms have been implicated in this transformation, studies that focus on the role of epigenetic alterations in this process are just emerging. DNA methylation controls gene expression in physiologic and pathologic states. Several studies show alterations in DNA methylation patterns in iAs-mediated pathogenesis, but these studies focused on single genes. We present a comprehensive genome-wide DNA methylation analysis using methyl-sequencing to measure changes between normal and iAs-transformed cells. Additionally, these differential methylation changes correlated positively with changes in gene expression and alternative splicing. Interestingly, most of these differentially methylated genes function in cell adhesion and communication pathways. To gain insight into how genomic DNA methylation patterns are regulated during iAs-mediated carcinogenesis, we show that iAs probably targets CTCF binding at the promoter of DNA methyltransferases, regulating their expression. These findings reveal how CTCF binding regulates DNA methyltransferase to reprogram the methylome in response to an environmental toxin.

Observation of Pneumococcal Phase Variation in Colony Morphology.

Streptococcus pneumoniae (pneumococcus) is an important human pathogen that causes pneumonia, meningitis, sepsis, and otitis media. This bacterium normally resides in the nasopharynx as a commensal, but sometimes disseminates to sterile sites of humans and causes local or systemic inflammation. This biphasic behavior of S. pneumoniae is correlated with a reversible switch between the opaque and transparent colony forms on agar plates, a phenomenon referred to as phase variation. The opaque variants appear to be more virulent in animal models of bacteremia but are deficient in nasopharyngeal colonization animal models. In contrast, the transparent variants display higher levels of nasopharyngeal colonization but relatively lower virulence in animal models. We have recently demonstrated that pneumococcal phase variation between these two colony types is caused by a reversible switch of genome DNA methylation (or epigenetic) patterns, which is driven by DNA inversions in the DNA methyltransferase genes. Observation of colony morphology is a simple and useful method to differentiate colonies with different characteristics, such as size, color, and opacity. This protocol describes how to study pneumococcal phase variation in colony morphology with a dissection microscope.

Nutritional programming of accelerated puberty in heifers: alterations in DNA methylation in the arcuate nucleus.

High rates of body weight gain during the juvenile period appear to program molecular events within the hypothalamus, leading to advancement of puberty. Methylation of DNA, an epigenetic mechanism that controls gene expression, is associated with metabolic programming events and is proposed to play a role in the pubertal process. In this study, DNA methylation was assessed in genomic DNA obtained from the arcuate nucleus (ARC) of juvenile heifers fed to gain body weight at low (0.5 kg/d; low-gain, LG, n = 4) or high (1 kg/d; high-gain, HG, n = 4) rates from 4.5 to 8.5 mo of age (earliest puberty expected at 9 mo of age in HG heifers). Using a custom-designed oligonucleotide array targeted to imprinted genes and genes associated with nutritional inputs and the control of puberty, a comparative-genomic-hybridization array was used to identify differentially methylated regions between LG and HG heifers. Differential methylation of genomic regions associated with altered mRNA expression was observed for genes whose activity has been reported to be involved in the modulation of growth and metabolism (GHR) and puberty (HMGA2). Hence, increased rates of body weight gain during the juvenile period alter the methylation pattern of genomic DNA obtained from the ARC and these changes may be involved in programming the age at puberty in heifers.

The nasal methylome and childhood atopic asthma

Given the strong environmental influence on both epigenetic marks and allergic asthma in children, the epigenetic alterations in respiratory epithelia might provide insight into allergic asthma. We sought to identify DNA methylation and gene expression changes associated with childhood allergic persistent asthma. We compared genomic DNA methylation patterns and gene expression in African American children with persistent atopic asthma (n=36) versus healthy control subjects (n=36). Results were validated in an independent population of asthmatic children (n=30) by using a shared healthy control population (n=36) and in an independent population of white adult atopic asthmatic patients (n=12) and control subjects (n=12). We identified 186 genes with significant methylation changes, differentially methylated regions or differentially methylated probes, after adjustment for age, sex, race/ethnicity, batch effects, inflation, and multiple comparisons. Genes differentially methylated included those with established roles in asthma and atopy and genes related to extracellular matrix, immunity, cell adhesion, epigenetic regulation, and airflow obstruction. The methylation changes were substantial (median, 9.5%; range, 2.6% to 29.5%). Hypomethylated and hypermethylated genes were associated with increased and decreased gene expression, respectively (P<2.8×10-6 for differentially methylated regions and P<7.8×10-10 for differentially methylated probes). Quantitative analysis in 53 differentially expressed genes demonstrated that 32 (60%) have significant methylation-expression relationships within 5 kb of the gene. Ten loci selected based on the relevance to asthma, magnitude of methylation change, and methylation-expression relationships were validated in an independent cohort of children with atopic asthma. Sixty-seven of 186 genes also have significant asthma-associated methylation changes in nasal epithelia of adult white asthmatic patients. Epigenetic marks in respiratory epithelia are associated with allergic asthma and gene expression changes in inner-city children.

DNA Methylation Variation Trends during the Embryonic Development of Chicken.

The embryogenesis period is critical for epigenetic reprogramming and is thus of great significance in the research field of poultry epigenetics for elucidation of the trends in DNA methylation variations during the embryonic development of birds, particularly due to differences in embryogenesis between birds and mammals. Here, we first examined the variations in genomic DNA methylation during chicken embryogenesis through high-performance liquid chromatography using broilers as the model organism. We then identified the degree of DNA methylation of the promoters and gene bodies involved in two specific genes (IGF2 and TNF-α) using the bisulfite sequencing polymerase chain reaction method. In addition, we measured the expression levels of IGF2, TNF-α and DNA methyltransferase (DNMT) 1, 3a and 3b. Our results showed that the genomic DNA methylation levels in the liver, heart and muscle increased during embryonic development and that the methylation level of the liver was significantly higher in mid-anaphase. In both the muscle and liver, the promoter methylation levels of TNF-α first increased and then decreased, whereas the gene body methylation levels remained lower at embryonic ages E8, 11 and 14 before increasing notably at E17. The promoter methylation level of IGF2 decreased persistently, whereas the methylation levels in the gene body showed a continuous increase. No differences in the expression of TNF-α were found among E8, 11 and 14, whereas a significant increase was observed at E17. IGF2 showed increasing expression level during the examined embryonic stages. In addition, the mRNA and protein levels of DNMTs increased with increasing embryonic ages. These results suggest that chicken shows increasing genomic DNA methylation patterns during the embryonic period. Furthermore, the genomic DNA methylation levels in tissues are closely related to the genes expression levels, and gene expression may be simultaneously regulated by promoter hypomethylation and gene body hypermethylation.

Etiology matters - Genomic DNA Methylation Patterns in Three Rat Models of Acquired Epilepsy.

This study tested the hypothesis that acquired epileptogenesis is accompanied by DNA methylation changes independent of etiology. We investigated DNA methylation and gene expression in the hippocampal CA3/dentate gyrus fields at 3 months following epileptogenic injury in three experimental models of epilepsy: focal amygdala stimulation, systemic pilocarpine injection, or lateral fluid-percussion induced traumatic brain injury (TBI) in rats. In the models studies, DNA methylation and gene expression profiles distinguished controls from injured animals. We observed consistent increased methylation in gene bodies and hypomethylation at non-genic regions. We did not find a common methylation signature in all three different models and few regions common to any two models. Our data provide evidence that genome-wide alteration of DNA methylation signatures is a general pathomechanism associated with epileptogenesis and epilepsy in experimental animal models, but the broad pathophysiological differences between models (i.e. pilocarpine, amygdala stimulation, and post-TBI) are reflected in distinct etiology-dependent DNA methylation patterns.

Genomic DNA Methylation Analyses Reveal the Distinct Profiles in Castor Bean Seeds with Persistent Endosperms.

Investigations of genomic DNA methylation in seeds have been restricted to a few model plants. The endosperm genomic DNA hypomethylation has been identified in angiosperm, but it is difficult to dissect the mechanism of how this hypomethylation is established and maintained because endosperm is ephemeral and disappears with seed development in most dicots. Castor bean (Ricinus communis), unlike Arabidopsis (Arabidopsis thaliana), endosperm is persistent throughout seed development, providing an excellent model in which to dissect the mechanism of endosperm genomic hypomethylation in dicots. We characterized the DNA methylation-related genes encoding DNA methyltransferases and demethylases and analyzed their expression profiles in different tissues. We examined genomic methylation including CG, CHG, and CHH contexts in endosperm and embryo tissues using bisulfite sequencing and revealed that the CHH methylation extent in endosperm and embryo was, unexpectedly, substantially higher than in previously studied plants, irrespective of the CHH percentage in their genomes. In particular, we found that the endosperm exhibited a global reduction in CG and CHG methylation extents relative to the embryo, markedly switching global gene expression. However, CHH methylation occurring in endosperm did not exhibit a significant reduction. Combining with the expression of 24-nucleotide small interfering RNAs (siRNAs) mapped within transposable element (TE) regions and genes involved in the RNA-directed DNA methylation pathway, we demonstrate that the 24-nucleotide siRNAs played a critical role in maintaining CHH methylation and repressing the activation of TEs in persistent endosperm development. This study discovered a novel genomic DNA methylation pattern and proposes the potential mechanism occurring in dicot seeds with persistent endosperm.

Poly(ADP-ribosyl)ation of Methyl CpG Binding Domain Protein 2 Regulates Chromatin Structure

The epigenetic information encoded in the genomic DNA methylation pattern is translated by methylcytosine binding proteins like MeCP2 into chromatin topology and structure and gene activity states. We have shown previously that the MeCP2 level increases during differentiation and that it causes large-scale chromatin reorganization, which is disturbed by MeCP2 Rett syndrome mutations. Phosphorylation and other posttranslational modifications of MeCP2 have been described recently to modulate its function. Here we show poly(ADP-ribosyl)ation of endogenous MeCP2 in mouse brain tissue. Consequently, we found that MeCP2 induced aggregation of pericentric heterochromatin and that its chromatin accumulation was enhanced in poly(ADP-ribose) polymerase (PARP) 1−/− compared with wild-type cells. We mapped the poly(ADP-ribosyl)ation domains and engineered MeCP2 mutation constructs to further analyze potential effects on DNA binding affinity and large-scale chromatin remodeling. Single or double deletion of the poly(ADP-ribosyl)ated regions and PARP inhibition increased the heterochromatin clustering ability of MeCP2. Increased chromatin clustering may reflect increased binding affinity. In agreement with this hypothesis, we found that PARP-1 deficiency significantly increased the chromatin binding affinity of MeCP2 in vivo. These data provide novel mechanistic insights into the regulation of MeCP2-mediated, higher-order chromatin architecture and suggest therapeutic opportunities to manipulate MeCP2 function.

MET18 Connects the Cytosolic Iron-Sulfur Cluster Assembly Pathway to Active DNA Demethylation in Arabidopsis.

DNA demethylation mediated by the DNA glycosylase ROS1 helps determine genomic DNA methylation patterns and protects active genes from being silenced. However, little is known about the mechanism of regulation of ROS1 enzymatic activity. Using a forward genetic screen, we identified an anti-silencing (ASI) factor, ASI3, the dysfunction of which causes transgene promoter hyper-methylation and silencing. Map-based cloning identified ASI3 as MET18, a component of the cytosolic iron-sulfur cluster assembly (CIA) pathway. Mutation in MET18 leads to hyper-methylation at thousands of genomic loci, the majority of which overlap with hypermethylated loci identified in ros1 and ros1dml2dml3 mutants. Affinity purification followed by mass spectrometry indicated that ROS1 physically associates with MET18 and other CIA components. Yeast two-hybrid and split luciferase assays showed that ROS1 can directly interact with MET18 and another CIA component, AE7. Site-directed mutagenesis of ROS1 indicated that the conserved iron-sulfur motif is indispensable for ROS1 enzymatic activity. Our results suggest that ROS1-mediated active DNA demethylation requires MET18-dependent transfer of the iron-sulfur cluster, highlighting an important role of the CIA pathway in epigenetic regulation.

Valproate administration to mice increases hippocampal p21 expression by altering genomic DNA methylation.

Although valproate (VPA) is used widely in the treatment of bipolar mood disorder and epilepsy, the precise mechanism of action in the brain remains elusive. In this study, we investigated the effects of subchronic VPA administrations on the expression of the cyclin-dependent kinase inhibitor (Cdkn) family in the hippocampus of adult mice. The administration of VPA specifically increased hippocampal p21 expression involving both mRNA and protein levels, but other members of the Cdkn family were not affected. We identified two CpG islands in the p21 gene regulatory region, located distal and proximal to the transcription start site. VPA altered genomic DNA methylation patterns in the distal region, but not in the proximal promoter region. However, no change was found in DNA methyltransferase (Dnmt) 1 or Dnmt3a protein levels, suggesting an involvement in active demethylation mechanisms. These findings suggest that VPA alters the gene expression of cell cycle regulators by modulating promoter DNA methylation, and this resulted in altered hippocampal cell proliferation. These findings promote understanding of the actions of VPA in the brain.

Chromatin modifications and genomic contexts linked to dynamic DNA methylation patterns across human cell types.

DNA methylation is related closely to sequence contexts and chromatin modifications; however, their potential differences in different genomic regions across cell types remain largely unexplored. We used publicly available genome-scale DNA methylation and histone modification profiles to study their relationships among different genomic regions in human embryonic stem cells (H1), H1-derived neuronal progenitor cultured cells (NPC), and foetal fibroblasts (IMR90) using the Random forests classifier. Histone modifications achieved high accuracy in modelling DNA methylation patterns on a genome scale in the three cell types. The inclusion of sequence features helped improve accuracy only in non-promoter regions of IMR90. Furthermore, the top six feature combinations obtained by mean decrease Gini were important indicators of different DNA methylation patterns, suggesting that H3K4me2 and H3K4me3 are important indicators that are independent of genomic regions and cell types. H3K9me3 was IMR90-specific and exhibited a genomic region-specific correlation with DNA methylation. Variations of essential chromatin modification signals may effectively discriminate changes of DNA methylation between H1 and IMR90. Genes with different co-variations of epigenetic marks exhibited genomic region-specific biological relevance. This study provides an integrated strategy to identify systematically essential epigenetic and genetic elements of genomic region-specific and cell type-specific DNA methylation patterns.

Genomic profiling of DNA methyltransferases reveals a role for DNMT3B in genic methylation.

DNA methylation is an epigenetic modification associated with transcriptional repression of promoters and is essential for mammalian development. Establishment of DNA methylation is mediated by the de novo DNA methyltransferases DNMT3A and DNMT3B, whereas DNMT1 ensures maintenance of methylation through replication. Absence of these enzymes is lethal, and somatic mutations in these genes have been associated with several human diseases. How genomic DNA methylation patterns are regulated remains poorly understood, as the mechanisms that guide recruitment and activity of DNMTs in vivo are largely unknown. To gain insights into this matter we determined genomic binding and site-specific activity of the mammalian de novo DNA methyltransferases DNMT3A and DNMT3B. We show that both enzymes localize to methylated, CpG-dense regions in mouse stem cells, yet are excluded from active promoters and enhancers. By specifically measuring sites of de novo methylation, we observe that enzymatic activity reflects binding. De novo methylation increases with CpG density, yet is excluded from nucleosomes. Notably, we observed selective binding of DNMT3B to the bodies of transcribed genes, which leads to their preferential methylation. This targeting to transcribed sequences requires SETD2-mediated methylation of lysine 36 on histone H3 and a functional PWWP domain of DNMT3B. Together these findings reveal how sequence and chromatin cues guide de novo methyltransferase activity to ensure methylome integrity.

Genome organization and DNA methylation patterns of B chromosomes in the red fox and Chinese raccoon dogs.

The molecular structure of B chromosomes (Bs) is relatively well studied. Previous research demonstrates that Bs of various species usually contain two types of repetitive DNA sequences, satellite DNA and ribosomal DNA, but Bs also contain genes encoding histone proteins and many others. However, many questions remain regarding the origin and function of these chromosomes. Here, we focused on the comparative cytogenetic characteristics of the red fox and Chinese raccoon dog B chromosomes with particular attention to the distribution of repetitive DNA sequences and their methylation status. We confirmed that the small Bs of the red fox show a typical fluorescent telomeric distal signal, whereas medium-sized Bs of the Chinese raccoon dog were characterized by clusters of telomeric sequences along their length. We also found different DNA methylation patterns for the B chromosomes of both species. Therefore, we concluded that DNA methylation may maintain the transcriptional inactivation of DNA sequences localized to B chromosomes and may prevent genetic unbalancing and several negative phenotypic effects.

Comparison of genomic DNA methylation pattern among septic and non-septic newborns - An epigenome wide association study.

DNA methylation is the current strategy in the field of biomarker discovery due to its prognostic efficiency. Its role in prognosis and early diagnosis has been recognized in various types of cancer. Sepsis still remains one of the major causes of neonatal mortality. Delay in diagnosis of sepsis leads to treatment difficulties and poor outcome. In this study, we have done an epigenome wide search to identify potential markers for prognosis of neonatal sepsis which may improve the treatment strategies. We analyzed the CpG methylation status in the epigenome of three septic and non-septic babies using Illumina Infinium HumanMethylation450K methylation microarray. The microarray data was analyzed with Illumina GenomeStudio v2011.1. After screening for biological and clinical significance, we found 81 differentially methylated CpGs located in 64 genes. Bioinformatic analysis using DAVID and GeneMania revealed a panel of differentially methylated protocadherin beta (PCDHB) genes that play vital role in leukocyte cell adhesion and Wnt signaling pathway. Apart, genes like CCS, DNAJA3, and DEGS2 were potentially hyper/hypo methylated which can be utilized in the development of novel biomarkers. This study will be helpful in exploring the role of DNA methylation in the pathophysiology of neonatal sepsis. The complete microarray data can be accessed from the public domain, Gene Expression Omnibus of NCBI (http://www.ncbi.nlm.nih.gov/geo/). The accession number is GSE58651.

Different genomic DNA methylation patterns between male and female adults of white-backed planthoppers Sogatella furcifera

DNA methylation plays a key role in gene regulation and phenotype variation in many organisms. The aim of this study was to survey the frequency and variation of cytosine methylation at CCGG sequences in adult male and female planthoppers Sogatella furcifera, a major rice pest in Asia, and to determine the occurrence of methylation changes associated with sexual dimorphism using methylation-sensitive amplification polymorphism. 1131 DNA fragments including CCGG sites were amplified using 36 pairs of selective primers: about 191 methylated bands were identified. In male planthoppers, we got a total of 581 bands, including 40 fully-methylated bands, 65 hemi-methylated bands and 476 none-methylated bands, so the fully-methylated ratio, hemi-methylated ratio and total methylated ratio were 6.88%, 11.19% and 18.07%, respectively. In the female planthopper, there were a total of 550 bands, including 44 fully-methylated bands, 42 hemi-methylated bands and 464 none-methylated bands. The fully-methylated ratio was 7.64% in female planthoppers, which was slightly higher than in the male planthoppers, however, the hemi-methylated ratio was lower (8.00%) in the female compared with the male planthopper. Altogether, 46 DNA bands displayed variable cytosine methylation patterns between male and female samples: 20 of them occurred only in male samples and 26 only in female samples. Thus, the genome methylation patterns are different between male and female adults. The results suggest that DNA methylation might be related to sexual differentiation and development in S. furcifera.

DNA methyltransferases 3a and 3b are differentially expressed in the early stages of a rat liver carcinogenesis model.

Carcinogenesis is driven by the accumulation of mutations and abnormal DNA methylation patterns, particularly the hypermethylation of tumor‑suppressor genes. Changes in genomic DNA methylation patterns are established by the DNA methyltransferases (DNMTs) family: DNMT1, DNMT3a and DNMT3b. The DNMTs are known to be overexpressed in tumors. However, when the DNMTs expression profile is altered in earlier stages of carcinogenesis remains to be elucidated. The resistant hepatocyte model (RHM) allows the analysis of the hepatocellular carcinoma (HCC) from the formation of altered cell foci to the appearance of tumors in rats. To investigate the DNMTs expression in this model, we first observed that timp3, rassf1a and p16 genes became methylated during cancer development by methylation‑specific PCR (MSP) and the bisulphate sequencing PCR (BSP) of timp3. The differential expression at the RNA and protein level of the three DNMTs was also assessed. dnmt1 expression was higher in tumors than in normal and early cancer stages. However, no evident overexpression of the enzyme was identified by immunohistochemistry. By contrast, DNMT3a and DNMT3b were consistently subexpressed in tumors. In the present study, we report a carcinogenesis model that does not feature the overexpression of DNMT1 but exhibits a transient expression of DNMT3a and DNMT3b.

Characteristics of cytosine methylation status and methyltransferase genes in the early development stage of cauliflower (Brassica oleracea L. var. botrytis)

DNA methylation is one of the most important epigenetic modifications involved in the development and differentiation in plants. Hypocotyl and cotyledon are the two major tissues of cauliflower (Brassica oleracea L. var. botrytis) seedlings. Both tissues show significantly different tissue specificity and regenerative abilities in vitro. However, the characteristics of DNA methylation modification and its roles in regulating the organ development in cauliflower remain largely unknown. In the present study, the DNA methylation status between the hypocotyl and cotyledon of cauliflower seedlings were analyzed. The results indicated that although the hypocotyl and cotyledon of cauliflower seedlings share the same genome, the genomic DNA methylation levels and patterns at CCGG sites were different. Compared with the cotyledon, the hypocotyl showed higher DNA methylation level, and more loci showing methylation pattern adjustments were also discovered. Twelve loci with changes of DNA methylation patterns were further explored. The quantitative expression analysis indicated that eight out of twelve sequenced fragments showed differential expression between the hypocotyl and cotyledon, of which the expression of six sequences was identified to be negative correlation with their DNA methylation status. In addition, three main DNA methyltransferase genes MET1, CMT3 and DRM were first explored in cauliflower. The results indicated that the expression of these three genes was closely associated with the different DNA methylation status in the hypocotyl and cotyledon. These findings provided more information to further explore the roles of DNA methylation modification in tissue differentiation and development of cauliflower.

Intergenerational genomic DNA methylation patterns in mouse hybrid strains.

BackgroundDNA methylation is a contributing factor to both rare and common human diseases, and plays a major role in development and gene silencing. While the variation of DNA methylation among individuals has been partially characterized, the degree to which methylation patterns are preserved across generations is still poorly understood. To determine the extent of methylation differences between two generations of mice we examined DNA methylation patterns in the livers of eight parental and F1 mice from C57BL/6J and DBA/2J mouse strains using bisulfite sequencing.ResultsWe find a large proportion of reproducible methylation differences between C57BL/6J and DBA/2J chromosomes in CpGs, which are highly heritable between parent and F1 mice. We also find sex differences in methylation levels in 396 genes, and 11% of these are differentially expressed between females and males. Using a recently developed approach to identify allelically methylated regions independently of genotypic differences, we identify 112 novel putative imprinted genes and microRNAs, and validate imprinting at the RNA level in 10 of these genes.ConclusionsThe majority of DNA methylation differences among individuals are associated with genetic differences, and a much smaller proportion of these epigenetic differences are due to sex, imprinting or stochastic intergenerational effects. Epigenetic differences can be a determining factor in heritable traits and should be considered in association studies for molecular and clinical traits, as we observed that methylation differences in the mouse model are highly heritable and can have functional consequences on molecular traits such as gene expression.