Cooperative activity of DNA methyltransferases for maintenance of symmetrical and non-symmetrical cytosine methylation in Arabidopsis thaliana

Maintenance of cytosine methylation in plants is controlled by three DNA methyltransferases. MET1 maintains CG methylation, and DRM1/2 and CMT3 act redundantly to enforce non-CG methylation. RPS, a repetitive hypermethylated DNA fragment from Petunia hybrida, attracts DNA methylation when transferred into Petunia or other species. In Arabidopsis thaliana, which does not contain any RPS homologues, RPS transgenes are efficiently methylated in all sequence contexts. To test which DNA methylation pathways regulate RPS methylation, we examined maintenance of RPS methylation in various mutant backgrounds. Surprisingly, CG methylation was lost in a drm1/2/cmt3 mutant, and non-CG methylation was almost completely eliminated in a met1 mutant. An unusual cooperative activity of all three DNA methyltransferases is therefore required for maintenance of both CG and non-CG methylation in RPS. Other unusual features of RPS methylation are the independence of its non-CG methylation from the RNA-directed DNA methylation (RdDM) pathway and the exceptional maintenance of methylation at a CC(m)TGG site in some epigenetic mutants. This is indicative of activity of a methylation system in plants that may have evolved from the DCM methylation system that controls CC(m)WGG methylation in bacteria. Our data suggest that strict separation of CG and non-CG methylation pathways does not apply to all target regions, and that caution is required in generalizing methylation data obtained for individual genomic regions.     

Genomic distribution and inter-sample variation of non-CpG methylation across human cell types

DNA methylation plays an important role in development and disease. The primary sites of DNA methylation in vertebrates are cytosines in the CpG dinucleotide context, which account for roughly three quarters of the total DNA methylation content in human and mouse cells. While the genomic distribution, inter-individual stability, and functional role of CpG methylation are reasonably well understood, little is known about DNA methylation targeting CpA, CpT, and CpC (non-CpG) dinucleotides. Here we report a comprehensive analysis of non-CpG methylation in 76 genome-scale DNA methylation maps across pluripotent and differentiated human cell types. We confirm non-CpG methylation to be predominantly present in pluripotent cell types and observe a decrease upon differentiation and near complete absence in various somatic cell types. Although no function has been assigned to it in pluripotency, our data highlight that non-CpG methylation patterns reappear upon iPS cell reprogramming. Intriguingly, the patterns are highly variable and show little conservation between different pluripotent cell lines. We find a strong correlation of non-CpG methylation and DNMT3 expression levels while showing statistical independence of non-CpG methylation from pluripotency associated gene expression. In line with these findings, we show that knockdown of DNMTA and DNMT3B in hESCs results in a global reduction of non-CpG methylation. Finally, non-CpG methylation appears to be spatially correlated with CpG methylation. In summary these results contribute further to our understanding of cytosine methylation patterns in human cells using a large representative sample set.     

Developmental changes in DNA methylation of pollen mother cells of David lily during meiotic prophase I

Epigenetic marks in the form of DNA methylation are involved in the development of germ cells and are important in the maintenance of fertility. However, the controlling system of the on-off switch for DNA methylation largely remains unclear. In this study, the extent of cytosine methylation during the meiotic prophase I in David lily is assessed using high pressure liquid chromatography to evaluate the DNA methylation rates. Comparing the degree of DNA methylation before, during, and after synizesis, both de novo methylation and demethylation occurred. Mainly the methylation level decreased by 21.3% (from 54.8 to 33.5%) during synizesis in the pollen mother cells. The developmental timing of genome-wide DNA methylation acquisition during pollen mother cell development is clarified in this paper. The relative amounts of 5-methyl-deoxycytidine of global methylation in leaf DNA in David lily were also higher than in other species reported.     

Association of ATRX with pericentric heterochromatin and the Y chromosome of neonatal mouse spermatogonia

Background  

Establishment of chromosomal cytosine methylation and histone methylation patterns are critical epigenetic modifications required for heterochromatin formation in the mammalian genome. However, the nature of the primary signal(s) targeting DNA methylation at specific genomic regions is not clear. Notably, whether histone methylation and/or chromatin remodeling proteins play a role in the establishment of DNA methylation during gametogenesis is not known. The chromosomes of mouse neonatal spermatogonia display a unique pattern of 5-methyl cytosine staining whereby centromeric heterochromatin is hypo-methylated whereas chromatids are strongly methylated. Thus, in order to gain some insight into the relationship between global DNA and histone methylation in the germ line we have used neonatal spermatogonia as a model to determine whether these unique chromosomal DNA methylation patterns are also reflected by concomitant changes in histone methylation.     

Methylation patterns of the human apoA-I/C-III/A-IV gene cluster in adult and embryonic tissues suggest dynamic changes in methylation during development.

We describe here a detailed analysis of the methylation patterns of the apoC-III and apoA-IV genes in adult and embryonic tissues. Together with previously reported data on the human apoA-I gene (4), the results presented here constitute a comprehensive study on the methylation pattern of the apoA-I/C-III/A-IV gene cluster. The two genes (apoC-III and apoA-IV) display tissue-specific methylation patterns that correlate with their activity. This gene-specific methylation pattern indicates that the apoA-I/C-III/A-IV gene cluster is not one entity with respect to methylation. The cluster is almost entirely methylated in tissues that do not express any of the genes; however, individual gene regions are unmethylated in the tissue of expression. A comparison of the observed methylation patterns in adult tissues with those in embryonic tissues suggests that the mature tissue-specific methylation patterns are a result of an interplay between demethylation and de novo methylation events in the embryo. These changes in DNA methylation include demethylation in the early embryo followed by de novo methylation at later stages. A second round of tissue-specific demethylation and methylation de novo occurs in the late embryo as well. Evidence presented here supports the idea that CpG islands are protected in general from methylation de novo by a built-in signal and not by CpG density per se.     

DNA甲基化微阵列

DNA甲基化微阵列是近年发展起来的高通量分析基因组水平DNA甲基化状态和模式的新型技术,已成为肿瘤表观遗传学组研究的重要工具之一。利用DNA甲基化微阵列研究某种疾病状态下异常甲基化的基因有利于进一步明确该疾病的表观遗传学异常机制,发现与之相关的表观遗传学标志物。现有的DNA甲基化微阵列主要包括CpG岛微阵列和甲基化寡聚核苷探针微阵列,根据已有的文献资料,较为详细地阐述了上述技术的原理、特点和适用范围,对于研究者根据自己的研究目的选择适当的DNA甲基化微阵列技术具有一定的指导价值。     

DNA methylation in plants: relationship to small RNAs and histone modifications, and functions in transposon inactivation

DNA methylation is a type of epigenetic marking that strongly influences chromatin structure and gene expression in plants and mammals. Over the past decade, DNA methylation has been intensively investigated in order to elucidate its control mechanisms. These studies have shown that small RNAs are involved in the induction of DNA methylation, that there is a relationship between DNA methylation and histone methylation, and that the base excision repair pathway has an important role in DNA demethylation. Some aspects of DNA methylation have also been shown to be shared with mammals, suggesting that the regulatory pathways are, in part at least, evolutionarily conserved. Considerable progress has been made in elucidating the mechanisms that control DNA methylation; however, many aspects of the mechanisms that read the information encoded by DNA methylation and mediate this into downstream regulation remain uncertain, although some candidate proteins have been identified. DNA methylation has a vital role in the inactivation of transposons, suggesting that DNA methylation is a key factor in the evolution and adaptation of plants.     

DNA methylation: the future of crime scene investigation?

Proper detection and subsequent analysis of biological evidence is crucial for crime scene reconstruction. The number of different criminal acts is increasing rapidly. Therefore, forensic geneticists are constantly on the battlefield, trying hard to find solutions how to solve them. One of the essential defensive lines in the fight against the invasion of crime is relying on DNA methylation. In this review, the role of DNA methylation in body fluid identification and other DNA methylation applications are discussed. Among other applications of DNA methylation, age determination of the donor of biological evidence, analysis of the parent-of-origin specific DNA methylation markers at imprinted loci for parentage testing and personal identification, differentiation between monozygotic twins due to their different DNA methylation patterns, artificial DNA detection and analyses of DNA methylation patterns in the promoter regions of circadian clock genes are the most important ones. Nevertheless, there are still a lot of open chapters in DNA methylation research that need to be closed before its final implementation in routine forensic casework.     

Regulation of imprinted DNA methylation

DNA methylation is an essential enzymatic modification in mammals. This common epigenetic mark occurs predominantly at the fifth carbon of cytosines within the palindromic dinucleotide 5'-CpG-3'. The majority of methylated CpGs are located within repetitive elements including centromeric repeats, satellite sequences and gene repeats encoding ribosomal RNAs. CpG islands, frequently located at the 5' end of genes, are typically unmethylated. DNA methylation also occurs at imprinted genes which exhibit parent-of-origin-specific patterns of methylation and expression. Imprinted methylation at differentially methylated domains (DMDs) is one of the regulatory mechanisms controlling the allele-specific expression of imprinted genes. Proper control of DNA methylation is needed for normal development and loss of methylation control can contribute to initiation and progression of tumorigenesis (reviewed in Plass and Soloway, 2002). Because patterns of imprinted DNA methylation are highly reproducible, imprinted loci make useful models for studying regulation of DNA methylation and may provide insights into how this regulation goes awry in cancer. Here, we review what is currently known about the mechanisms regulating imprinted DNA methylation. We will focus on cis-acting DNA sequences, trans-acting protein factors and the possible involvement of RNAs in control of imprinted DNA methylation.     

Widely variable endogenous retroviral methylation levels in human placenta

It is generally assumed that transposable elements, including endogenous retroviruses (ERVs), are silenced by DNA methylation/chromatin structure in mammalian cells. However, there have been very few experimental studies to examine the methylation status of human ERVs. In this study, we determined and compared the methylation status of the 5′ long terminal repeats (LTRs) of different copies of the human endogenous retrovirus (HERV) family HERV-E, which are inserted in various genomic contexts. We found that three HERV-E LTRs which function as alternative gene promoters in placenta are unmethylated in that tissue but heavily methylated in blood cells, where these LTRs are not active promoters. This difference is not solely due to global hypomethylation in placenta, since two general measures of methylation levels of HERV-E and HERV-K LTRs suggest only 10–15% lower overall HERV methylation in placenta compared to blood. Comparisons between methylation levels of the LTR-derived gene promoters and six random HERV-E LTRs in placenta showed that the former display significantly lower methylation levels than random LTRs. Moreover, the differences in methylation between LTRs cannot always be explained by their genomic environment, since methylation of flanking sequences can be very different from methylation of the LTR itself.