Epigenetics of Neural Development
An enlightening Point of View by Yeo et al. in this issue of Epigenetics highlights recent finding in the area of environmental toxicology and developmental neuroscience relating to toxicity mechanisms of bisphenol A (BPA). The authors summarize recent findings, including their discovery that BPA disrupts the chloride shift in a sex-specific manner via epigenetics mechanisms.
Psychotropic Induced Obesity: A Role for Epigenetics
Psychotropic agents are notorious for their ability to increase fat mass in psychiatric patients. As epigenetic programs have a prominent role in the cell fate commitments and differentiation required for adipogenesis and the epigenetic effects on energy metabolism are well supported by several genetic models, Chase and Sharma propose that several psychotropics may utilize a common epigenetic effector path causing an increase in adipogenesis or reduction in energy metabolism. Recently, G protein-coupled signaling cascades have been shown to directly modify epigenetic regulatory enzymes, implicating surface receptor activity by psychotropic medications and pointing to potential therapeutic targets. The potential therapeutic implications are also suggested by the effects of the clinically approved antidepressant tranylcypromine, a histone demethylase inhibitor, which has impressive therapeutic effects on metabolism in the obese phenotype.
HumanMethylation450 BeadChip: Reducing Technical Variability
Analysis of DNA methylation is of increasing interest for epidemiologic studies examining disease etiology and impacts of environmental exposures. The Infinium HumanMethylation450 BeadChip (450K) has become a popular tool to characterize the DNA methylome. For large-scale studies, minimizing technical variability and potential bias is of utmost importance. Yousefi et al. have evaluated the performance of several existing and novel color channel normalizations designed to reduce technical variability and batch effects in 450K analyses from a large population study. The authors performed comparative assessment of 10 normalization procedures, including the GenomeStudio Illumina procedure, the lumi smooth quantile approach, and the newly proposed all samples normalization (ASMN). They concluded that the ASMN procedure can improve existing color channel normalization, especially for large epidemiologic studies, and can be successfully implemented to enhance a 450K DNA methylation data pipeline.
DNA Hypermethylation as Indicator of Gastric Premalignant Processes
In gastric cancers and premalignant lesions of the gastric mucosae, DNA methylation changes in specific genes have been observed. In this issue of Epigenetics, Schneider et al. have quantitatively evaluated DNA methylation in tumors, non-tumor gastric mucosae, and gastric biopsies at the promoters of 5 genes (EN1, PCDH10, RSPO2, ZIC1, and ZNF610) that show methylation alterations that discriminate gastric cancers from non-tumor epithelia. The authors report that for these genes, levels of methylation in precancerous lesions are related to Helicobacter pylori virulence, geographic region and measures of chronic inflammation. These genes seem predisposed to sustain significant quantitative changes in DNA methylation at early stages of the gastric precancerous process.
SUMOylation of the KDM5B Histone Lysine Demethylase
The histone lysine demethylase KDM5B plays key roles in gene repression by demethylating H3K4me3, a modification commonly found at the promoter region of actively transcribed genes. KDM5B is known to regulate the expression of genes involved in cell cycle progression. Now, Bueno and Richard report that KDM5B is SUMOylated at two lysine residues and that the ectopic expression of the hPC2 SUMO E3 ligase enhances this SUMOylation. Interestingly, the levels of KDM5B and its SUMOylated forms are regulated during the cell cycle. The authors propose that SUMOylation by hPC2 regulates the activity of KDM5B.
Site-Specific Methylated Reporters for the Analysis of DNA Methylation
In this issue of Epigenetics, Han et al. describe a site-specific DNA methylation method that uses synthetically methylated primers and high fidelity PCR coupled with ligation of reporter constructs. The authors used this method to introduce methylated cytosines into fragments of the respective DAPK and RASSF1A promoters that had been cloned into luciferase reporters. Their results suggest that site-specific DNA methylation of a reporter construct can be used for functional analysis of commonly observed gene promoter methylation patterns and that the reporter system contains key elements of the endogenous chromatin machinery.
Epigenome-Wide Profiling of DNA Methylation in T- and B-Lymphocytes
Approaches that analyze methylation in mixed cell population from whole blood may preclude the identification of cell type-specific methylation, which may subsequently bias identification of disease-specific changes. To address this problem, Glossop et al. conducted genome-wide DNA methylation profiling using HumanMethylation450 BeadChips to identify differences within matched pairs of T-lymphocytes and B-lymphocytes isolated from peripheral blood. Hierarchical clustering revealed perfect segregation of samples into two distinct clusters based on cell type. Differentially methylated genes showed enrichment for biological functions/pathways associated with leukocytes and T-lymphocytes. This work therefore shows that T-lymphocytes and B-lymphocytes possess intrinsic differences in DNA methylation within a restricted set of functionally related genes, providing a foundation for investigating DNA methylation in diseases in which these cell types play important and distinct roles.
Identifying Biomarkers in Ewing Sarcoma
In order to identify effective biomarkers in Ewing sarcoma (ES) on a genome-wide scale, Alholle et al. used a functionally proven epigenetic approach, in which gene expression was induced in ES cell lines by treatment with a demethylating agent followed by hybridization onto high density gene expression microarrays. After following a strict selection criterion, eight genes expressed in human bone marrow-derived mesenchymal stem cells showed methylation frequencies of > 20% in ES tumors and hypermethylation was associated with transcriptional silencing. In addition, six of those genes also showed high methylation frequency in osteosarcomas. These genes may provide insights into bone cancer tumorigenesis and be useful in the development of epigenetic biomarkers for prognosis and detection of these rare tumor types.
Transcriptional Enhancement by Targeted Promoter Demethylation
The ability to specifically reactivate epigenetically silenced genes would have great utility in experimental studies and potential therapeutic value. In this issue, Gregory et al. describe the specific targeting of thymidine DNA glycosylase (TDG), an enzyme involved in the mechanism of methyl cytosine demethylation, to the promoter of Nos2, a gene silenced by methylation in fibroblasts, using artificial zinc finger DNA binding domains. Individual targeted TDG constructs had a small effect on Nos2 expression and methylation, but simultaneous targeting of a quartet of TDG constructs significantly restored responsiveness to LPS and IFN stimuli in association with marked cytosine demethylation at the promoter and CpG island; catalytically inactive TDG complexes had no effect. This study shows highly specific, directed reactivation of a single, silenced gene by targeting of a demethylase to the promoter.
Novel Candidate Biomarker Genes in Cervical Cancer
Oncogenic human papilloma viruses are associated with precancerous cervical lesions and development of cervical cancer. As DNA methylation signatures of the host genome in normal, precancerous and cervical cancer tissue may indicate tissue-specific perturbation in carcinogenesis, Farkas et al. aimed to identify new candidate genes that were differentially methylated in squamous cell carcinoma compared with DNA samples from cervical intraepithelial neoplasia grade 3 (CIN3) and normal cervical scrapes. Their results showed an extensive differential methylation signature in cervical cancer compared with the CIN3 or normal cervical tissues. These insights into the functional role of DNA methylome alterations in cervical cancer could be clinically applicable in diagnostics and prognostics, and may guide the development of new epigenetic therapies.
The “ImmunoCRIT” as a Marker for Solid Tumor Aggressiveness
The adaptive immune system is involved in tumor establishment and aggressiveness. Tumors of the ovaries, an immune-privileged organ, spread via transceolomic routes and rarely to distant organs. This is contrary to tumors of non-immune privileged organs, which often disseminate hematogenously to distant organs. Epigenetics-based immune cell quantification allows direct comparison of the immune status in benign and malignant tissues and in blood. In this issue of Epigenetics, Türbachova et al. introduce the “cellular ratio of immune tolerance” (immunoCRIT) as defined by the ratio of regulatory T cells to total T lymphocytes. The immunoCRIT was analyzed on benign tissue samples of colorectal, bronchial, renal and ovarian origin as well as in samples from primary colorectal, bronchial, mammary and ovarian cancers. ImmunoCRIT is strongly increased in all cancerous tissues and gradually augmented strictly dependent on tumor aggressiveness. The authors propose that the immunoCRIT is a powerful marker for tumor aggressiveness and disease dissemination.
Integrated Analysis of Genome-Wide Genetic and Epigenetic Association Data
Many human diseases are multifactorial, involving multiple genetic and environmental factors impacting on one or more biological pathways. Genome-wide genetic and epigenetic association studies have been conducted for different diseases and traits, but no clear associations are usually achieved between the genomic loci and the biological pathways. In this issue, Ke et al. describe a protocol for the integrated analysis of genome-wide genetic and epigenetic data based on permutation of a sum statistic for the combined effects in a locus or pathway. The method has been applied to published type 1 diabetes (T1D) genome-wide- and epigenome-wide-association studies data to identify genomic loci and biological pathways that are associated with T1D genetically and epigenetically. Through combined analysis, the authors identified novel loci and pathways, which could add to our understanding of disease mechanisms of T1D as well as complex diseases in general.