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Epigenetic systems involved in the dynamics of gene expression, which are fundamental to cell determination and function without alteration in DNA sequences, are based on methylation of the N-terminal tails of lysine residues and DNA methylation. We demonstrate the vital importance for genetic transfer by different (hydrogen) networks, suggesting a complex interaction between the two epigenetic modifications. In other words, the methylation of local lysines can prescribe CPG methylation, which requires that methylation of histones and DNA are cooperative in carrying out an epigenetic instruction for integrating gene-silencing networks. To give a bio-organic description of the epigenetic coherence between histone and base methylation, we used the well-known B- into Z-DNA dynamics in combination with the unique properties of phosphatemethylated DNA on different levels of chemistry. [ABSTRACT FROM AUTHOR]

Abstract: We investigated the transcription levels, promoter methylation status and role as a tumor suppressor gene (TSG) of the cadherin CDH4 in nasopharyngeal carcinoma (NPC). The expression of CDH4 was decreased in NPC cell lines, xenografts and primary tumor biopsies. Promoter hypermethylation of CDH4 was detected in all five NPC cell lines, both NPC xenograft lines and 94.3% of primary tumors but not in any of the 12 normal epithelial samples. Loss of CDH4 expression could be restored by the methyltransferase inhibitor 5-aza-2′-deoxycytidine in NPC cell lines. Ectopic expression of CDH4 in the NPC cell lines inhibits cell proliferation, colony formation, migration and elicit cell communication. CDH4 may be a novel putative TSG that can be frequently and tumor-specifically inactivated by its promoter methylation in NPC. [Copyright &y& Elsevier]

Abstract: Gene silencing by epigenetic mechanisms is frequent in prostate cancer (PCA). The link between DNA hypermethylation and histone modifications is not completely understood. We chose the GSTP1 gene which is silenced by hypermethylation to analyze the effect of the histone deacetylase inhibitor depsipeptide on DNA methylation and histone modifications at the GSTP1 promoter site. Prostate cell lines (PC-3, LNCaP, and BPH-1) were treated with depsipeptide; apoptosis (FACS analysis), GSTP1 mRNA levels (quantitative real-time PCR), DNA hypermethylation (methylation-specific PCR), and histone modifications (chromatin immunoprecipitation) were studied. Depsipeptide induced apoptosis in PCA cells, but not a cell cycle arrest. Depispeptide reversed DNA hypermethylation and repressive histone modifications (reduction of H3K9me2/3 and H3K27me2/3; increase of H3K18Ac), thereby inducing GSTP1 mRNA re-expression. Successful therapy requires both, DNA demethylation and activating histone modifications, to induce complete gene expression of epigenetically silenced genes and depsipeptide fulfils both criteria. [Copyright &y& Elsevier]

Abstract: Epigenetic mechanisms maintain heritable changes in gene expression and chromatin organization over many cell generations. Importantly, deregulated epigenetic mechanisms play a key role in a wide range of human malignancies, including liver cancer. Hepatocellular carcinoma (HCC), which originates from the hepatocytes, is by far the most common liver cancer, with rates and aetiology that show considerable geographic variation. Various environmental agents and lifestyles known to be risk factors for HCC (such as infection by hepatitis B virus (HBV) and hepatitis C virus (HCV), chronic alcohol intake, and aflatoxins) are suspected to promote its development by eliciting epigenetic changes, however the precise gene targets and underlying mechanisms have not been elucidated. Many recent studies have exploited conceptual and technological advances in epigenetics and epigenomics to investigate the role of epigenetic events induced by environmental factors in HCC tumors and non-tumor precancerous (cirrhotic) lesions. These studies have identified a large number of genes and pathways that are targeted by epigenetic deregulation (changes in DNA methylation, histone modifications and RNA-mediated gene silencing) during the development and progression of HCC. Frequent identification of aberrant epigenetic changes in specific genes in cirrhotic tissue is consistent with the notion that epigenetic deregulation of selected genes in pre-malignant lesions precedes and promotes the development of HCC. In addition, several lines of evidence argue that some environmental factors (such as HBV virus) may abrogate cellular defense systems, induce silencing of host genes and promote HCC development via an “epigenetic strategy”. Finally, profiling studies reveal that HCC tumors and pre-cancerous lesions may exhibit epigenetic signatures associated with specific risk factors and tumor progression stage. Together, recent evidence underscores the importance of aberrant epigenetic events induced by environmental factors in liver cancer and highlights potential targets for biomarker discovery and future preventive and therapeutic strategies. [Copyright &y& Elsevier]

Abstract: DNA methylation is a gene-silencing and host defense system that can down-regulate viral gene expression in mammalian cells. An established targeted DNA methylation method was used to demonstrate that genome-integrated CMV and adenovirus type 5 E1A promoters were hypermethylated after MCF7 and HEK293 cells were transfected with in vitro methylated viral promoter fragments. In both cases, the targeted methylation-induced gene silencing could be reversed by addition of 5-aza-2′-deoxycytidine, confirming that the CMV and E1A promoters are regulated by DNA methylation. The kinetics of the targeted DNA methylation was determined using a reporter system in live cells. In conclusion, targeted DNA methylation is able to efficiently silence susceptible viral promoters and provides an alternative strategy to study the impact of loci-specific DNA methylation in viral gene expression. [Copyright &y& Elsevier]

One of the key questions in the study of mammalian gene regulation is how epigenetic methylation patterns on histones and DNA are initiated and established. These stable, heritable, covalent modifications are largely associated with the repression or silencing of gene transcription, and when deregulated can be involved in the development of human diseases such as cancer. This article reviews examples of viruses and bacteria known or thought to induce epigenetic changes in host cells, and how this might contribute to disease. The heritable nature of these processes in gene regulation suggests that they could play important roles in chronic diseases associated with microbial persistence; they might also explain so-called ‘hit-and-run’ phenomena in infectious disease pathogenesis. [ABSTRACT FROM AUTHOR]

The neuronal repellent SLIT2 is repressed in a number of cancer types primarily through promoter hypermethylation. SLIT2, however, has not been studied in prostate cancer. Through genome-wide location analysis we identified SLIT2 as a target of polycomb group (PcG) protein EZH2. The EZH2-containing polycomb repressive complexes bound to the SLIT2 promoter inhibiting its expression. SLIT2 was downregulated in a majority of metastatic prostate tumors, showing a negative correlation with EZH2. This repressed expression could be restored by methylation inhibitors or EZH2-suppressing compounds. In addition, a low level of SLIT2 expression was associated with aggressive prostate, breast and lung cancers. Functional assays showed that SLIT2 inhibited prostate cancer cell proliferation and invasion. Thus, this study showed for the first time the epigenetic silencing of SLIT2 in prostate tumors, and supported SLIT2 as a potential biomarker for aggressive solid tumors. Importantly, PcG-mediated repression may serve as a precursor for the silencing of SLIT2 by DNA methylation in cancer. [ABSTRACT FROM AUTHOR]

Methylation of promoter CpG islands has been associated with gene silencing and demonstrated to lead to chromosomal instability. Therefore, some postulate that aberrantly methylated CpG regions may be important biomarkers indicative of cancer development. In this study we used the Illumina GoldenGate Methylation BeadArray Cancer Panel I for simultaneously profiling methylation of 1,505 CpG sites in order to identify methylation differences in 76 liver tissues ranging from normal to pre-neoplastic and neoplastic states. CpG sites for ESR1, GSTM2, and MME were significantly differentially methylated when comparing the pre-neoplastic tissues from patients with concomitant hepatocellular carcinoma (HCC) to the pre-neoplastic tissues from patients without HCC. When comparing paired HCC tissues to their corresponding pre-neoplastic non-tumorous tissues, eight CpG sites, including one CpG site that was hypermethylated ( APC) and seven ( NOTCH4, EMR3, HDAC9, DCL1, HLA- DOA, HLA- DPA1, and ERN1) that were hypomethylated in HCC, were identified. Our study demonstrates that high-throughput methylation technologies may be used to identify differentially methylated CpG sites that may prove to be important molecular events involved in carcinogenesis. [ABSTRACT FROM AUTHOR]

Background: Silencing of transgenes in mice is a common phenomenon typically associated with short multi-copy transgenes. We have investigated the regulation of the highly inducible human granulocyte-macrophage colonystimulating-factor gene (Csf2) in transgenic mice. Results: In the absence of any previous history of transcriptional activation, this transgene was expressed in T lineage cells at the correct inducible level in all lines of mice tested. In contrast, the transgene was silenced in a specific subset of lines in T cells that had encountered a previous episode of activation. Transgene silencing appeared to be both transcription-dependent and mediated by epigenetic mechanisms. Silencing was accompanied by loss of DNase I hypersensitive sites and inability to recruit RNA polymerase II upon stimulation. This pattern of silencing was reflected by increased methylation and decreased acetylation of histone H3 K9 in the transgene. We found that silenced lines were specifically associated with a single pair of tail-to-tail inverted repeated copies of the transgene embedded within a multi-copy array. Conclusions: Our study suggests that epigenetic transgene silencing can result from convergent transcription of inverted repeats which can lead to silencing of an entire multi-copy transgene array. This mechanism may account for a significant proportion of the reported cases of transgene inactivation in mice. [ABSTRACT FROM AUTHOR]

Abstract: The Ras-Association Domain Family (RASSF) comprises ten members, termed RASSF1 to RASSF10. RASSF1 to RASSF6 harbor a C-terminal Ras-association (RA) domain and RASSF7 to RASSF10 contain an N-terminal RA domain. Interestingly, it was observed that in various tumor types distinct RASSFs transcripts (e.g. RASSF1A and RASSF2A) are missing due to hypermethylation of their CpG island promoter. Since methylation of the RASSF1A promoter is described as an early and frequent event in tumorigenesis, RASSF1A could serve as a useful diagnostic marker in cancer screens. RASSFs are implicated in various cellular mechanisms including apoptosis, cell cycle control and microtubule stabilization, though little is known about the underlying mechanisms. Tumor suppressing functions were reported for several members. Here we review the current literature on RASSF members focusing on structural, functional and epigenetic aspects. Characterizing the cellular mechanisms that regulate the signaling pathways RASSFs are involved in, could lead to a deeper understanding of tumor development and, furthermore, to new strategies in cancer treatment. [Copyright &y& Elsevier]

X inactivation is the process of a chromosome-wide silencing of the majority of genes on the X chromosome during early mammalian development. This process may be aberrant in cloned animals. Here we show that repressive modifications, such as methylation of DNA, and the presence of methylated histones, H3K9me2 and H3K27me3, exhibit distinct aberrance on the inactive X chromosome in live clones. In contrast, H3K4me3, an active gene marker, is obviously missing from the inactive X chromosome in all cattle studied. This suggests that the disappearance of active histone modifications (H3K4me3) seems to be more important for X inactivation than deposition of marks associated with heterochromatin (DNA methylation, H3K27me3 and H3K9me2). It also implies that even apparently normal clones may have subtle abnormalities in repressive, but not activating epigenetic modifications on the inactive X when they survive to term. We also found that the histone H3 methylations were enriched and co-localized at q21-31 of the active X chromosome, which may be associated with an abundance of LINE1 repeat elements. [ABSTRACT FROM AUTHOR]

Abstract: Epigenetic silencing of sFRP genes has been shown to lead to constitutive activation of the canonical Wnt-signaling pathway. The first description of deregulated Wnt-signaling activation in a hematological malignancy was reported in chronic myeloid leukemia (CML). To investigate whether epigenetic silencing of sFRP is responsible for the observed Wnt activation in CML, we studied the methylation and mutational status of the sFRP1 promoter in 48 chronic phase CML patients. Of the 48 CML patients 41 were shown to be unmethylated, 6 patients hemi-methylated and 1 patient fully methylated at the sFRP1 promoter. Albeit observed infrequently in chronic phase CML, sFRP1 promoter methylation correlated with primary cytogenetic resistance to imatinib mesylate. sFRP1 promoter methylation may indicate a genetically more unstable form of disease resistant to therapy and provide a key biological difference in therapy resistant patients, in addition to a possible mechanism for the observed activation of canonical Wnt signaling in CML. [Copyright &y& Elsevier]

Abstract: Hexavalent chromium [Cr(VI)] is a mutagen and carcinogen, and occupational exposure can lead to lung cancers and other adverse health effects. Genetic changes resulting from DNA damage have been proposed as an important mechanism that mediates chromate''s carcinogenicity. Here we show that chromate exposure of human lung A549 cells increased global levels of di- and tri-methylated histone H3 lysine 9 (H3K9) and lysine 4 (H3K4) but decreased the levels of tri-methylated histone H3 lysine 27 (H3K27) and di-methylated histone H3 arginine 2 (H3R2). Most interestingly, H3K9 dimethylation was enriched in the human MLH1 gene promoter following chromate exposure and this was correlated with decreased MLH1 mRNA expression. Chromate exposure increased the protein as well as mRNA levels of G9a a histone methyltransferase that specifically methylates H3K9. This Cr(VI)-induced increase in G9a may account for the global elevation of H3K9 dimethylation. Furthermore, supplementation with ascorbate, the primary reductant of Cr(VI) and also an essential cofactor for the histone demethylase activity, partially reversed the H3K9 dimethylation induced by chromate. Thus our studies suggest that Cr(VI) may target histone methyltransferases and demethylases, which in turn affect both global and gene promoter specific histone methylation, leading to the silencing of specific tumor suppressor genes such as MLH1. [Copyright &y& Elsevier]

Abstract: To identify novel methylation-silenced genes in gastric cancer, we carried out a genome-wide search for genes that are up-regulated after treatment with the demethylating agent, 5-aza-2′-deoxycytidine (5Aza-dC). When three gastric cancer cell lines (SNU-1,-601, and -719) were treated with 5Aza-dC, 143 genes were found to be upregulated by twofold or more using oligonucleotide microarrays. Six of these genes, i.e. TFPI2, GPX3, GPX1, IGFBP6, IRF7 and DMRT1, showed promoter hypermethylation in one or more gastric cancer cell lines, but were unmethylated in normal gastric mucosa by bisulphite sequencing and methylation-specific PCR analysis. The following percentages of these genes were found to be aberrantly methylated in gastric cancer samples; TFPI2 (80.9%), GPX3 (30.1%), DMRT1 (46.9%), GPX1 (16.7%), IGFBP6 (22.6%) and IRF7 (32.1%). Interestingly, the survival of patients possessing methylated alleles of TFPI2 (123/152, 80.9%) was poorer than that of patients with unmethylated alleles (p =0.023). Multivariate analysis confirmed that TFPI2 methylation is a significant and independent prognostic factor in gastric carcinoma. Furthermore, altered TFPI2 expression, as demonstrated by immunohistochemistry in 566 consecutive gastric cancer tissues, was found to be significantly associated with sex (p =0.003), WHO classification (p <0.001), and a mixed subtype by Lauren’s classification (p <0.001). Thus, the present study identified several novel genes, which were methylated in gastric cancer and among them, methylation of TFPI2 was an unfavourable prognostic marker. [Copyright &y& Elsevier]

Abstract: Tissue specific differentially methylated regions (TDMRs) were identified and localized in the mouse genome using second generation virtual RLGS (vRLGS). Sequenom MassARRAY quantitative methylation analysis was used to confirm and determine the fine structure of tissue specific differences in DNA methylation. TDMRs have a broad distribution of locations to intragenic and intergenic regions including both CpG islands, and non-CpG islands regions. Somewhat surprising, there is a strong bias for TDMR location in non-promoter intragenic regions. Although some TDMRs are within or close to repeat sequences, overall they are less frequently associated with repetitive elements than expected from a random distribution. Many TDMRs are methylated at early developmental stages, but unmethylated later, suggesting active or passive demethylation, or expansions of populations of cells with unmethylated TDMRs. This is notable during postnatal testis differentiation where many testis specific TDMRs become progressively “demethylated”. These results suggest that methylation changes during development are dynamic, involve demethylation and methylation, and may occur at late stages of embryonic development or even postnatally. [Copyright &y& Elsevier]

Abstract: HIC1 (Hypermethylated in Cancer 1), as it name implied, was originally isolated as a new candidate tumor suppressor gene located at 17p13.3 because it resides in a CpG island that is hypermethylated in many types of human cancers. HIC1 encodes a transcription factor associating an N-terminal BTB/POZ domain to five C-terminal Krüppel-like C2H2 zinc finger motifs. In this review, we will begin by providing an overview of the current knowledge on HIC1 function, mainly gained from in vitro studies, as a sequence-specific transcriptional repressor interacting with a still growing range of HDAC-dependent and HDAC-independent corepressor complexes. We will then summarize the studies that have demonstrated frequent hypermethylation changes or losses of heterozygosity of the HIC1 locus in human cancers. Next, we will review animal models which have firmly established HIC1 as a bona fide tumor suppressor gene epigenetically silenced and functionally cooperating notably with p53 within a complex HIC1-p53-SIRT1 regulatory loop. Finally, we will discuss how this epigenetic inactivation of HIC1 might “addict” cancer cells to altered survival and signaling pathways or to lineage-specific transcription factors during the early stages of tumorigenesis. [Copyright &y& Elsevier]

Objective: Multiple myeloma (MM) is presently an incurable malignant plasma cell tumor. The objective of this study was to investigate expression of the interferon regulatory factor family (IRF1-9) and the potential role of DNA methylation in silencing IRF genes in MM cell lines and purified MM cells from patients. Materials and Methods: Using a panel of 13 human MM cell lines and purified CD138+ cells from nine MM patients, expression of IRF genes was investigated by quantitative reverse transcriptase polymerase chain reaction and Western blot. DNA methylation of the interferon consensus sequence–binding protein (ICSBP/IRF8) gene was measured using pyrosequencing, and the effect of promoter methylation on expression was analyzed by in vitro methylation of a cloned ICSBP/IRF8 promoter, and treatment of MM cells with 5-aza-2′-deoxycytidine (DAC). Results: Eight of thirteen of the MM cell lines were found to lack ICSBP/IRF8 expression, associated with hypermethylation of the CpG island in the ICSBP/IRF8 promoter. We also found that ICSBP/IRF8 was significantly underexpressed in primary MM cells, whereas the ICSBP/IRF8 promoter was methylated in only one of nine of primary purified CD138+ MM samples. DAC-mediated demethylation restored endogenous ICSBP/IRF8 expression, whereas in vitro methylation silenced the promoter. Conclusion: Expression of the ICSBP/IRF8 gene is silenced in a majority of MM cell lines and primary CD138+ MM cells. DNA methylation of the ICSBP/IRF8 gene is a frequent event in MM cell lines, but silencing is also observed in the absence of methylation. These results suggest that silencing of ICSBP/IRF8 expression, by DNA methylation or other epigenetic mechanisms, may be associated with the malignant phenotype of MM. [Copyright &y& Elsevier]

Advances in molecular biology have provided tools for studying the epigenetic factors that modulate gene expression. DNA methylation is an epigenetic modification that can have sustained effects on transcription and is associated with long-term gene silencing. In this review, we focus on the regulation of estrogen receptor alpha (ERα) expression by hormonal and environmental cues, the consequences of these cues for female maternal and sexual behavior, and recent studies that explore the role of DNA methylation in mediating these developmental effects, with particular focus on the mediating role of maternal care. The methylation status of ERα has implications for reproductive behavior, cancer susceptibility, and recovery from ischemic injury, suggesting an epigenetic basis for risk and resilience across the life span. [Copyright &y& Elsevier]

Abstract: Gene inactivation by promoter hypermethylation has been demonstrated in the colonic mucosa of colorectal cancer (CRC) patients. However, current data do not prove direct involvement of this epigenetic modification in the early stages of CRC. Promoter methylation profiles of E-cadherin, hMLH1, MGMT, p16INK4a, p15INK4b and p14ARF ; mutations of K-ras, B-raf and TP53 and microsatellite instability (MSI) were examined in normal and cancerous colonic mucosal tissue in 82 CRC patients using methylation-specific PCR assays. Methylation of hMLH1 and MGMT in normal mucosa correlated significantly with MSI and K-ras activation in neighbouring cancerous mucosal tissues. Similarly, poorly differentiated tumours were associated with methylated p16INK4a and E-cadherin in neighbouring normal colonic tissues (NCTs). Our results indicate that epigenetic changes in mucosa surrounding colorectal neoplastic lesions may describe a ‘field cancerisation’ phenomenon that may occur previous to genetic alterations in early stages of carcinogenesis. [Copyright &y& Elsevier]

Abstract: Knowledge on heritable changes in gene expression that result from epigenetic events is of increasing relevance in the development of strategies for prevention, early diagnosis and treatment of cancer. Histone acetylation and DNA methylation are epigenetic modifications whose patterns can be regarded as heritable marks that ensure accurate transmission of the chromatin states and gene expression profiles over many cell generations. Importantly, patterns and levels of DNA methylation and histone acetylation are profoundly altered in human cancers. Accumulating evidence suggests that an epigenetic cross-talk, i.e. interplay between DNA methylation and histone acetylation, may be involved in the process of gene transcription and aberrant gene silencing in tumours. Although the molecular mechanism of gene activation is relatively well understood, the hierarchical order of events and dependencies leading to gene silencing in the course of cancer development remain largely unknown. While some studies suggest that DNA methylation patterns guide histone modifications (including histone acetylation and methylation) during gene silencing, other studies argue that DNA methylation takes its cues primarily from histone modification states. In this review, we summarize current knowledge on the interplay between DNA methylation and histone modifications during gene silencing and its importance in the integration of environmental and intrinsic stimuli in the control of gene expression. We also discuss the importance of an epigenetic cross-talk in the protection against genetic changes in response to environmental genotoxins as well as the implication for cancer therapy and prevention. [Copyright &y& Elsevier]

A central question in genomic imprinting is how a specific sequence is recognized as the target for epigenetic marking. In both mammals and plants, imprinted genes are often associated with tandem repeats and transposon-related sequences, but the role of these elements in epigenetic gene silencing remains elusive. FWA is an imprinted gene in Arabidopsis thaliana expressed specifically in the female gametophyte and endosperm. Tissue-specific and imprinted expression of FWA depends on DNA methylation in the FWA promoter, which is comprised of two direct repeats containing a sequence related to a SINE retroelement. Methylation of this element causes epigenetic silencing, but it is not known whether the methylation is targeted to the SINE-related sequence itself or the direct repeat structure is also necessary. Here we show that the repeat structure in the FWA promoter is highly diverse in species within the genus Arabidopsis. Four independent tandem repeat formation events were found in three closely related species. Another related species, A. halleri, did not have a tandem repeat in the FWA promoter. Unexpectedly, even in this species, FWA expression was imprinted and the FWA promoter was methylated. In addition, our expression analysis of FWA gene in vegetative tissues revealed high frequency of intra-specific variation in the expression level. In conclusion, we show that the tandem repeat structure is dispensable for the epigenetic silencing of the FWA gene. Rather, SINE-related sequence is sufficient for imprinting, vegetative silencing, and targeting of DNA methylation. Frequent independent tandem repeat formation events in the FWA promoter led us to propose that they may be a consequence, rather than cause, of the epigenetic control. The possible significance of epigenetic variation in reproductive strategies during evolution is also discussed. [ABSTRACT FROM AUTHOR]

Transposable elements (TEs) are the single largest component of most eukaryote genomes. Rice and wheat offer ideal systems to study epigenetic regulation of TEs and to investigate the evolution of TEs following allopolyploidization. Almost 40% of the rice genome and ~90% of the wheat genome are derived from TEs. Most elements contain inactivating mutations, but others are reversibly silenced by epigenetic mechanisms. The presence of TE sequences in EST and cDNA databases indicates that those elements escaped silencing and expressed. The transcriptional activation of TEs might impact the expression of adjacent host genes. Allopolyploidization in wheat activates long terminal repeat (LTR) retrotransposon promoters that initiate readout transcription into adjacent genes. Moreover, LTR methylation and readout transcription initiated from promoters in LTRs change during plant development. The power of whole genome analysis of TE methylation and transcription coupled with the availability of virtually a complete genome sequence for rice resulted in much higher resolution than in previously reported studies. The underlying mechanism(s) whereby TEs alter expression of adjacent genes through readout transcription and the potential significant biological role of transcriptional interference between retroelements and cellular genes are discussed. [ABSTRACT FROM AUTHOR]

Hypermethylation of CpG islands is a common epigenetic alteration associated with cancer. Global patterns of hypermethylation are tumor-type specific and nonrandom. The biological significance and the underlying mechanisms of tumor-specific aberrant promoter methylation remain unclear, but some evidence suggests that this specificity involves differential sequence susceptibilities, the targeting of DNA methylation activity to specific promoter sequences, or the selection of rare DNA methylation events during disease progression. Using restriction landmark genomic scanning on samples derived from tissue culture and in vivo models of T cell lymphomas, we found that MYC overexpression gave rise to a specific signature of CpG island hypermethylation. This signature reflected gene transcription profiles and was detected only in advanced stages of disease. The further inactivation of the Pten, p53, and E2f2 tumor suppressors in MYC-induced lymphomas resulted in distinct and diagnostic CpG island methylation signatures. Our data suggest that tumor-specific DNA methylation in lymphomas arises as a result of the selection of rare DNA methylation events during the course of tumor development. This selection appears to be driven by the genetic configuration of tumor cells, providing experimental evidence for a causal role of DNA hypermethylation in tumor progression and an explanation for the tremendous epigenetic heterogeneity observed in the evolution of human cancers. The ability to predict genome-wide epigenetic silencing based on relatively few genetic alterations will allow for a more complete classification of tumors and understanding of tumor cell biology. [ABSTRACT FROM AUTHOR]

Hypermethylation of tumor suppressor genes is central to the pathogenesis of human malignancy, yet this alteration's etiology remains unclear, and its clinical impact has not yet been studied using an approach that can yield directly generalizable results. Therefore, we sought to examine both of these issues in bladder cancer, seeking to understand the characteristics of epidemiologically well-defined patient tumors with differing levels of methylation silencing. We analyzed epigenetic silencing of 16 genes in a population-based incident case series of 331 bladder transitional cell carcinomas. We utilized a novel item response theory (IRT) model, to examine, in an unbiased fashion, the relationship of patient characteristics, carcinogen exposure history and tumor characteristics with the underlying propensity for gene hypermethylation. Age, male gender and current cigarette smoking were significantly positively associated with the methylation latent trait. Promoter hypermethylation as a latent trait significantly predicted both non-invasive/high-grade and invasive stage disease and was also significantly associated with survival, with each unit increase in the latent trait resulting in a 30% increase in the risk of death. This work, studying all stages and grades of incident bladder cancer, provides definitive evidence that carcinogen exposures play a critical role in selecting these alterations in tumorous clones and that epigenetic silencing is a strong and significant predictor of tumor stage and overall patient survival. Finally, our novel approach provides insight into the etiology of promoter hypermethylation, suggesting that selected, carcinogen-exposed individuals have a greater propensity for hypermethylation that is associated with more aggressive, fatal disease. [ABSTRACT FROM PUBLISHER]

14-3-3σ proteins regulate numerous cellular processes that are important to cancer development. One of its biological roles involves G2 cell-cycle arrest following DNA damage. It has also been reported that the loss of 14-3-3σ expression via CpG methylation may contribute to malignant transformation by impairing the G2 cell-cycle checkpoint function, thereby allowing an accumulation of genetic defects. However, how the CpG methylation-dependent silencing mechanism works in relation to promoter methylation associated with methyl-CpG-binding proteins (MeCPs) is still unclear. To better understand the mechanism, we first examined the methylation status of the 14-3-3σ promoter-associated CpG islands and 14-3-3σ gene expression in a subset of prostate cancer cell lines using methylation-specific PCR (MSP), an HhaI-based DNA methylation assay, and reverse transcription–PCR (RT–PCR). We found that the 14-3-3σ expression is lost in LNCaP and Tramp-C1 prostate cancer cell lines and that this expression is restored after treatment with epigenetic silencing modifiers 5-aza-2′-deoxycytidine (5-aza) and trichostatin A (TSA). These results imply transcriptional silencing via promoter-associated CpG methylation. Chromatin immunoprecipitation analysis revealed that methyl-CpG-binding protein 2 (MBD2) is associated preferentially to the methylated CpG island in the 14-3-3σ promoter in LNCaP and Tramp-C1 cells but not in 14-3-3σ-expressing PC3 and DU145 cells, which contain an unmethylated CpG island in the 14-3-3σ promoter region. The 14-3-3σ gene silencing because of CpG methylation correlates with binding of MBD2. In addition, the activation of 14-3-3σ gene expression by a combination of 5-aza and TSA also involves the release of the MBD2 from the 14-3-3σ promoter-methylated CpG island in LNCaP and Tramp-C1 cells. Furthermore, MBD2 knockdown by siRNA stimulated 14-3-3σ expression in LNCaP cells. We also investigated whether the loss of 14-3-3σ expression in LNCaP and Tramp-C1 cells affects cell proliferation by MTT assays. Interestingly, we observed that 14-3-3σ-inactivated LNCaP and Tramp-C1 cells had markedly decreased cell proliferation and protein expression of proliferation cell nuclear antigen (PCNA) after restoration of 14-3-3σ expression with 5-aza and TSA treatment. On the other hand, the same treatment did not significantly affect 14-3-3σ-active PC3 and DU145 cells, which normally express 14-3-3σ. Finally, 14-3-3σ knockdown by siRNA resulted in increased proliferation in PC3 and DU145 cells. These findings suggest that the transcriptional silencing of the 14-3-3σ gene is caused by promoter CpG island methylation associated with MBD2, and that this may play an important role in prostate cancer progression during the invasive and metastatic stages of the disease.Oncogene (2006) 25, 4559–4572. doi:10.1038/sj.onc.1209462; published online 19 June 2006 [ABSTRACT FROM AUTHOR]

Epigenetic alterations like DNA methylation and the resulting inactivation of cancer-related genes often contribute to the development of various cancers. To identify the genes that are silenced by aberrant methylation in renal cell carcinoma (RCC), we subjected two RCC lines to methylated CpG island amplification/representational difference analysis. This identified 27 CpG islands. Combined bisulfite restriction analysis of these CpG islands in primary RCC cases revealed that four were methylated in a tumor-specific manner. One of these was identified as the human homeo-box gene B13 (HOXB13) gene, but the remaining three CpG islands were not associated with known genes. The methylation frequencies of HOXB13 in primary RCC samples and lines were 30 and 73%, respectively. The methylation status of HOXB13 correlated with the loss of its expression both in RCC lines and primary tumors, and methyltransferase inhibitor treatment induced the recovery of its expression. Exogenous expression of HOXB13 in RCC cells that lacked endogenous HOXB13 expression suppressed colony formation and induced apoptotic features. Furthermore, HOXB13 methylation correlated positively with tumor grade and microvessel invasion. These results suggest that HOXB13 is a novel candidate tumor suppressor gene in RCC and that its inactivation may play an important role in both RCC tumorigenesis and progression.Oncogene (2006) 25, 1733–1742. doi:10.1038/sj.onc.1209200; published online 7 November 2005 [ABSTRACT FROM AUTHOR]

DNA methylation, histone deacetylation, and methylation of histone H3 at lysine 9 are the three best-characterized covalent modifications associated with a repressed chromatin state. Recent advances highlight an essential, intricate web of interactions among these processes, generating a self-reinforcing, self-perpetuating cycle of epigenetic events that lead to long-term transcriptional repression. Histone deacetylation and methylation at lysine 9 of H3 might also contribute to the establishment of DNA methylation patterns, a long-standing mystery in epigenetics. What''s more, recent clues suggest a potential link between CpG methylation and other histone modifications. A complex picture is emerging in which DNA methylation and histone modifications work hand-in-hand as parts of an epigenetic program that integrates gene-silencing networks within the cell. [Copyright &y& Elsevier]

Epigenetic modification of plant gene and transposon activity, which correlates with their methylation, is often heritable over many generations. Such heritable properties allow conventional genetic linkage analysis to identify the sequences affected in epigenetic variants. Machinery controlling the establishment of the epigenetic state and role of the epigenetic controls in plant development are also discussed. [ABSTRACT FROM PUBLISHER]

Despite prolonged antiretroviral therapy, HIV-1 persists as transcriptionally inactive proviruses. The HIV-1 latency remains a principal obstacle in curing AIDS. It is important to understand mechanisms by which HIV-1 latency is established to make the latent reservoir smaller. We present a molecular characterization of distinct populations at an early phase of infection. We developed an original dual-color reporter virus to monitor LTR kinetics from establishment to maintenance stage. We found that there are two ways of latency establishment i.e., by immediate silencing and slow inactivation from active infection. Histone covalent modifications, particularly polycomb repressive complex 2 (PRC2)-mediated H3K27 trimethylation, appeared to dominate viral transcription at the early phase. PRC2 also contributes to time-dependent LTR dormancy in the chronic phase of the infection. Significant differences in sensitivity against several stimuli were observed between these two distinct populations. These results will expand our understanding of heterogeneous establishment of HIV-1 latency populations. [ABSTRACT FROM AUTHOR]

The article discusses inbreeding effects in epigenesis. It mentions about the omic technology which enhances the knowledge of inbreeding effects. It discovers epigenetics to the differential regulation of gene expression, imprinting and gene silencing. Epigenetic differences among natural population is being linked with the variations of DNA methylation.

Abstract: The implication of DNA hypermethylation in the pathogenesis of myelodysplastic syndromes (MDS) provides a rationale for using hypomethylating agents such as azacitidine. Growing evidence suggests that azacitidine may reverse epigenetic gene silencing at specific genomic targets. AZA-001 established azacitidine as the first agent to provide a significant overall survival benefit in MDS patients. These data confirmed that azacitidine has a progressively cumulative effect on the MDS clone and support the value of maintenance therapy. Prolonged survival was independent of achieving complete response. Azacitidine in combination with histone deacetylase inhibitors might offer better efficacy by modulating the methylation and acetylation states of silenced genes. [Copyright &y& Elsevier]

Chromosomal loss of 18q21 is a frequent event in colorectal cancer (CRC) development, suggesting that this region harbors tumor suppressor genes (TSGs). Several candidate TSGs, among which methyl-CpG-binding domain protein 1 (MBD1), CpG-binding protein CXXC1, Sma- and Mad-related protein 4 (SMAD4), deleted in colon cancer (DCC) and methyl-CpG-binding domain protein 2 (MBD2) are closely linked on a 4-Mb DNA region on chromosome18q21. As TSGs can be epigenetically silenced, this study investigates whether MBD1, CXXC1, SMAD4, DCC and MBD2 are subject to epigenetic silencing in CRC. Methylation-specific polymerase chain reaction and sodium bisulfite sequencing of these genes show that DCC, but not MBD1, CXXC1, SMAD4 and MBD2, has promoter CpG island methylation in CRC cell lines and tissues {normal mucosa [29.5% (18/61)], adenomas [81.0% (47/58)] and carcinomas [82.7% (62/75)] (P = 8.6 × 10−9)} that is associated with reduced DCC expression, independent of 18q21 loss analyzed by multiplex ligation-dependent probe amplification. Reduced gene expression of CXXC1, SMAD4 and MBD2 correlates with 18q21 loss in CRC cell lines (P = 0.04, 0.02 and 0.02, respectively). Treatment with the demethylating agent 5-aza-2′-deoxycytidine, but not with the histone deacetylase inhibitor trichostatin A exclusively restored DCC expression in CRC cell lines. Chromatin immunoprecipitation studies reveal that the DCC promoter is marked with repressive histone-tail marks H3K9me3 and H3K27me3, whereas activity related H3K4me3 was absent. Only active epigenetic marks were detected for MBD1, CXXC1, SMAD4 and MBD2. This study demonstrates specific epigenetic silencing of DCC in CRC as a focal process not affecting neighboring genes on chromosomal region 18q21. [ABSTRACT FROM AUTHOR]

Within the human genome there are hundreds of copies of the rRNA gene, but only a fraction of these genes are active. Silencing through epigenetics has been extensively studied; however, it is essential to understand how active rRNA genes are maintained. Here, we propose a role for the methyl-CpG binding domain protein MBD3 in epigenetically maintaining active rRNA promoters. We show that MBD3 is localized to the nucleolus, colocalizes with upstream binding factor, and binds to unmethylated rRNA promoters. Knockdown of MBD3 by small interfering RNA results in increased methylation of the rRNA promoter coupled with a decrease in RNA polymerase I binding and pre-rRNA transcription. Conversely, overexpression of MBD3 results in decreased methylation of the rRNA promoter. Additionally, overexpression of MBD3 induces demethylation of nonreplicating plasmids containing the rRNA promoter. We demonstrate that this demethylation occurs following the overexpression of MBD3 and its increased interaction with the methylated rRNA promoter. This is the first demonstration that MBD3 is involved in inducing and maintaining the demethylated state of a specific promoter. [ABSTRACT FROM AUTHOR]

The pharmacological effect of morphine as a painkiller is mediated mainly via the mu opioid receptor (MOR) and is dependent on the number of MORs in the cell surface membrane. While several studies have reported that the MOR gene is regulated by various cis- and trans-acting factors, many questions remain unanswered regarding in vivo regulation. The present study shows that epigenetic silencing and activation of the MOR gene are achieved through coordinated regulation at both the histone and DNA levels. In P19 mouse embryonal carcinoma cells, expression of the MOR was greatly increased after neuronal differentiation. MOR expression could also be induced by a demethylating agent (5′-aza-2′-deoxycytidine) or histone deacetylase inhibitors in the P19 cells, suggesting involvement of DNA methylation and histone deacetylation for MOR gene silencing. Analysis of CpG DNA methylation revealed that the proximal promoter region was unmethylated in differentiated cells compared to its hypermethylation in undifferentiated cells. In contrast, the methylation of other regions was not changed in either cell type. Similar methylation patterns were observed in the mouse brain. In vitro methylation of the MOR promoters suppressed promoter activity in the reporter assay. Upon differentiation, the in vivo interaction of MeCP2 was reduced in the MOR promoter region, coincident with histone modifications that are relevant to active transcription. When MeCP2 was disrupted using MeCP2 small interfering RNA, the endogenous MOR gene was increased. These data suggest that DNA methylation is closely linked to the MeCP2-mediated chromatin structure of the MOR gene. Here, we propose that an epigenetic mechanism consisting of DNA methylation and chromatin modification underlies the cell stage-specific mechanism of MOR gene expression. [ABSTRACT FROM AUTHOR]