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Results for “DNA methylation profiling”

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At least 667 records · Page 37Linked to original sources
PubMed2003

Upregulation of the truncated basic hair keratin 1(hHb1-DeltaN) in carcinoma cells by Epstein-Barr virus (EBV).

To investigate the role of Epstein-Barr virus (EBV) in epithelial tumors, we compared the expression pattern of cellular genes in the EBV-infected gastric carcinoma cell line, NU-GC-3, and its uninfected control. Subtractive suppression hybridization (SSH) was combined with high-density DNA array screening to identify differentially expressed genes. We have discovered that EBV infection upregulated a truncated variant of human basic hair keratin 1 (hHb1-DeltaN), a gene that had previously been identified in metastatic breast carcinoma. We verified the differential expression of hHb1-DeltaN in 3 independent EBV-positive and -negative NU-GC-3 clones by Northern blotting. We further verified the EBV-dependent upregulation of hHb1-DeltaN in 3 other carcinoma cell lines (AGS, TWO3 and DLD1) by RT-PCR. Inhibition of CpG methylation by 5-Aza-CdR induced hHb1-DeltaN mRNA expression in the EBV-negative clones but did not alter the expression in the EBV-positive clones. The expression of hHb1-DeltaN protein was detectable by immunofluorescence and Western blotting in EBV-positive but not in EBV-negative NU-GC-3 clones after proteasome inhibitor (MG132) treatment. hHb1-DeltaN protein formed fibrous structures in the cytoplasm and accumulated in distinct nuclear bodies in the euchromatic areas of the cell nucleus. We suggest that the unstable hHb1-DeltaN protein may inhibit some of the functions of the keratin cytoskeleton and/or interfere with transcription regulation. It also may establish a link between EBV and the low differentiated or anaplastic status of the carcinomas that carry the virus.

Azacitidine↗
PubMed2026

Maternal high-fat diet modulates lupus nephritis through fetal Wnt-steroid hormone and epigenetic reprogramming in MRL/lpr mouse offspring.

We previously investigated whether maternal high-fat diet (HFD) exposure alters lupus nephritis (LN) progression in MRL/lpr offspring. Contrary to expectation, maternally HFD-exposed offspring showed delayed and attenuated nephritic progression compared with control diet offspring. The maternal HFD developmental impact on LN remains unclear. Here, integrated amniotic fluid metabolomics and fetal liver transcriptomics revealed that maternal HFD reshaped the intrauterine molecular environment, particularly involving steroid hormone biosynthesis and Wnt/β-catenin-associated regulatory networks. Methylome profiling further demonstrated broad CpG hypomethylation, immune-related differentially methylated region enrichment, and an inverse association between global CpG methylation and oxidative genomic DNA damage. Among candidate regulatory nodes, Axin2, a canonical Wnt/β-catenin target and feedback regulator, emerged as a potential link between fetal nutritional exposure, epigenetic remodeling, and persistent pathway modulation. Although whole-locus and gene body methylation of Axin2 were not markedly altered, promoter-region methylation showed an increasing tendency under maternal HFD exposure. In adult offspring, maternal HFD was associated with reduced Axin2 protein expression, decreased Wnt-responsive transcripts, increased peripheral corticosterone levels, and attenuation of LN progression. The inverse association between Axin2 expression and corticosterone further suggested coupling between suppressed Wnt pathway output and steroid hormone remodeling. Together, these findings support a developmental model in which maternal HFD reshapes the fetal intrauterine environment and establishes a persistent Wnt-steroid hormone-epigenetic regulatory axis that unexpectedly attenuates LN progression in genetically susceptible offspring.

Axin2↗
PubMed2005

Distinct expression profiles of Mecp2 transcripts with different lengths of 3'UTR in the brain and visceral organs during mouse development.

Four different transcripts of the Mecp2 gene can be distinguished by the length of the 3' untranslated region generated by usage of alternative polyadenylation sites. In situ hybridization analyses encompassing embryonic to 20-week postnatal age showed that transcripts are expressed in the central nervous system, with a progressive restriction during development culminating in localized strong expression in the cerebral cortex, olfactory bulb, hippocampal formation, and internal granule and Purkinje layer of the cerebellum. Real-time RT-PCR measurements of Mecp2 transcript levels showed variations with mouse age in two distinctive patterns that are unique to the central nervous system and the visceral organs, respectively. The 10-kb mRNA is the predominant form expressed in the brain in contrast to the shorter species expressed in the lung and liver. The developmental profile of Mecp2 mRNA highlights a potential tissue-specific function of the 3'UTR in the regulation of MeCP2 protein synthesis in response to the age-specific requirement of MeCP2 function during the life of the mouse.

3' Untranslated Regions↗
PubMed2004

Nocardioides aquiterrae sp. nov., isolated from groundwater in Korea.

A bacterial strain, GW-9T, which was isolated from groundwater in Korea, was subjected to a polyphasic taxonomic study using phenotypic characterization and phylogenetic and genetic methods. Phylogenetic analysis based on 16S rDNA sequences showed that strain GW-9T forms an evolutionary lineage within the radiation enclosing Nocardioides species and, in particular, a coherent cluster with Nocardioides pyridinolyticus. The cell-wall peptidoglycan type of strain GW-9T was based on LL-diaminopimelic acid as the diamino acid, indicating wall chemotype I. The predominant menaquinone was MK-8(H4). Strain GW-9T had a cellular fatty acid profile containing straight-chain, branched, unsaturated and 10-methyl fatty acids. The major fatty acid was iso-C(16:0). The DNA G+C content of strain GW-9T was 73 mol%. The 16S rDNA of strain GW-9T was 99.2% similar to that of the type strain of Nocardioides pyridinolyticus and 94.9-96.0% similar to sequences of the type strains of other Nocardioides species. Differences in phenotypic characteristics and genetic distinctiveness indicate that strain GW-9T is separate from previously described Nocardioides species. Therefore, on the basis of the data presented, a novel species of the genus Nocardioides, Nocardioides aquiterrae sp. nov., is proposed. The type strain is strain GW-9T (=KCCM 41647T=JCM 11813T).

DNA, Ribosomal↗
PubMed2026

Genome-wide chromatin recording resolves dynamic cell state changes.

Understanding how the chromatin state of a cell influences its future behavior is a major challenge throughout biology. However, most chromatin profiling methods are limited to endpoint assays. Here, we present LagTag, a method for recovery of earlier and endpoint chromatin states in the same mammalian cells. In this approach, transient expression of bacterial adenine methyltransferase fusions records the DNA binding profiles of chromatin-associated proteins of interest at earlier timepoints. Subsequent tagmentation and sequencing recovers the earlier chromatin profile from adenine methylation profiles, alongside endpoint profiles of endogenous chromatin-associated proteins. We verified that LagTag profiles aligned with those from established methods in mouse and human cells. We then applied LagTag to record and recover dynamic chromatin state transitions during mouse embryonic stem cell differentiation, capturing transcriptional signatures from pre- and post-differentiation timepoints within the same cell population. LagTag thus provides a foundation for temporally resolved chromatin profiling.

Journal Article↗
PubMed2002

Applications of CpG island microarrays for high-throughput analysis of DNA methylation.

Differential methylation hybridization (DMH) is a high-throughput microarray technique designed to identify changes in DNA methylation patterns commonly observed in cancer and other disease states. The DMH methodology comprises three fundamental components: the arraying of CpG island clones on glass slides, the preparation of the sample amplicons under investigation, and the hybridization of amplicon targets onto the CpG island microarray. Herein, we outline the DMH protocol and illustrate its utility and the validation approaches used in analyzing the hypermethylation profile of breast tumor and normal samples.

CpG Islands↗
PubMed1999

Gene silencing results in instability of antibody production in transgenic plants.

The stability of antibody and Fab expression was assessed in five different homozygous transgenic Arabidopsis lines. Each of these lines showed silencing of the transgenes that encode the antibody polypeptides, leading to instability of antibody production. However, each line had a different and specific instability profile. The characteristic variation in the level of antibody accumulation in each line as a function of developmental stage indicated that the T-DNA integration pattern played a role in triggering silencing, and also that the history and the integration position of simple transgene loci can influence the susceptibility to epigenetic silencing. In different lines with low antibody accumulation levels, methylation was found either in the promoter alone, in both the promoter and the transcribed region, in the transcribed region only, or in the transcribed region and downstream sequences. In conclusion, our data suggest that epigenetic effects result in different transgene expression profiles in each of the five Arabidopsis lines analyzed.

Antibody Formation↗
PubMed2004

Hairy transcriptional repression targets and cofactor recruitment in Drosophila.

Members of the widely conserved Hairy/Enhancer of split family of basic Helix-Loop-Helix repressors are essential for proper Drosophila and vertebrate development and are misregulated in many cancers. While a major step forward in understanding the molecular mechanism(s) surrounding Hairy-mediated repression was made with the identification of Groucho, Drosophila C-terminal binding protein (dCtBP), and Drosophila silent information regulator 2 (dSir2) as Hairy transcriptional cofactors, the identity of Hairy target genes and the rules governing cofactor recruitment are relatively unknown. We have used the chromatin profiling method DamID to perform a global and systematic search for direct transcriptional targets for Drosophila Hairy and the genomic recruitment sites for three of its cofactors: Groucho, dCtBP, and dSir2. Each of the proteins was tethered to Escherichia coli DNA adenine methyltransferase, permitting methylation proximal to in vivo binding sites in both Drosophila Kc cells and early embryos. This approach identified 40 novel genomic targets for Hairy in Kc cells, as well as 155 loci recruiting Groucho, 107 loci recruiting dSir2, and wide genomic binding of dCtBP to 496 loci. We also adapted DamID profiling such that we could use tightly gated collections of embryos (2-6 h) and found 20 Hairy targets related to early embryogenesis. As expected of direct targets, all of the putative Hairy target genes tested show Hairy-dependent expression and have conserved consensus C-box-containing sequences that are directly bound by Hairy in vitro. The distribution of Hairy targets in both the Kc cell and embryo DamID experiments corresponds to Hairy binding sites in vivo on polytene chromosomes. Similarly, the distributions of loci recruiting each of Hairy's cofactors are detected as cofactor binding sites in vivo on polytene chromosomes. We have identified 59 putative transcriptional targets of Hairy. In addition to finding putative targets for Hairy in segmentation, we find groups of targets suggesting roles for Hairy in cell cycle, cell growth, and morphogenesis, processes that must be coordinately regulated with pattern formation. Examining the recruitment of Hairy's three characterized cofactors to their putative target genes revealed that cofactor recruitment is context-dependent. While Groucho is frequently considered to be the primary Hairy cofactor, we find here that it is associated with only a minority of Hairy targets. The majority of Hairy targets are associated with the presence of a combination of dCtBP and dSir2. Thus, the DamID chromatin profiling technique provides a systematic means of identifying transcriptional target genes and of obtaining a global view of cofactor recruitment requirements during development.

Adenine↗
PubMed2026

Integration of methylome and transcriptome reveals age-associated signatures of stage-specific dynamics and regulatory remodeling in dogs.

Aging in mammals is characterized by widespread yet coordinated epigenetic alterations. However, integrative analyses of DNA methylation and gene expression in dogs remain largely unexplored, particularly within genetically homogeneous single-breed populations. To elucidate the molecular signatures of canine aging, we profiled the genome-wide methylome and transcriptome of clinically healthy beagle dogs representing three distinct age groups. Global methylation levels were highly conserved across individuals, yet both methylation and gene expression variability increased progressively with aging. This epigenetic drift was not stochastic but structured by genomic context, as reduced CpG-gene associations indicated a gradual loss of epigenetic control over transcription. We further observed stage-related methylation change patterns across the three age groups, including early-shift, late-shift, and progressive patterns. These groups showed partially overlapping but distinct hallmark associations, suggesting links to different age-related biological processes. Together, these findings indicate that canine epigenetic aging involves multifaceted molecular changes across adulthood and support dogs as a useful model for investigating conserved molecular signatures of aging.

Beagle dog↗
PubMed2026

Genome-wide chromatin recording resolves dynamic cell state changes.

Understanding how the chromatin state of a cell influences its future behavior is a major challenge throughout biology. However, most chromatin profiling methods are limited to endpoint assays. Here, we present LagTag, a method for recovery of earlier and endpoint chromatin states in the same mammalian cells. In this approach, transient activity of bacterial adenine methyltransferase fusions records the DNA-binding profiles of chromatin-associated proteins of interest at earlier time points. Subsequent tagmentation and sequencing recover the earlier chromatin profile from adenine methylation profiles, alongside endpoint profiles of endogenous chromatin-associated proteins. We verified that LagTag profiles aligned with those from established methods in mouse and human cells. We then applied LagTag to record and recover dynamic chromatin state transitions during mouse embryonic stem cell differentiation, capturing transcriptional signatures from pre- and post-differentiation time points within the same cell population. LagTag thus provides a foundation for temporally resolved chromatin profiling. A record of this paper's transparent peer review process is included in the supplemental information.

chromatin organization↗
PubMed2000

Rhodococcus koreensis sp. nov., a 2,4-dinitrophenol-degrading bacterium.

A 2,4-dinitrophenol-degrading bacterial strain, DNP505T, which was isolated from an industrial wastewater, was taxonomically studied by a polyphasic approach using phenotypic, chemotaxonomic and genetic methods. Strain DNP505T has a cell wall of chemotype IV containing meso-diaminopimelic acid, arabinose and galactose. The predominant menaquinone is MK-8(H2). Mycolic acids contain 43-53 carbon atoms. Strain DNP505T has a cellular fatty acid profile containing straight-chain saturated, unsaturated and 10-methyl-branched fatty acids and has C16:0 as the major fatty acid. The DNA G+C content is 66 mol%. Strain DNP505T formed a coherent cluster with Rhodococcus species in a phylogenetic inference based on 16S rDNA sequences. Interestingly, strain DNP505T was found to have two types of 16S rDNA sequence, which showed 10 bp sequence differences (99.3% nucleotide similarity). Its differences in some phenotypic characteristics and its genetic distinctiveness indicate that strain DNP505T is separate from Rhodococcus species described previously. It is therefore proposed that strain DNP505T should be placed in the genus Rhodococcus as a new species, Rhodococcus koreensis. The type strain of the new species is strain DNP505T (= KCTC 0569BPT = JCM 10743T).

2,4-Dinitrophenol↗
PubMed1995

Loss of heterozygosity in chromosomes 1, 5, 7 and 13 in mouse hepatoma detected by systematic genome-wide scanning using RLGS genetic map.

We have developed an RLGS-based scanning system to detect DNA alteration in tumor tissues, using 575 mapped spots/loci in a single gel. This system is very powerful for screening and identifying not only loss of heterozygosity (LOH) but also DNA methylation change. In this study, we applied this system to search for the LOH of hepatoma from an interspecific F1 hybrid between Mus spretus and C57BL/6 with SV40 early T antigen transgene connected to a mouse major urinary protein enhancer/promoter. Comparing the RLGS profiles of each tumor to that of the normal tissue showed significant LOH in chromosomes 1, 5, 7 and 13.

Animals↗
PubMed2005

Stress-specific signatures: expression profiling of p53 wild-type and -null human cells.

Gene expression responses of human cell lines exposed to a diverse set of stress agents were compared by cDNA microarray hybridization. The B-lymphoblastoid cell line TK6 (p53 wild-type) and its p53-null derivative, NH32, were treated in parallel to facilitate investigation of p53-dependent responses. RNA was extracted 4 h after the beginning of treatment when no notable decrease in cell viability was evident in the cultures. Gene expression signatures were defined that discriminated between four broad general mechanisms of stress agents: Non-DNA-damaging stresses (heat shock, osmotic shock, and 12-O-tetradecanoylphorbol 13-acetate), agents causing mainly oxidative stress (arsenite and hydrogen peroxide), ionizing radiations (neutron and gamma-ray exposures), and other DNA-damaging agents (ultraviolet radiation, methyl methanesulfonate, adriamycin, camptothecin, and cis-Platinum(II)diammine dichloride (cisplatin)). Within this data set, non-DNA-damaging stresses could be discriminated from all DNA-damaging stresses, and profiles for individual agents were also defined. While DNA-damaging stresses showed a strong p53-dependent element in their responses, no discernible p53-dependent responses were triggered by the non-DNA-damaging stresses. A set of 16 genes did exhibit a robust p53-dependent pattern of induction in response to all nine DNA-damaging agents, however.

B-Lymphocytes↗
PubMed2001

DNAinsight: a web based image processing system for large scale RLGS analysis.

DNA methylation occurring within the context of CpG site in the promoter element generally correlates with transcriptional silencing, delayed replication, condensed chromatin and mammalian genome imprinting (Nature Genet 24:101-2, 2000). We have been focusing attention on Restriction Landmark Genomic Scanning (RLGS) method as the powerful experimental procedure which can used to determine DNA methylation status change occurring on a genome simultaneously. To support systematic analysis with RLGS method, we have developed the automated processing algorithms for two-dimensional electrophoretograms of genomic DNA based on RLGS method (RLGS profile). Our powerful processing algorithms realize the automated spot recognition from RLGS profiles and the automated comparison of a huge number of such profiles. To make systematic RLGS analysis more easy and objective, we equipped our automated RLGS image processing algorithms with a WWW interface and relational database which stores the RLGS profile images together with their accompanying information, such as DNA sample specification, two-dimensional electrophoresis conditions and so forth. The web based RLGS image processing system "DNAinsight" can be accessed via URL http://www.methylome.jp/DNAinsight/.

Algorithms↗
PubMed2005

Methylation of tumor suppressor genes p16(INK4a), p27(Kip1) and E-cadherin in carcinogenesis.

Not only genomic mutations but also abnormal epigenetic methylation can significantly contribute to gene silencing and carcinogenesis. Methylation is particularly often observed in the CpG islands of the promoter regions in the regulatory genes. However, there are considerable differences in the incidence of methylation e.g. in the tumor suppressor genes, so that aberrant methylation of p16(INK4a) is relatively frequently observed in tumors, p27(Kip1) methylation is rare, and the incidence of E-cadherin methylation occurs at an intermediate rate. Although true genomic defects are generally much less common than methylation, parallel tendencies for both are often observed, probably reflecting the different levels of evolutionary advantage for tumor cells from inactivation of different genes. This also suggests that loss of p27 expression could be more a consequence of carcinogenesis, while lost p16 expression is a true oncogenic event. Due to the role of p27 in maintaining cellular quiescence, however, loss of its expression can still be a useful partial indicator of the aggressiveness of cancer. Loss of E-cadherin or its catenin partners of cellular adhesion will result in increasing invasiveness and metastatic potential of neoplastic cells but, because of several alternative routes to the same effect, incidence of lost expression for one component gene like E-cadherin does not need to be very high. Similarly, there must be a relatively high number of genes with modest or low incidence of aberrant silencing by methylation, to reflect multiple alternatives for the multistep process of carcinogenesis. Nevertheless, methylation of different genes also shows characteristic differences between different cancer and tumor types, and the epigenetic methylation patterns therefore have considerable diagnostic and prognostic potential. Realising this potential requires efficient methods for profiling the status of methylation. Such profiling methods have only recently become available and are still under relatively rapid development.

Cadherins↗
PubMed2003

Expression profiling reveals that methylation of TIMP3 is involved in uveal melanoma development.

Uveal melanoma is associated with a high tumor-related mortality due to the propensity to develop metastases. The mechanisms that are responsible for malignant dissemination are largely unknown and need to be explored to facilitate diagnosis and treatment of metastases. To identify molecules involved in dissemination, we used cell lines derived from a primary uveal melanoma and 2 liver metastases from the same patient as a model for tumor progression. Using a microarray representing 1,176 genes, we identified 63 differentially expressed genes. Forty genes showed a higher expression and 23 showed a lower expression in the primary cell line compared to the metastasis cell lines. These genes are involved in processes like angiogenesis, apoptosis, macrophage stimulation, and extracellular matrix regulation. In contrast, the 2 liver metastasis cell lines produced nearly identical expression profiles. Demethylation of the primary melanoma cell line by 5-aza-2'deoxycytidine treatment revealed that 19 genes were suppressed by hypermethylation. An important finding was the 5-fold decreased expression of TIMP3 in the metastatic cell lines, a molecule involved in extracellular matrix degradation. We demonstrate in the cell lines that TIMP3 expression is regulated by methylation. These observations were confirmed in primary uveal melanoma and suggests a role for TIMP3 in uveal melanoma development.

Azacitidine↗
PubMed2003

Methyl-CpG binding proteins identify novel sites of epigenetic inactivation in human cancer.

Methyl-CpG binding proteins (MBDs) mediate histone deacetylase-dependent transcriptional silencing at methylated CpG islands. Using chromatin immunoprecitation (ChIP) we have found that gene-specific profiles of MBDs exist for hypermethylated promoters of breast cancer cells, whilst a common pattern of histone modifications is shared. This unique distribution of MBDs is also characterized in chromosomes by comparative genomic hybridization of immunoprecipitated DNA and immunolocalization. Most importantly, we demonstrate that MBD association to methylated DNA serves to identify novel targets of epigenetic inactivation in human cancer. We combined the ChIP assay of MBDs with a CpG island microarray (ChIP on chip). The scenario revealed shows that, while many genes are regulated by multiple MBDs, others are associated with a single MBD. These target genes displayed methylation- associated transcriptional silencing in breast cancer cells and primary tumours. The candidates include the homeobox gene PAX6, the prolactin hormone receptor, and dipeptidylpeptidase IV among others. Our results support an essential role for MBDs in gene silencing and, when combined with genomic strategies, their potential to 'catch' new hypermethylated genes in cancer.

5-Methylcytosine↗