Unknown,Transcriptomics,Genomics,Proteomics

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Methylation profiling of differentially expressed regions between C57BL/6 and BALB/c in bone marrow derived macrophages

ABSTRACT: DNA-methylation is a vital epigenetic mark that participates in establishing and maintaining chromatin structures and in regulating gene transcription during mammalian development and cellular differentiation. Inter-individual differences in methylation patterns may represent a major source of phenotypic variation, however, the determinants, inheritance, extent, and consequences of such differences are poorly understood. Here we have analysed methylation profiles of immune cells from two inbreed mouse strains (C57BL/6 & BALB/c) that represent prototypic models for Th1- or Th2-dominated immune responses. Using a methyl-CpG immunoprecipitation approach, genomes were fractionated into methylated and unmethylated genome pools and separately analysed at 180 genomic regions (covering 28 Mb of the mouse genome) that were selected based on differential gene expression between both mouse strains. Differentially methylated regions are detected by analyzing array probes for diametrically opposed enrichment behaviour between both hybridizations (e.g. a region that is relatively enriched in the unmethylated pool of BALB/c and shows reverse enrichment behaviour in the methylated pool is considered hypomethylated in BALB/c). The integrated analysis of hypo and hypermethylation profiles allowed the identification of several hundred differentially methylated regions, but also uncovered regions that were duplicated in one strain, contained single nucleotide polymorphisms or micro- and macro-deletions. Keywords: MCIp-on-Chip; comparative genomic hybridization Methylation profiles of regions, harbouring differentially expressed genes in BMM, were compared between C57BL/6 and BALB/c. Two independently grown, harvested and MCIp treated gDNA preparations were used for each mouse strain (Two biological replicates; One replicate per array). Hypo- and hypermethylated pools serve as diametrically oposed technical replicates (mirror approach).

ORGANISM(S): Mus musculus

SUBMITTER: Michael Rehli

PROVIDER: E-GEOD-14463 | biostudies-arrayexpress |

REPOSITORIES: biostudies-arrayexpress

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Publications

Allele-specific DNA methylation in mouse strains is mainly determined by cis-acting sequences.

Schilling Elmar E El Chartouni Carol C Rehli Michael M

Genome research 20090817 11

DNA methylation is a vital epigenetic mark that participates in establishing and maintaining chromatin structures and regulates gene transcription during mammalian development and cellular differentiation. Differences in epigenetic patterns between individuals may contribute to phenotypic variation and disease susceptibility; however, little is known about the extent of such variation or how different epigenetic patterns are established. Here we have compared DNA methylation profiles of macropha ...[more]

PMID: 19687144

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Project description:We have previously developed an approach that fractionates genomic DNA fragments depending on their CpG density (methyl-CpG-immunoprecipitation, MCIp), and adapted this approach to identify regions that are differentially methylated in the two closely related regulatory T-cells (Treg cells) and conventional T-cells (Tconv cells). Because Treg cells naturally occur at a relatively low frequency, we used a previously established protocol to expand Treg cells from a stable naÃ¯ve Treg population that is characterized by the co-expression of CD4, CD25 and CD45RA. We separated gDNA of both expanded T cell lineages (Tregexp and Tconvexp) into unmethylated (CpG) and methylated pools (mCpG) using MCIp and compared cell type-specific differences in DNA methylation by co-hybridization of the two umethylated or the two methylated DNA subpopulations of Treg and Tconv, respectively, to these locus-wide custom tiling arrays. As enriched DNA-fragments from a cell type in the methylated fraction should be depleted in the unmethylated fraction, the signal intensities in CpG pool and mCpG pool hybridizations should complement themselves (âMirror-Imageâ approach) and thereby allow the identification of differentially methylated regions (DMR). Because we expected to find lineage-specific methylation differences with greater probability in regions associated with differential transcriptional activity, we limited our analysis to gene loci that showed cell type-specific gene expression in Treg versus Tconv cells plus a handful of control regions that were equally expressed in both cell types. The microarray used in this study covered 12 megabases of the human genome and contained 69 regions (with a median size of 100.000 kb) and 128 proximal promoter regions and 181 genes, which included a number of well known and functionally relevant genes like CD40LG, IFNG, FOXP3, IL2RA and CTLA4. Keywords: MCIp-on-chip; comparative genomic hybridization With MCIp gDNA from Treg or Tconv cells was separated into hypo- and hypermethylated pools. On each array, wheather the two hypomethylated fractions- one from Treg, the other from Tconv cells- or the two hypermethylated fractions were cohybridized. Two biological replicates.

Dataset Information

Methylation profiling of differentially expressed regions between C57BL/6 and BALB/c in bone marrow derived macrophages

Publications

Allele-specific DNA methylation in mouse strains is mainly determined by cis-acting sequences.

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