Dataset Information

Genome-wide methylation analysis of retrocopy-associated CpG islands and their genomic environment.

ABSTRACT: Gene duplication by retrotransposition, i.e., the reverse transcription of an mRNA and integration of the cDNA into the genome, is an important mechanism in evolution. Based on whole-genome bisulfite sequencing of monocyte DNA, we have investigated the methylation state of all CpG islands (CGIs) associated with a retrocopy (n = 1,319), their genomic environment, as well as the CGIs associated with the ancestral genes. Approximately 10% of retrocopies are associated with a CGI. Whereas almost all CGIs of the human genome are unmethylated, 68% of the CGIs associated with a retrocopy are methylated. In retrocopies resulting from multiple retrotranspositions of the same ancestral gene, the methylation state of the CGI often differs. There is a strong positive correlation between the methylation state of the CGI/retrocopy and their genomic environment, suggesting that the methylation state of the integration site determined the methylation state of the CGI/retrocopy, or that methylation of the retrocopy by a host defense mechanism has spread into the adjacent regions. Only a minor fraction of CGI/retrocopies (n = 195) has intermediate methylation levels. Among these, the previously reported CGI/retrocopy in intron 2 of the RB1 gene (PPP1R26P1) as well as the CGI associated with the retrocopy RPS2P32 identified in this study carry a maternal methylation imprint. In conclusion, these findings shed light on the evolutionary dynamics and constraints of DNA methylation.

SUBMITTER: Grothaus K

PROVIDER: S-EPMC4854546 | biostudies-literature | 2016 Mar

REPOSITORIES: biostudies-literature

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Genome-wide methylation analysis of retrocopy-associated CpG islands and their genomic environment.

Grothaus Katrin K Kanber Deniz D Gellhaus Alexandra A Mikat Barbara B Kolarova Julia J Siebert Reiner R Wieczorek Dagmar D Horsthemke Bernhard B

Epigenetics 20160218 3

Gene duplication by retrotransposition, i.e., the reverse transcription of an mRNA and integration of the cDNA into the genome, is an important mechanism in evolution. Based on whole-genome bisulfite sequencing of monocyte DNA, we have investigated the methylation state of all CpG islands (CGIs) associated with a retrocopy (n = 1,319), their genomic environment, as well as the CGIs associated with the ancestral genes. Approximately 10% of retrocopies are associated with a CGI. Whereas almost all ...[more]

PMID: 26890210

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Project description:BackgroundHypermethylation of promoter CpG islands is strongly correlated to transcriptional gene silencing and epigenetic maintenance of the silenced state. As well as its role in tumor development, CpG island methylation contributes to the acquisition of resistance to chemotherapy. Differential Methylation Hybridisation (DMH) is one technique used for genome-wide DNA methylation analysis. The study of such microarray data sets should ideally account for the specific biological features of DNA methylation and the non-symmetrical distribution of the ratios of unmethylated and methylated sequences hybridised on the array. We have therefore developed a novel algorithm tailored to this type of data, Methylation Linear Discriminant Analysis (MLDA).ResultsMLDA was programmed in R (version 2.7.0) and the package is available at CRAN 1. This approach utilizes linear regression models of non-normalised hybridisation data to define methylation status. Log-transformed signal intensities of unmethylated controls on the microarray are used as a reference. The signal intensities of DNA samples digested with methylation sensitive restriction enzymes and mock digested are then transformed to the likelihood of a locus being methylated using this reference. We tested the ability of MLDA to identify loci differentially methylated as analysed by DMH between cisplatin sensitive and resistant ovarian cancer cell lines. MLDA identified 115 differentially methylated loci and 23 out of 26 of these loci have been independently validated by Methylation Specific PCR and/or bisulphite pyrosequencing.ConclusionMLDA has advantages for analyzing methylation data from CpG island microarrays, since there is a clear rational for the definition of methylation status, it uses DMH data without between-group normalisation and is less influenced by cross-hybridisation of loci. The MLDA algorithm successfully identified differentially methylated loci between two classes of samples analysed by DMH using CpG island microarrays.

Project description:BackgroundThe development and application of CRISPR technologies for the modification of the genome are rapidly expanding. Advances in the field describe new CRISPR components that are strategically engineered to improve the precision and reliability of CRISPR editing within the genome sequence. Genome modification using induced genome breaks that are targeted and mediated by CRISPR components leverage cellular mechanisms for repair like homology directed repair (HDR) to incorporate genomic edits with increased precision.ResultsIn this report, we describe the gain of methylation at typically hypomethylated CpG island (CGI) locations affected by the CRISPR-mediated incorporation of donor DNA using HDR mechanisms. With characterization of CpG methylation patterns using whole genome bisulfite sequencing, these CGI methylation disruptions trace the insertion of the donor DNA during the genomic edit. These insertions mediated by homology-directed recombination disrupt the generational methylation pattern stability of the edited CGI within the cells and their cellular lineage within the animal strain, persisting across generations. Our approach describes a statistically based workflow for indicating locations of modified CGIs and provides a mechanism for evaluating the directed modification of the methylome of the affected CGI at the CpG-level.ConclusionsWith advances in genome modification technology comes the need to detect the level and persistence of methylation change that modifications to the genomic sequence impose upon the collaterally edited methylome. Any modification of the methylome of somatic or germline cells could have implications for gene regulation mechanisms governed by the methylation patterns of CGI regions in the application of therapeutic edits of more sensitively regulated genomic regions. The method described here locates the directed modification of the mouse epigenome that persists over generations. While this observance would require supporting molecular observations such as direct sequence changes or gene expression changes, the observation of epigenetic modification provides an indicator that intentionally directed genomic edits can lead to collateral, unintentional epigenomic changes post modification with generational persistence.

Dataset Information

Genome-wide methylation analysis of retrocopy-associated CpG islands and their genomic environment.

Publications

Genome-wide methylation analysis of retrocopy-associated CpG islands and their genomic environment.

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