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Global DNA hypermethylation in down syndrome placenta.

Jin S, Lee YK, Lim YC, Zheng Z, Lin XM, Ng DP, Holbrook JD, Law HY, Kwek KY, Yeo GS, Ding C - PLoS Genet. (2013)

Bottom Line: We hypothesize that such global hypermethylation may be mediated by down-regulation of TET family genes involved in DNA demethylation, and down-regulation of REST/NRSF involved in transcriptional and epigenetic regulation.Genes located on chr21 were up-regulated by an average of 53% in DS compared to normal villi, while genes with promoter hypermethylation were modestly down-regulated.Our data suggest that global epigenetic changes may occur early in development and contribute to DS phenotypes.

View Article: PubMed Central - PubMed

Affiliation: Growth, Development and Metabolism Program, Singapore Institute for Clinical Sciences, Agency for Science, Technology and Research, Singapore.

ABSTRACT
Down syndrome (DS), commonly caused by an extra copy of chromosome 21 (chr21), occurs in approximately one out of 700 live births. Precisely how an extra chr21 causes over 80 clinically defined phenotypes is not yet clear. Reduced representation bisulfite sequencing (RRBS) analysis at single base resolution revealed DNA hypermethylation in all autosomes in DS samples. We hypothesize that such global hypermethylation may be mediated by down-regulation of TET family genes involved in DNA demethylation, and down-regulation of REST/NRSF involved in transcriptional and epigenetic regulation. Genes located on chr21 were up-regulated by an average of 53% in DS compared to normal villi, while genes with promoter hypermethylation were modestly down-regulated. DNA methylation perturbation was conserved in DS placenta villi and in adult DS peripheral blood leukocytes, and enriched for genes known to be causally associated with DS phenotypes. Our data suggest that global epigenetic changes may occur early in development and contribute to DS phenotypes.

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Gene expression changes in DS.For each gene, average expression values (RPKM) were calculated for both normal and DS samples. Only genes with RPKM ≥0.5 in at least one sample group were used for further analysis. Gene expression changes in DS were represented by log2 (Average DS samples expression/Average normal samples expression). (A) PDF distribution for gene expression changes for all genes, chr21 genes, hypermethylated genes and REST target genes. Genes located on chr21 were up-regulated by an average of 53% in DS. The hypermethylated genes were down-regulated, as evidenced by a left shift of the PDF curve (p<0.05, Wilcoxon rank-sum test, two-sided). The genes targeted by REST were marginally up-regulated (p = 0.06, Wilcoxon rank-sum test, two-sided). (B) Repression of gene expression by promoter hypermethylation was more prominent in promoters that were originally at lower methylation levels in normal samples. Each data point represents one hypermethylated promoter. X axis is the average methylation level in the normal samples for each promoter. Y axis is the gene expression ratio between DS and normal (log2 transformed).
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pgen-1003515-g003: Gene expression changes in DS.For each gene, average expression values (RPKM) were calculated for both normal and DS samples. Only genes with RPKM ≥0.5 in at least one sample group were used for further analysis. Gene expression changes in DS were represented by log2 (Average DS samples expression/Average normal samples expression). (A) PDF distribution for gene expression changes for all genes, chr21 genes, hypermethylated genes and REST target genes. Genes located on chr21 were up-regulated by an average of 53% in DS. The hypermethylated genes were down-regulated, as evidenced by a left shift of the PDF curve (p<0.05, Wilcoxon rank-sum test, two-sided). The genes targeted by REST were marginally up-regulated (p = 0.06, Wilcoxon rank-sum test, two-sided). (B) Repression of gene expression by promoter hypermethylation was more prominent in promoters that were originally at lower methylation levels in normal samples. Each data point represents one hypermethylated promoter. X axis is the average methylation level in the normal samples for each promoter. Y axis is the gene expression ratio between DS and normal (log2 transformed).

Mentions: We next performed RNA-Seq analysis in five normal and four DS placenta villi samples (Table S1). Genes located on chr21 were up-regulated by an average of 53% in DS (Figure 3A), consistent with previous reports [2]–[4]. _ENREF_10_ENREF_10Many well-studied genes such as BACH1, SOD1, TIAM1, ITSN1, DSCR1/RCAN1, and DYRK1A located on chr21 were up-regulated (Table S4). A total of 589 genes across all autosomes were hypermethylated in the promoters in DS. Out of the 589 genes, 207 genes passed the expression threshold (reads per kilobase per million mapped reads, RPKM≥0.5) and are located on autosomes other than chr21. Significant down-regulation of gene expression was observed for the 207 genes (p<0.05, Wilcoxon rank-sum test, two-sided). Interestingly, the association between promoter hypermethylation and gene expression repression was more pronounced for promoters with lower DNA methylation in the normal samples, suggesting that increased methylation in originally unmethylated promoters is likely to have a bigger impact on gene expression (Figure 3B). We further validated four genes (CES1, TFAP2E, CDH13, NDN) that showed increased promoter methylation and decreased gene expression, with EpiTYPER assays and quantitative real-time PCR, with a new set of gestational age matched samples (Table S3, Figure S6A–S6B).


Global DNA hypermethylation in down syndrome placenta.

Jin S, Lee YK, Lim YC, Zheng Z, Lin XM, Ng DP, Holbrook JD, Law HY, Kwek KY, Yeo GS, Ding C - PLoS Genet. (2013)

Gene expression changes in DS.For each gene, average expression values (RPKM) were calculated for both normal and DS samples. Only genes with RPKM ≥0.5 in at least one sample group were used for further analysis. Gene expression changes in DS were represented by log2 (Average DS samples expression/Average normal samples expression). (A) PDF distribution for gene expression changes for all genes, chr21 genes, hypermethylated genes and REST target genes. Genes located on chr21 were up-regulated by an average of 53% in DS. The hypermethylated genes were down-regulated, as evidenced by a left shift of the PDF curve (p<0.05, Wilcoxon rank-sum test, two-sided). The genes targeted by REST were marginally up-regulated (p = 0.06, Wilcoxon rank-sum test, two-sided). (B) Repression of gene expression by promoter hypermethylation was more prominent in promoters that were originally at lower methylation levels in normal samples. Each data point represents one hypermethylated promoter. X axis is the average methylation level in the normal samples for each promoter. Y axis is the gene expression ratio between DS and normal (log2 transformed).
© Copyright Policy
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC3675012&req=5

pgen-1003515-g003: Gene expression changes in DS.For each gene, average expression values (RPKM) were calculated for both normal and DS samples. Only genes with RPKM ≥0.5 in at least one sample group were used for further analysis. Gene expression changes in DS were represented by log2 (Average DS samples expression/Average normal samples expression). (A) PDF distribution for gene expression changes for all genes, chr21 genes, hypermethylated genes and REST target genes. Genes located on chr21 were up-regulated by an average of 53% in DS. The hypermethylated genes were down-regulated, as evidenced by a left shift of the PDF curve (p<0.05, Wilcoxon rank-sum test, two-sided). The genes targeted by REST were marginally up-regulated (p = 0.06, Wilcoxon rank-sum test, two-sided). (B) Repression of gene expression by promoter hypermethylation was more prominent in promoters that were originally at lower methylation levels in normal samples. Each data point represents one hypermethylated promoter. X axis is the average methylation level in the normal samples for each promoter. Y axis is the gene expression ratio between DS and normal (log2 transformed).
Mentions: We next performed RNA-Seq analysis in five normal and four DS placenta villi samples (Table S1). Genes located on chr21 were up-regulated by an average of 53% in DS (Figure 3A), consistent with previous reports [2]–[4]. _ENREF_10_ENREF_10Many well-studied genes such as BACH1, SOD1, TIAM1, ITSN1, DSCR1/RCAN1, and DYRK1A located on chr21 were up-regulated (Table S4). A total of 589 genes across all autosomes were hypermethylated in the promoters in DS. Out of the 589 genes, 207 genes passed the expression threshold (reads per kilobase per million mapped reads, RPKM≥0.5) and are located on autosomes other than chr21. Significant down-regulation of gene expression was observed for the 207 genes (p<0.05, Wilcoxon rank-sum test, two-sided). Interestingly, the association between promoter hypermethylation and gene expression repression was more pronounced for promoters with lower DNA methylation in the normal samples, suggesting that increased methylation in originally unmethylated promoters is likely to have a bigger impact on gene expression (Figure 3B). We further validated four genes (CES1, TFAP2E, CDH13, NDN) that showed increased promoter methylation and decreased gene expression, with EpiTYPER assays and quantitative real-time PCR, with a new set of gestational age matched samples (Table S3, Figure S6A–S6B).

Bottom Line: We hypothesize that such global hypermethylation may be mediated by down-regulation of TET family genes involved in DNA demethylation, and down-regulation of REST/NRSF involved in transcriptional and epigenetic regulation.Genes located on chr21 were up-regulated by an average of 53% in DS compared to normal villi, while genes with promoter hypermethylation were modestly down-regulated.Our data suggest that global epigenetic changes may occur early in development and contribute to DS phenotypes.

View Article: PubMed Central - PubMed

Affiliation: Growth, Development and Metabolism Program, Singapore Institute for Clinical Sciences, Agency for Science, Technology and Research, Singapore.

ABSTRACT
Down syndrome (DS), commonly caused by an extra copy of chromosome 21 (chr21), occurs in approximately one out of 700 live births. Precisely how an extra chr21 causes over 80 clinically defined phenotypes is not yet clear. Reduced representation bisulfite sequencing (RRBS) analysis at single base resolution revealed DNA hypermethylation in all autosomes in DS samples. We hypothesize that such global hypermethylation may be mediated by down-regulation of TET family genes involved in DNA demethylation, and down-regulation of REST/NRSF involved in transcriptional and epigenetic regulation. Genes located on chr21 were up-regulated by an average of 53% in DS compared to normal villi, while genes with promoter hypermethylation were modestly down-regulated. DNA methylation perturbation was conserved in DS placenta villi and in adult DS peripheral blood leukocytes, and enriched for genes known to be causally associated with DS phenotypes. Our data suggest that global epigenetic changes may occur early in development and contribute to DS phenotypes.

Show MeSH
Related in: MedlinePlus