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. 2017 Feb;9(2):189-207.
doi: 10.2217/epi-2016-0138. Epub 2016 Dec 2.

Trans-acting Epigenetic Effects of Chromosomal Aneuploidies: Lessons From Down Syndrome and Mouse Models

Free PMC article

Trans-acting Epigenetic Effects of Chromosomal Aneuploidies: Lessons From Down Syndrome and Mouse Models

Catherine Do et al. Epigenomics. .
Free PMC article

Erratum in

  • Erratum.
    Epigenomics. 2017 Mar;9(3):369. doi: 10.2217/epi-2016-0138e1. Epub 2017 Feb 17. Epigenomics. 2017. PMID: 28234027 Free PMC article. No abstract available.


An important line of postgenomic research seeks to understand how genetic factors can influence epigenetic patterning. Here we review epigenetic effects of chromosomal aneuploidies, focusing on findings in Down syndrome (DS, trisomy 21). Recent work in human DS and mouse models has shown that the extra chromosome 21 acts in trans to produce epigenetic changes, including differential CpG methylation (DS-DM), in specific sets of downstream target genes, mostly on other chromosomes. Mechanistic hypotheses emerging from these data include roles of chromosome 21-linked methylation pathway genes (DNMT3L and others) and transcription factor genes (RUNX1, OLIG2, GABPA, ERG and ETS2) in shaping the patterns of DS-DM. The findings may have broader implications for trans-acting epigenetic effects of chromosomal and subchromosomal aneuploidies in other human developmental and neuropsychiatric disorders, and in cancers.

Keywords: CTCF; DNA methylation; Down syndrome; aneuploidy; cancer; chromatin; developmental disorders; methyltransferases; mouse models; transcription factors.

Conflict of interest statement

Financial & competing interests disclosure

This work was supported by grants from the NIH to B Tycko (R01AG036040, R01AG035020, P01-HD035897), by a grant from the Down Syndrome Research Foundation-UK to B Tycko, by a grant to the Herbert Irving Comprehensive Cancer Center at Columbia University (P30CA013696), by grants from the NIH to E Yu (R01NS66072 and R01HL91519) and to Roswell Park Cancer Institute (P3016056), and by a grant from the Children's Guild Foundation to E Yu. The authors have no other relevant affiliations or financial involvement with any organization or entity with a financial interest in or financial conflict with the subject matter or materials discussed in the manuscript apart from those disclosed.

No writing assistance was utilized in the production of this manuscript.


<b>Figure 1.</b>
Figure 1.. Chromosomal aneuploidies and disease pathogenesis: role of trans-acting epigenetic effects.
Diagram of chromosome 21 (Hsa21), which is trisomic in Down syndrome. Effector genes on the triplicated Hsa21 act on downstream target genes, mostly on other chromosomes, both by acute transcriptional effects and via epigenetic effects, including alterations in DNA methylation that can propagate to daughter cells in growing and self-renewing tissues, to produce biological phenotypes. DS-DM: Differential CpG methylation.
<b>Figure 2.</b>
Figure 2.. Altered DNA methylation in whole blood cells and T lymphocytes with Ts21: consistency in independent studies and examples of biologically relevant genes with differential CpG methylation.
(A) Heatmap showing reproducibility of DS-DM loci in whole blood from the study by Bacalini et al. [57] and T cells from our study [59]. Raw data from each dataset were processed separately using the same pipeline: after normalization by β-mixture quantile normalization using the ChAMP Bioconductor package, probes with more than 20% of missing values or detection p-value > 0.05, probes mapping to the XY chromosomes and probes querying CpGs that overlap with common single-nucleotide polymorphisms (dbSNP138, minor allele frequency >1%) were filtered out. Raw data were downloaded from NCBI-GEO (GSE74486 for T cells and GSE52588 for whole-blood data). Differentially methylated CpGs were defined for all datasets as nominal p-value <0.005, absolute methylation difference >0.15. In our meta-analysis, we only considered strong DS-DM CpGs, with at least two DS-DM CpGs in 500 bp windows. We identified 1083 DS-DM CpGs in PBL and 1300 in T cells, with 21% overlap. Although all enrichment analyses were done using these entire DS-DM CpG sets, for clarity, the heatmap only shows the top ranked DS-DM CpGs (based on p-values), all with an absolute methylation difference >0.2. There are more hypermethylated loci (red color in DS) than hypomethylated loci (blue color in DS) in the whole blood cells and T cells, although a greater bias toward hypermethylation is found in brain tissues [59]. (B) Scatter plots showing strong DS-DM in RUNX1, SH3BP2, TMEM131 and ZDHHC14 in both whole blood and T cells. For each locus, fractional methylation values for the strongest DS-DM CpG are shown. The horizontal lines represent the average fractional methylation. T-test p-values for DM in DS versus controls are indicated. (C) Map and Bis-Seq validations of DS-DM in ZDHHC14. The bis-seq amplicons cover the DS-DM CpGs identified by Illumina 450K BeadChip array and flanking regions. The data reveal a discrete DS-DM region (amplicons 2 & 3) spanning 600 bp and overlapping an enhancer region marked by H3K4me1, immediately adjacent to a CTCF-bound insulator element. C: Control; DS: Down syndrome; DS-DM: Differential CpG methylation; PBL: Peripheral blood leukocyte.
<b>Figure 3.</b>
Figure 3.. Pan-tissue and multitissue differential CpG methylation loci with hypermethylation are enriched in CTCF motifs, and T lymphocyte differential CpG methylation loci with hypomethylation are enriched in RUNX motifs.
(A) Bar graph showing enrichment in TF motifs among the whole blood hypomethylated DS-DM CpGs. Coordinates of occurrences of 90 motifs were downloaded from ENCODE to identify TF motifs within 100 bp of each CpG queried by the 450K BeadChips. Unlike in Figure 2 where only the top-ranked DM-CpGs are shown, we assessed enrichment using logistic regressions on all DS-DM CpGs in clusters of at least two DS-DM CpGs within 500 bp passing nominal p-value < 0.005 and fractional methylation differences <0.15. Only significant enrichment (nominal p-value < 0.05) of TF motifs with more than five occurrences in the DS-DM regions are shown. (B) Bar graph showing enrichment in TF motifs among the T-cell hypomethylated DS-DM CpGs. Methods as in (A). (C) List of pan and multitissue DS-DM genes. Raw data from five public datasets were processed separately using the same pipeline and criteria as described in Figure 1. Raw data were downloaded from NCBI-GEO (GSE74486, GSE63347, GSE73747, GSE52588 and GSE66210). For each study, only tissues with at least three control and three DS samples were assessed. DM CpGs were defined for all datasets as CpGs in clusters of at least two DS-DM CpGs within 500 bp passing nominal p-value < 0.005 and methylation difference < 0.15. We merged the DM sets into five tissue sets: the brain datasets include samples from adult frontal cortex (FC) and cerebellum; fetal brain datasets samples from fetal cerebrum, FC and temporal cortex (TC); epithelial dataset samples from buccal epithelial cells; and blood dataset samples from T cells and PBL. Hyper and hypomethylated CpG sets from each tissue were overlapped using 1 kb windows, since differential methylation at adjacent CpGs often reflects methylation changes in the same regulatory element. Multitissue DS-DM CpGs were defined as CpGs with DS-DM within 500 bp present in at least three different tissues. Overall, we found 157 multitissue DS-DM CpGs, located in two genes (8 CpGs) with pan-tissue hypermethylation, 22 genes (141 CpGs) with multitissue hypermethylation and one gene (8 CpGs) with multitissue hypomethylation. (D) Bar graph showing enrichment in TF motifs among the pan or multitissues hypermethylated DS-DM CpGs. Methods were as in (A). DS-DM: Differential CpG methylation; PBL: Peripheral blood leukocyte; TF: Transcription factor.
<b>Figure 4.</b>
Figure 4.. Mouse models for dissecting the mechanisms and biological consequences of differential CpG methylation.
(A) The Hsa21 syntenic regions triplicated in major mouse models of DS are shown, which can serve as models for dissecting the mechanisms and biological consequences of DS-DM. The indicated regions of Hsa21 are syntenically conserved with three genomic regions separately located on mouse chromosome 10 (Mmu10), Mmu16 and Mmu17. Dp(16)1Yey and Ts65Dn mice carry more triplicated Hsa21 gene orthologs than any other triplication mouse models of DS. The genomic locations of methylation pathway genes and transcription factor genes, and their presence or absence in each of these models, are indicated. (B) Enrichment of Hsa21-orthologous TF gene consensus-binding motifs among Ts65Dn DS-DM CpGs. Enrichment analysis was performed using HOMER software applied to whole-genome bisulfite sequencing (WGBS) data from cerebral gray matter of Ts65Dn versus wt littermate mice [do c et al. 2016, manuscript in preparation]. DS-DM CpGs were defined as in [59]. As discussed in the main text, the WGBS data from these mouse brains showed a significant excess of hypermethylated DM regions, but revealed a somewhat larger percentage of hypomethylated loci than were seen in the 450K Beadchip data from human brains. Strong enrichment is observed in sites recognized by TFs encoded by genes in the Mmu16 duplicated regions (bold font). Enrichment among hypomethylated DS-DM is suggestive of passive demethylation due to the TF occupancy, while enrichment among hypermethylated DS-DM is suggestive of active methylation by recruitment of DNMTs, as described in other settings [74]. Significant, though weaker, enrichment in the E26 transformation-specific family of transcription factor (ETS) motif was also observed in our prior WGBS data [59] from whole cerebrums of Dp(16) (p-value = 10-03) but not of Dp(10). Enrichment in transcription factor-binding sites (TFBSs) was defined as FDR <0.05 and fold-enrichment >1.2. The 10 top-ranked enriched TFBS sites are shown for hypermethylated and hypomethylated DS-DM CpGs. The finding of enrichment of three of these TFBS classes (all ETS family) among the hypermethylated DM loci and one (not ETS family) among the hypomethylated DM loci further supports a nonrandom mechanistic connection. DS-DM: Differential CpG methylation; TF: Transcription factor.
<b>Figure 5.</b>
Figure 5.. Similarities between differential CpG methylation in human Down syndrome versus control brains and differential methylation in brains from the chromosomally engineered mouse models of Down syndrome illustrated by results in the PCDHGA2/Pcdhga2 gene clusters.
Map of the PCDHGA cluster. Differential CpG methylation in human DS (DS-DM) CpGs in the human 450K data and DM CpGs in the whole-genome bisulfite sequencing (WGBS) data from the mouse models were defined as in [59]. WGBS for Dp(10)1Yey and Dp(16)1Yey was performed on whole cerebrums, while WGBS for Ts65Dn was performed on macrodissected cerebral gray matter. The Y-axes show fractional differences in CpG methylation. WGBS median read depth = 30 in all experiments; a total of 1074 CpGs were covered at >20 read depth in this chromosomal segment; only the CpGs passing our published criteria for significant differential methylation [59] are indicated by the bars. DS-DM regions coincide with CTCF ChIP-seq peaks – consistent with the enrichment in CTCF motif sites that we and others have found among DS-DM loci. CTCF ChIP-seq data from human astrocytes and mouse whole brain were downloaded from ENCODE. C: Control; DS: Down syndrome; WGBS: Whole-genome bisulfite sequencing; wt: Wild-type.

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