Hvilsom2013

Hvilsom201

Differentially methylated CpG sites in each great ape genus.

<p>Heat maps showing genus specific differentially methylated CpG sites. We found 2,284 human-specific differentially methylated CpGs, 1,245 specific to <i>Pan</i> species, 1,374 specific to <i>Gorilla</i> species and 5,501 changes specific to <i>Pongo</i> species. Each vertical line represents a single CpG, with each row showing the β-value obtained in each individual tested.</p

A significant relationship between changes in promoter methylation and protein evolution between human and chimpanzee.

<p>We performed a comparison of alterations in promoter methylation with (A) the frequency of amino-acid alterations and (B) the relative rate of coding to non-coding variation with genes (K_A/K_I) between human and chimpanzee. Using both metrics we observed a significant association between the rate of protein evolution and epigenetic regulatory changes. P-values are based on 1,000 permutations (Differentially methylated genes, n = 745; genes without significant changes in methylation, n = 6,507).</p

Location of differential methylation in primate genomes.

<p>(A) Distribution of 99,191 CpG sites interrogated in all great ape species. Left: Gene-centric functional distribution of methylation changes : 1,500 bp upstream of gene TSSs, 200 bp upstream of TSSs, 5′UTR, 1^st exon, gene body, 3′ UTR and intergenic. Right: CpG-island centric distribution: CpG island, shore (±2 kb flanking CpG islands), shelf (2–4 kb from CpG islands). (B) A non-random distribution of methylation changes in recent primate evolution. We observe an excess of differential CpG methylation within the first 1,500 bp upstream of gene TSSs, gene bodies, intergenic regions, shore regions flanking CpG islands and non-CpG island regions. In contrast DNA methylation tends to be relatively conserved close to gene transcription start sites (−200 bp of TSS to 1^st exon), and in the body of CpG islands. Each bar shows the relative enrichment for differential methylation versus that expected under a null distribution. * denotes p<0.0001 (permutation test). (C) Density plot showing the distribution of methylation levels of differentially methylated sites compared to that in the rest of the genome. Sites of evolutionary change among great apes have a significantly different distribution (p = 2.2×10⁻¹⁶ Kolmogorov-Smirnov test).</p

Examples of differentially methylated genes.

<p>Each data point represents the mean β-value of the group and whiskers show 2 standard deviations above and below the mean. (A) <i>ARTN</i> is a neurotrophic factor and it shows hypermethylation of 3 CpG sites associated with the long isoform specifically in human. (B) <i>COL2A1</i> shows hypermethylation of 4 CpG sites at the promoter specifically in human. This gene encodes the alpha-1 chain of type II collagen, which is found primarily in cartilage, the inner ear and the vitreous humor of the eye. (C) <i>PGAM2</i> shows hypomethylation of CpG sites at the promoter specifically in human. <i>PGAM2</i> is an enzyme involved in the glycolitic pathway, mutations in which are associated with muscle cramping and intolerance for strenuous exercise. (D) <i>GABBR1</i> shows a complex methylation pattern in which human and gorilla shows similar pattern of methylation, orangutan shows relative hypomethylation, while chimpanzees and bonobos show increased methylation levels at TSS and intermediate levels associated with the short isoform of this gene. <i>GABBR1</i> is a neuronal receptor involved in synaptic inhibition, slow wave sleep, muscle relaxation and sensitivity to pain.</p

Example gene showing methylation differences between human and chimpanzee at promoter level.

<p><i>BRCA1</i> provides an example of co-occurrence between protein sequence evolution and gene regulation. The <i>BRCA1</i> gene shows changes in DNA methylation in a regulatory region upstream of the TSS <a href="http://www.plosgenetics.org/article/info:doi/10.1371/journal.pgen.1003763#pgen.1003763-Rice1" target="_blank">[40]</a> and a K_A/K_I ratio of 0.69 between human and chimpanzee.</p

Methylation patterns mimic sequence based phylogenetic relationships.

<p>(A) Methylation changes correlate with DNA sequence changes. x-axis shows the number of nucleotide substitutions between two species per kb, y-axis shows the changes in methylation based on β-values. (B) Neighbor-joining tree based on methylation data from probes with a perfect match in all reference genomes (31,853 autosomal CpGs). Bootstrap values (1,000 permutations) are shown for each node.</p

Candidate genes exclusively deregulated and targeted by human miR-541-3p.

<p>Candidate genes exclusively deregulated and targeted by human miR-541-3p.</p

miRNA conservation in clustered and non-clustered miRNAs.

<p>(<b>A</b>) Single nucleotide variant (SNV) density in clustered or non-clustered miRNAs, calculated as the average number of fixed substitutions in any of the great ape populations across the precursor miRNA. (<b>B</b>) Molecular age of clustered and non-clustered miRNAs. Molecular age is taken from Iwama et al. [<a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0154194#pone.0154194.ref017" target="_blank">17</a>] were each integer represents a period of origin with the oldest miRNAs having a value of -1 (right after the split between mammals and birds) and the youngest a value of 13 (after the split between humans and chimpanzees). (<b>C</b>) Correlation between SNV density and expression, calculated as the average expression values for miRNAs across five human tissues (cerebellum, brain, heart, kidney and testis) taken from Meunier et al. [<a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0154194#pone.0154194.ref016" target="_blank">16</a>].</p

Hairpin structures and stabilities of the studied miRNAs.

<p>Minimum free energy (MFE) and secondary structure predictions for human (hsa), chimpanzee (ptr) and macaque (mml) miRNA precursor sequences (miRBase, release 21), according to RNAfold. Grey circles represent nucleotide changes between miRNAs.</p