Pop-CpGs for GWAS, DNA Methylation Contributing to Human Diversity

Phenotypes are traits expressed by organisms in a specific environment. When you think of your own ethnicity, you might mostly be considering your appearance. These visible characteristics are associated with where your ancestors came from, of course. Now, don’t take this personally. But, biomedical researchers associate your ethnicity with risks. Why?! Because ethnicity can be associated with phenotypes with health risks.

We once expected that by sequencing the entire human genome, the basis of human diversity would be revealed. After all, your traits are based on your genes, right? Unfortunately, it has been difficult to demonstrate how our genomes produce the varieties of human phenotypes. Case in point, most human mutations / gene variants are located in non-coding regions of our DNA. Research focus has shifted to look behind the “genome sequence” curtain, so to speak. Behind the curtain, there is a complicated system of inter-connected epigenetic machinery, modulating gene expression. Human genome databases are the basis for this exploration.

How does epigenetics relate with phenotypes? For the paper, Holger Heyn et al. DNA Methylation contributes to natural human variation (2013) Genome Research, the authors interrogated the methylomes of three ethnic groups using methylation sequencing arrays. They report several fascinating findings that struck me.

  • Their method of GWAS targeting methylation clusters blood & tissue samples by ethnicity.
  • Methylation appears to act as an evolutionary established mediator, within the relationship between genotype and epitype.
  • Local selective pressure may induce independent epigenetic variants.
  • “Pop-CpGs”* could aid genetic variance GWAS. (*definition below)


In this study, three ethnic groups were sorted by methyomes alone. The researchers started with 288 samples B-lymphoblastoid cell lines, produced by EBV immortalization. B-lymphoblastoid cell line genomic DNA samples were sourced evenly from Caucasian-American, African-American and Han Chinese American people. (BTW Fun fact: Han Chinese are the largest ethnic group in the world! )Methylation profiles were assessed using Illumina’s DNA methylation BeadChip platform. This array covers 96% of CpG islands, totaling 485,000 genes, at single nucleotide resolution. Libraries of genomic DNA are bisulfite treated. This process converts un-methylated cytosines to uracil. Samples are each exposed to methylated bead probes, and un-methylated bead probes. A matched DNA probe causes single-base extension of that sample sequence, incorporating a labeled ddNTP, that is then detected through fluorescence staining. By this sequencing array method, the group found 1373 differentially methylated CpG sites that clustered primarily by donor ancestral geographical origin. Termed by the authros as , “Pop-CpGs”, these are cytosine phosphodiester bond guanine sites that have population specific differential methylation. Of the 1373 sites, 439 pop-CpG sites for ethnicity were also shared in primary blood samples. They also present results with regard to human tissues, Hepatitis B infection risk and xenobiotic response factors.

The researchers also discovered evidence for other types of gene expression regulation. Their analysis of pop-CpGs showed an enrichment with two histone enhancer marks and a histone heterochromatin mark. … transcription factor binding sites. They performed expression studies to show the DNA methylation at some of these pop-CpGs was associated with gene repression and activation. Some pop-CpGs are located in those non-coding areas. Therefore it appears that regulatory elements other than promoters, are tapped for human variation.

Their results are a small survey of how the human epigenetic system works with our genetic system. A third of pop-CpG were not directly associated with any underlying genetic variation. Two thirds were. So, most of the time the DNA sequence sets up the context for methylation. Ethnicity in these study groups, is linked to differences in drug metabolism, response to external stimuli, sensory perception and disease risk. The authors’ suggestion that independent pop-CpGs could be the result of things like diet, pathogen, drugs, pesticides, carcinogens and hormone exposures makes sense. More GWAS studies using Pop-CpGs should lend support the epitype induction idea.

Heyn H, Moran S, Hernando-Herraez I, Sayols S, Gomez A, Sandoval J, Monk D, Hata K, Marques-Bonet T, Wang L, & Esteller M (2013). DNA methylation contributes to natural human variation. Genome research, 23 (9), 1363-72 PMID: 23908385

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