Summary of DNA methylation profile of the human major histocompatibility complex: A pilot study for the human epigenome project.

1. Trevor Lythgoe
Biology Lab
Wed 1900
Summary of DNA Methylation Profiling of the Human Major Histocompatibility
Complex: A Pilot Study for the Human Epigenome Project
The following is a summary of the Pilot Study for the Human Epigenome as was
conducted by Vardhman K. Rakyan, Thomas Hildmann, Karen L. Novik, Jorn Lewin,
Jorg Tost, Antony V. Cox, T. Dan Andrews, Kevin L. Howe, Thomas Otto, Alexander
Olek, Judith Fischer, Ivo G. Gut, Kurt Berlin, and Stephan Beck.
Introduction
The Human Epigenome Project is intended to shed light on the complex process of gene
expression by mechanisms other than changes in the underlying DNA sequence. In short,
the Project hopes to understand what process allows for a single fertilized egg cell to
become neurons, muscle cells, blood cells etc. For instance, while the daughter cell
acquires new functions from the parent cell, it retains the DNA code necessary to
continue in its role of reproduction and growth. As a cell replicates it goes through a
process called chromatin remodeling, which is accomplished through two main
mechanisms, one of which addressed in this study is the addition of methyl groups to the
CpG sites, which convert cytosine to 5-methylcytosine. 5-methylscyotsine acts much the
same as cytosine in pairing with guanine, however, highly methylated areas tend to be
less transcriptionally active. The question of what importance this step is to the
development of the human cell expression was addressed in the study. The pilot study
was specifically aimed at the Human Major Histocompatibility Complex (MHC), a
cluster of genes located on chromosome 6 necessary to antigen production and critical to
the success of transplantation.
Citation: Rakyan VK, Hildmann T, Novik KL, Lewin J, Tost J, et al. (2004) DNA methylation profiling of the
human major histocompatibility complex: A pilot study for the human epigenome project. PLoS Biol 2(12):
e405.

2. Methods
Rakyan et al analyzed DNA methylation in seven human tissues (healthy and their
diseased variants) from 32 separate individuals– adipose, brain, breast, liver, lung, muscle
and prostate– using bisulphate sequencing and created a software program ESME to
determine the DNA methylation levels from the sequence trace files. This they compared
with a known system called matrix-assisted laser desorption/ionization mass
spectrometry (MALDI-MS) and found that their results had a concordance rate of 88%.
The concordance rate of 88% suggests the validity of the new method compared with the
older, but slower, known methods.
Results
A major goal of the pilot study was to create a database that would be publicly accessible
and adequately display the data learned through the study. The 253 unique amplicons
successfully analyzed were mapped to the human genome assembly using BLAST (Basic
Local Alignment Search Tool– an algorithm used for comparing primary biological
sequence information such as the amino-acid sequences of different proteins or the
nucleotides of DNA sequences.) (http://blast.ncbi.nlm.nih.gov/Blast.cgi, 18 March 2011)
The results of the study were made available on the internet at
http://www.epigenome.org.
Rakyan et al discovered that the methylation profile of the MHC region appears to be
strongly bimodal, 90% of the amplicons being hyper- (more than 70% median
methylation of amplicon) or hypomethylated (less than 30% of median methylation of
Citation: Rakyan VK, Hildmann T, Novik KL, Lewin J, Tost J, et al. (2004) DNA methylation profiling of the
human major histocompatibility complex: A pilot study for the human epigenome project. PLoS Biol 2(12):
e405.

3. amplicon.) They confirmed the idea that these extremes of methylation profiles are the
most stable states within the genome, finding also that the majority of the CpG islands
were hypomethylated throughout development. Hypermethalated regions result in
transcriptional silencing effectively turning off the gene through an environment of
heterochromatin.
While a majority of the genes that were associated with the tissue-specific methylation
profiles in this study showed no corresponding tissue-specific expression profiles in a
previous analysis, a significant portion (10%) of all amplicons displayed differential
methylation between the tissue types. Of these 31% were located in the upstream regions,
a proportion that is in the same range as the total number of upstream amplicons relative
to intragenic amplicons analyzed in this study. One reason for the lack of significant data
supporting the identification of tissue specificity of methylation profiles is that the
detection of some genes that are known to be associated with various mRNA isoforms is
quite difficult with conventional microarray analysis. Another theory is that there are
only a small proportion of genes affected by methylation result in tissue specificity.
Increasing evidence suggests that an individual’s epigenetic profile can influence
phenotype and susceptibility to various diseases such as cancer. This is supported by the
amount of intragenic amplicons found despite the fact that nearly all loci studied
evidenced some degree of heterogeneity.
The primary purpose of epigenetic modifications is to control gene expression. An
Citation: Rakyan VK, Hildmann T, Novik KL, Lewin J, Tost J, et al. (2004) DNA methylation profiling of the
human major histocompatibility complex: A pilot study for the human epigenome project. PLoS Biol 2(12):
e405.

4. abnormal epigenetic modification, as in many cancers, occurs within in the CpG islands
that overlap promoters, which may result in aberrant transcription of the cognate gene, a
tumor suppressor. If it were possible to control epigenetic modification the possibility of
eliminating genetic mutations like cancer, or even in the more moderate form of
controlling genetic disease in plants, would be an incredible boon to society as a whole.
Conclusion
The biggest challenge of the Human Epigenome Project is acquiring all the elements
from several large-scale studies and obtaining a single holistic view of the human
genome. The HEP database will provide a central location to collect the results from
various studies. The results from the pilot study show a significant proportion of the
analyzed loci within the MHC show tissue-specific methylation profiles, which may help
us to understand certain cell phenotypes. Understanding certain cell phenotypes may lead
to the ability to control epigenetic modification and the cure for cancer.
Citation: Rakyan VK, Hildmann T, Novik KL, Lewin J, Tost J, et al. (2004) DNA methylation profiling of the
human major histocompatibility complex: A pilot study for the human epigenome project. PLoS Biol 2(12):
e405.