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Review
. 2013 Oct;15(4):373-81.
doi: 10.1177/1099800412444785. Epub 2012 Jun 3.

Epigenetic regulation and measurement of epigenetic changes

Kimberly E Stephens  1 , Christine A Miaskowski, Jon D Levine, Clive R Pullinger, Bradley E Aouizerat
Affiliations
Review

Epigenetic regulation and measurement of epigenetic changes

Kimberly E Stephens et al. Biol Res Nurs. 2013 Oct.

Abstract

Epigenetic mechanisms provide an adaptive layer of control in the regulation of gene expression that enables an organism to adjust to a changing environment. Epigenetic regulation increases the functional complexity of deoxyribonucleic acid (DNA) by altering chromatin structure, nuclear organization, and transcript stability. These changes may additively or synergistically influence gene expression and result in long-term molecular and functional consequences independent of the DNA sequence that may ultimately define an individual's phenotype. This article (1) describes histone modification, DNA methylation, and expression of small noncoding RNA species; (2) reviews the most common methods used to measure these epigenetic changes; and (3) presents factors that need to be considered when choosing a specific tissue to evaluate for epigenetic changes.

Keywords: DNA methylation; epigenetics; histone modification; small untranslated RNA.

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Figures

Figure 1
Figure 1
Chromatin organization. Chromatin is arranged into nucleosomes as its first level of organization. A nucleosome is made up of histone proteins and double-stranded deoxyribonucleic acid (DNA) that wraps around a histone octomer 1.7 times in a left-handed coil. Hydrogen bonds hold the phosphodiester backbone of DNA to the amino acid side chains of the histones. Each nucleosome is separated from the next by a short segment of DNA. The conformation resembles a “beads-on-a-string” structure. Further condensation of chromatin structure is facilitated by histone “tails” that extend from the nucleosome and help to stack nucleosomes into a more compact chromatin structure by linking them to neighboring nucleosomes. These circular stacks of nucleosomes compact further to form a solenoid that results in a chromatin fiber that measures 30 nanometers in diameter. As the final chromatin structure, chromosomes are thought to be the result of a mesh formed through fibers connected by cross-linked proteins or as a result of hierarchical packaging of chromatin around a central axis mediated by structural maintenance of chromosome proteins. Reprinted by permission from Macmillan Publishers Ltd: Felsenfeld & Groudine, 2003.
Figure 2
Figure 2
Posttranslational modifications of the N-terminal region of histone H4. The majority of the known histone modifications affect the amino acids that make up the unstructured N-terminal region of the histone core protein. Covalent modifications of the human H4 histone protein include phosphorylation (P), acetylation (Ac), and methylation (Me). Single letter abbreviations for amino acid residues: A = alanine; G = glycine; H = histidine; K = lysine; L = leucine; R = arginine; S = serine.
Figure 3
Figure 3
Following amplification of the region of interest by polymerase chain reaction (PCR), array-based methods rely on hybridization of a sample of deoxyribonucleic acid (DNA) to oligonucleotides contained within the array. Sodium bisulfite treatment of DNA produces a change in the nucleotide sequence of all unmethylated cytosines. Amplification of a specific locus of interest is performed, and a “tag” is incorporated into the PCR products using modified PCR primers that feature one of two unique sequence tails (i.e., one primer detects the methylated state and has one specific tag, while the other primer detects the unmethylated state and has a different tag). This procedure results in fluorescently labeled PCR products with a nucleotide sequence specific to its methylation status. A set of two oligonucleotides is synthetically made with their sequences complementary to the unmethylated and methylated sequence of the labeled PCR products and is attached to the surface of a glass slide. These immobilized oligonucleotides serve as a target for the labeled PCR products and are referred to as probes. The labeled PCR products are incubated with an array of immobilized probes to permit complementary sequences to hybridize to a given probe. A high-resolution camera captures the position of emitted fluorescence. The difference in signal intensities between the paired methylated and unmethylated alleles is used to calculate the percentage of methylation for the sequence associated with the probe. Modified from “Methylation-specific oligonucleotide microarray: A new potential for high-throughput methylation analysis,” by R. S. Gitan, H. Shi, C.-M. Chen, P. S. Yan, & T. H.-M. Huang, Genome Research, 12, p. 159. Copyright 2002 by Cold Springs Harbor Laboratory Press.
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