Applying whole-genome studies of epigenetic regulation to study human disease

Epigenetics may be broadly defined as the study of processes that produce a heritable phenotype that is not strictly dependent on DNA sequence. The definition has traditionally been restricted to processes that occur in the cell’s nucleus, with the term ‘heritable’ having a loose meaning that can be applied to either the entire organism or single cells. For example, a process that produces a phenotype only in a specific cell type (for instance, chromatin-mediated maintenance of a differentiated state) is usually considered epigenetic even if it is not directly inherited, but instead must be re-established or actively maintained at each cell division. Given this definition, the field of epigenetics has long focused on proteins that affect DNA packaging, and thereby affect the utilization of the genetic information encoded in the DNA template. This focus extends to the enzymatic modification of those proteins, and to the enzymatic modification of the DNA template itself, primarily DNA methylation. This review is written in conjunction with the international symposium on Genome-wide Epigenetics 2005, held at the University of Tokyo, Japan on November 8–10, 2005. Over the past decade, the field of epigenetics has undergone an exciting expansion in the number of researchers, techniques available and our understanding of epigenetic phenomena. The purpose of this short review is not to summarize all of these advances, but rather to guide the reader to more detailed sources of information by sketching an outline of the major thrusts in the field, emphasizing mammalian epigenetics in particular.Epigenetic information manifests itself by regulating gene expression, often reflected by biological phenotypes. The mechanistic mediators of heritable gene expression appear to be similar whether one is considering epigenetic inheritance on the organismal level (from parent to offspring) or the cellular level (from parent to daughter cells). For example, a mediator of epigenetic regulation is cytosine methylation. The action of maintenance methyltransferases ensures that the methylation of pairs of cytosines on complementary DNA strands at CpG dinucleotides is propagated to both pairs of daughter DNA molecules following DNA replication (Bestor, 2000). The inheritance of gene regulatory information from parental gametes defines the phenomenon of genomic imprinting, for which cytosine methylation is a central mediator (Li et al., 1993). Cytosine methylation is the best-studied mediator of epigenetic regulation. In mammalian genomes, the vast majority of methylation of cytosines occurs when the cytosine is followed by a guanine, a so-called CpG dinucleotide (Clark et al., 1995). The methyl group protrudes into the major groove of the DNA double helix, preventing the binding of transcription factors that would otherwise bind locally in a sequence-specific manner (Bell and Felsenfeld, 2000; Hark et al., 2000), and facilitating the binding of methyl-binding proteins that generally attach to DNA in a sequence non-specific manner (Jorgensen and Bird, 2002). The methylation of cytosines may not be the initiating event in a change from activity to silencing of a locus, but it is certainly associated with a number of molecular changes. These include the alteration of covalent modifications of histone tails (El-Osta and Wolffe, 2000), the recruitment of methyl-binding proteins (Jorgensen and Bird, 2002) and proteins normally associated with distinctive chromatin states (such as SWI/SNF components; Harikrishnan et al., 2005), and the compaction of the chromatin with silencing of gene expression locally (Nguyen et al., 2001). It is possible that cytosine methylation is induced by primary events involving the covalent modification of histone tails. The histone octamers that form the nucleosome protrude N-terminal tails, where it was thought that various combinations of methylation, acetylation, phosphorylation and other post-translational modifications could be established to create what has been referred to as the histone code (Jenuwein and Allis, 2001). However, this combinatorial concept is being challenged by data suggesting that the pattern of modifications may be more simple than the potential complexity, establishing what is more like a binary on or off pattern (Schubeler et al., 2004; Dion et al., 2005; Liu CL et al., 2005). As described for methylcytosine, these modified histone molecules are capable of recruiting further molecules and complexes that influence chromatin structure and local gene activity. Whether histone tail modifications induce or are induced by cytosine methylation, it seems that the disruption of one component of this system can induce changes in other components also (Ben-Porath and Cedar, 2001). The recent finding in fission yeast that small RNAs are targeted to heterochromatin (Verdel et al., 2004) suggests that epigenetic gene regulation in mammalian cells may also involve the actions of such short RNAs. In addition, it has been observed that a subset of DNA methyltransferases and methyl-binding proteins have RNA-binding properties (Jeffery and Nakielny, 2004), supporting the idea that RNA molecules may influence the regulation of expression of their own or other loci. However, a point worth stressing is that the major consequence of all these biochemical processes, which ultimately alter gene expression, is the regulation of transcription factor binding. Transcription factors with activating effects bind in modules at specific loci to facilitate the loading of the RNA polymerase complex locally, a prerequisite for gene expression. The relationship between epigenetic regulation and transcription factor binding tends to be overlooked in reviews of either field. However, as we emphasize below, techniques to study chromatin components and transcription factors in vivo are equally advanced, allowing integration of data and major advances in the understanding of the relationship of epigenetic alterations with transcription factor binding. An example of such a study shows a relationship between binding of the CTCF nuclear protein and cytosine methylation in a large-scale analysis (Mukhopadhyay et al., 2004). 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