DNA Methylation: Enzymology

Abstract

Methylated bases (C5‐methylcytosine, N4‐methylcytosine and N6‐methyladenine) are found in the DNA of most species, where they play important biological roles. There exist two families of DNA methyltransferases with specificity for methylation of the C5 position of cytosine or exocyclic amino groups of cytosine or adenine. They all use S‐adenosyl‐l‐methionine as the methyl group donor and contain a catalytic domain with a similar fold. In bacteria, methylation of DNA is used to distinguish self and foreign DNA, to direct post‐replicative mismatch repair, co‐ordinate DNA replication and cell cycle and for regulation of gene expression. In vertebrates, DNA methylation is established by three DNA methyltransferases (Dnmt1, Dnmt3a and Dnmt3b) in a cell‐type‐specific pattern, mainly at the CG sites. It functions as an epigenetic mark and is involved in the regulation of chromatin accessibility in many biological contexts. Bacterial DNA methyltransferases are mainly directed by the DNA sequence of their target site, whereas mammalian and plant Dnmts are recruited and regulated by several factors and other epigenetic signals, including modification of histone tails or RNA molecules.

Key Concepts

  • DNA methylation adds information to the DNA that is not encoded in the DNA sequence.
  • DNA methyltransferases use AdoMet as methyl group donor and flip their target base out of the DNA helix.
  • DNA methylation in bacteria reflects the sequence specificity of the bacterial methyltransferases, whereas eukaryotic enzymes are targeted and regulated by a network of different mechanisms.
  • Maintenance of mammalian DNA methylation patterns is mainly encoded in the palindromic nature of the CG site and is coupled to DNA replication.
  • Eukaryotic DNA methyltransferases are recruited by specific histone modifications generating a network of interconnected epigenetic signals.

Keywords: DNA methylation; DNA methyltransferase; epigenetic modification; Dnmt1 ; Dnmt3 ; enzyme mechanism; enzyme regulation; 5‐methylcytosine; 6‐methyladenine

Figure 1. Types of methylated bases found in the DNA: C5‐methylcytsosine, N4‐methylcytosine and N6‐methyladenine. The methyl group is coloured in green. dR – deoxyribose.
Figure 2. Catalysis of the DNA methylation reaction. (a) he transfer of the methyl group from the cofactor S‐adenosyl‐l‐methionine (AdoMet) to the target base (adenine or cytosine) is catalyzed out by DNA methyltransferases. It leads to the generation of the methylated base and the cofactor product S‐adenosyl‐l‐homocysteine (AdoHcy). AdoMet and AdoHcy are shown in stick models, and the methyl group is coloured in green. (b) Structure of the bacterial M. HhaI cytosine‐C5 methyltransferase in complex with a specific DNA duplex (PDB 1MHT), illustrating the characteristic AdoMet‐dependent methyltransferase fold and base flipping. The enzyme is shown schematically coloured by secondary structure, and AdoHcy is shown in space fill representation coloured by atom type and the two strands of the DNA are labelled in green and orange. The target base is rotated out of the DNA helix and inserted into the catalytic pocket of the enzyme.
Figure 3. The methylation cycle in prokaryotes: dynamics and roles (for details refer to text).
Figure 4. The methylation cycle in higher eukaryotes: dynamics and roles (for details refer to text).
Figure 5. Structures of mammalian DNA MTases. (a) Domain architecture of mammalian DNA MTases. The functional domains in the N‐terminal parts of the proteins are indicated and the conserved C5 DNA MTase motifs in the C‐terminal part are labelled. (b) Crystal structure of the C‐terminal domain of Dnmt3L in complex with the Dnmt3a‐ADD in space‐filling representation (PDB 4U7T). A tetramer consisting of two molecules of Dnmt3a (in the centre in light and dark blue) and two molecules of Dnmt3L (at the edges in light and dark green) is shown. The ADD domains of Dnmt3a (in red) interact with the catalytic domain of the enzyme and regulate its activity. The H3 peptides bound to the ADD domains are shown in stick representation and AdoHcy is shown in space fill. (c) Crystal structure of Dnmt1 with the functional domains shown in different colours. The replication foci targeting domain (RFTS) and CXXC domain are located in the vicinity of the catalytic domain and regulate the activity and function of the enzyme (PDB 3AV4).
Figure 6. Examples of mechanisms contributing to the targeting of Dnmt3a in cells. (a) Model of the Dnmt3a/3L complex bound to the nucleosome. The ADD and PWWP domains of Dnmt3a directly read the modification stat of the histone H3 tails, recruit and activate the enzyme for methylation of DNA wrapped around histones carrying the specific marks. In this image, the ADD and PWWP domains were modelled. For simplicity, they were only included for one Dnmt3a catalytic domain (labelled with *). (b) Model of the polymerisation of the Dnmt3a/3L heterotetramers on DNA. Polymerisation of the enzyme stimulates its catalytic activity and contributes to the methylation pattern generated by Dnmt3a.
Figure 7. Modified cytosine bases found in genomic DNA of eukaryotes. 5‐hydroxymetylcytosine (5hmC), 5‐formylcytosine (5fC) and 5‐carboxylcytosine (5caC) are formed by TET‐mediated stepwise oxidation of the methyl groups of 5‐methylcytosine and are involved in the process of active DNA demethylation.
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Further Reading

Cheng X and Roberts RJ (2001b) AdoMet‐dependent methylation, DNA methyltransferases and base flipping. Nucleic Acids Research 29 (18): 3784–3795.

Jeltsch A and Jurkowska RZ (2014b) New concepts in DNA methylation. Trends in Biochemical Sciences 39 (7): 310–318.

Jeltsch A (2002b) Beyond Watson and Crick: DNA methylation and molecular enzymology of DNA methyltransferases. ChemBioChem 3: 274–293.

Jones PA (2012b) Functions of DNA methylation Islands, start sites, gene bodies and beyond. Nature Reviews Genetics 13 (7): 484–492.

Jurkowska RZ , Jurkowski TP and Jeltsch A (2011c) Structure and Function of Mammalian DNA Methyltransferases. ChemBioChem 12: 206–222.

Baylin SB and Jones PA (2011b) A decade of exploring the cancer epigenome – biological and translational implications. Nature Reviews Cancer 11 (10): 726–734.

Feng S , Jacobsen SE and Reik W (2010b) Epigenetic reprogramming in plant and animal development. Science 330 (6004): 622–627.

Wu H and Zhang Y (2014b) Reversing DNA methylation: mechanisms, genomics, and biological functions. Cell 156 (1–2): 45–68.

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Jurkowska, Renata Z, and Jeltsch, Albert(Apr 2015) DNA Methylation: Enzymology. In: eLS. John Wiley & Sons Ltd, Chichester. http://www.els.net [doi: 10.1002/9780470015902.a0006156.pub2]