ReviewSuicidal function of DNA methylation in age-related genome disintegration
Introduction
Although 5-methylcytosine (5mC, mC) was discovered in DNA sixty years ago (Hotchkis, 1948, Wyatt, 1951), the biological role of this enzymatic cytosine modification in eukaryotic organisms continues to be one of the most intriguing mysteries in molecular genetics. DNA methylation affects many genetic and epigenetic processes, including regulation of gene activity and genome imprinting, by inducing chromatin remodeling (Bird, 2002, Laird, 2005). At the same time, the methylation of cytosine residues produces mutation hotspots that can lead to human genetic diseases and cancer (Cooper and Youssoufian, 1988, Rideout et al., 1990). A paradox emerges from the recognition that the genetic regulation usually requires stabile and reliable mechanisms, but methylation destabilizes structure and inactivates genes throughout the organism.
A number of other puzzling facts about DNA methylation have accrued since its discovery (Jones, 1999). For example, some eukaryotes lack DNA methyltransferase genes (MTase) (Ponger, 2005), and the 5mC content in the genome varies by more than 100-fold across species (Lyko et al., 2000, Vanyushin, 2006). Most of the genes that are methylated in vertebrates are unmethylated in lower eukaryotic species (Bird, 2002, Selker et al., 2003). The bulk of the available 5′-CpG-3′ (CG) sites are methylated in embryos (Monk et al., 1987), whereas most 5mC are lost from the genome of various species over their ontogenesis (Mazin, 1993a, Lyko et al., 2000). DNA methylation patterns are extraordinarily labile and substantial inter-individual variation has been demonstrated, even for identical twins (Fraga et al., 2005, Heijmans et al., 2007). Inactivation of the dnmt1 MTase gene causes DNA demethylation and death of homozygous mouse embryos in early development (Li et al., 1992, Takebayashi et al., 2007). However, heterozygous DNMT1-knockdown mice show increased 5mC content with aging (Yung et al., 2001), and embryonic stem (ES) cells grow normally in the absence of MTases (Li et al., 1992, Tsumura et al., 2006). About 80% of ancestral CG sites have disappeared from the human genome during evolution (Josse et al., 1961) and many spontaneous mutations result in new CG sites (Mazin, 1994a, Yang et al., 1996). The detection of asymmetrically methylated mCpNpN sites (Meyer et al., 1994, Ramsahoye et al., 2000), massive methylation of post-replicative DNA (Vanyushin, 1984) and aberrant methylation of genes (Esteller, 2005, Kobayashi et al., 2007) all contradict the mechanism of semi-conservative inheritance of methylation patterns (Holliday and Pugh, 1975, Riggs, 1975). The age-related loss of most 5mC from the genome of cultured normal cells coincides with their Hayflick limit (Mazin, 1993c, Mazin, 1995), but DNA hypermethylation is observed in cancer cells growing in vitro (Wilson and Jones, 1983, Gray et al., 1991). Conversely, a global reduction in methylation is typical for the genome of cancer cells in vivo (Gama-Sosa et al., 1983, Cadieux et al., 2006). Resolution of these paradoxes is a prerequisite for understanding the biological role of DNA methylation, but no existing hypothesis can explain all of these contradictory observations. The reasons may be hidden in the poly-functionality of methylation.
In this review, the methodology of the systems analysis was used to compose a multicolored portrait of the mutagenic function of DNA methylation from the fractured, yet key, experimental facts accumulated over the years. Each section of the article reflects an integral part of the whole picture and discusses how this mechanism works on molecular, cellular, organismal, and evolutionary levels and over cell cycle, ontogenesis, and specific. The synthesis of these component parts into coherent model allowed the proposal of the hypothesis, which considers a role of DNA methylation in the age-related genome self-destruction as the universal genetic mechanism for programming cell's and organism's aging and death. The epigenetic function of methylation has been discussed in many recent reviews (Doerfler, 2008, Feinberg, 2008, Ooi and Bestor, 2008, Calvanese et al., 2009, Sinclair and Oberdoerffer, 2009).
Section snippets
DNA methylation induces most of spontaneous mutations of multiple classes
The mutagenic activity of cytosine methylation was studied in detail, and it is commonly accepted that CG methylation sites are hotspots for most C > T and G > A (G:C > A:T) transition mutations in human genes (Cooper and Krawczak, 1989). CG-containing restriction sites are often polymorphic and have a high frequency of 5mC > T mutations in both Esherichia coli and human DNA (Barker et al., 1984, Coulondre et al., 1985). Most single-base substitutions (BS) that are responsible for genetic diseases and
Cytosine methylation as a mutagen
Why does DNA methylation produce so many mutations? Pathways for the several of several different classes of mutations at CG sites are presented in Fig. 1. Half-methylated CG/mCG (hmCG) sites arise in newly synthesized DNA during replication (I). These sites are recognized by the “maintenance” MTase, DNMT1, which is integrated into the DNA replication enzymatic complex and restores symmetrically methylated CG sites (1). A fraction of the newly formed 5mC residues are usually deaminated during
The origin of polymorphism of methylation sites and asymmetric mCpNpN sites
One of the conclusions that follows from the model presented on the Fig. 1 is that a final product of cytosine methylation may be asymmetrically methylated TG/mCA sites (1 > 2 > 6 > 7). The unusually methylated mCpNpN (mCNN, N is A, T, C, or mC) sites have been found in the DNA of fungi, insects, plants, and animals over the last decade (Selker and Stevens, 1985, Meyer et al., 1994, Ramsahoye et al., 2000, Wada et al., 2003). It was shown that these non-palindromic sites can result from the special
Relict 5mC > T mutations at CpG sites in DNA
5mC > T transitions may occur not only in somatic cells, but also in germline cells of various species. Since these mutations are generally irreversible, owing to the lower chance of inverse substitutions (T > C > 5mC) and the fact that most are neutral, these mutations have accumulated in the genome of vertebrate germlines over millions years of evolution (Cooper and Krawczak, 1989). Thus, the observed frequency of CG sites in human DNA is only eight dinucleotides per 1000n (kbp) of DNA instead of
The genome loses most 5mC over ontogenesis in various species and the hayflick limit of normal cells
One of the most intriguing results, which follows from the model of accumulation of methylation-dependent mutations, is the possible mechanism of reduction in 5mC content in genome with age (Fig. 1). Age-related DNA demethylation was demonstrated for differentiating mammalian cells (Jost et al., 2001, Kress et al., 2001, Kress et al., 2006), normal cells cultured in vitro (Wilson and Jones, 1983, Fairweather et al., 1985, Gray et al., 1991), growing higher plants (Kirnos et al., 1983),
What are the functions of DNA methylation?
The data considered above allow us to assume that cytosine methylation is the special mutagenic mechanism, which may be implicated in various biological processes. Eukaryotic MTases may have evolved from prokaryotic methylation-restriction enzymes or tRNA methyltransferases (Bestor, 1990, Jurkowski et al., 2008) and have served diverse functions throughout genome evolution.
Evolution would be impossible without the spontaneous mutations that are a source of variability on which natural selection
DNA methylation as a mechanism of aging
Let us consider in more detail a role of DNA methylation in the age-related genome self-destruction, which may be one of the mechanisms of aging in higher eukaryotic organisms. Fifteen years ago, the hypothesis was put forward that DNA methylation is the source of the genome instability that is implicated in the aging process (Mazin, 1994b). As expected, cytosine methylation works as a special counter of cell divisions. About 50 millions new 5mC residues arise in the haploid mammalian genome (3 ×
Paradoxes of DNA methylation
The poly-functionality of DNA methylation and its crosstalk with various genetic processes might explain many of the paradoxical facts referred to in the Introduction. For example, there is a “5mC value paradox”, as the 5mC content in DNA can vary from undetectable levels to more than10 mol% across eukaryotic species. Free-living soil worms (Caenorhabditis elegans) and baker's yeast (S. cerevisiae) lack MTase genes and their genomes do not appear contain 5mC (Ponger, 2005). DNA methylation was
Conclusion
The focus of this review has been that DNA methylation is a potent endogenous generator of mutations that may induce high level of DNA damage and genome disintegration responsible for cell apoptosis and organism aging and death. The hypothesis about suicidal function of methylation is presented here as an integral model, which has been examined in detail and from a number of different perspectives. This function may be involved in various genetic processes. As a generator of mutations and a
Acknowledgements
I am grateful to P.J. Allenberg, N.E. Braudo, C.D. Jackson, and T.I. Vlasova for comments, T.I. Heifetz for proofreading the manuscript, L.A. Mazin for design of figures and web site www.DNAgeing.net and my colleagues in Moscow State University for their support.
References (188)
- et al.
Overproduction of DNA cytosine methyltransferases causes methylation and C > T mutations at non-canonical sites
J. Biol. Chem.
(1996) - et al.
Restriction sites containing CpG show a higher frequency of polymorphism in human DNA
Cell
(1984) - et al.
Characterization of Dnmt3b:thymine-DNA glycosylase interaction and stimulation of thymine glycosylase-mediated repair by DNA methyltransferase(s) and RNA
J. Mol. Biol.
(2008) - et al.
Hemimethylation and non-CpG methylation levels in a promoter region of human LINE-1 (L1) repeated elements
J. Biol. Chem.
(2005) - et al.
The role of epigenetics in aging and age-related diseases
Ageing Res. Rev.
(2009) - et al.
LC/ESI-MS demonstrates the absence of 5-methyl-2′-desoxycytosine in Plasmodium falciparum genomic DNA
Mol. Biochem. Parasitol.
(2006) - et al.
DNA methylation and CpG suppression
Cell Differ.
(1985) - et al.
Inactivation of Dnmt3b in mouse embryonic fibroblasts results in DNA hypomethylation, chromosomal instability, and spontaneous immortalization
J. Biol. Chem.
(2005) - et al.
The in vitro lifespan of MRC-5 cells is shortened by 5-azacytidine-induced deamination
Exper. Cell Res.
(1987) - et al.
Dynamic CpG and non-CpG methylation of the Peg1/Mest gene in the mouse oocyte and preimplantated embryo
J. Biol. Chem.
(2005)
Evolutionary changes in CpG and methylation levels in the genome of vertebrates
Gene
The DNA methylation paradox
Trends Genet.
Enzymatic synthesis of deoxyribonucleic acid. VIII. Frequency of nearest neighbour base sequences in deoxyribonucleic acid
J. Biol. Chem.
Genomic and p16-specific DNA methylation of the mouse colon: elder age and dietary folate as interactive determinants
J. Nutrish.
Apurinic sites are position-specific topoisomerase II poisons
J. Biol. Chem.
Local DNA demethylation in vertebrates: how could it be performed and targeted?
FEBS Lett.
Targeted mutation of the DNA methyltransferase gene results in embryonic lethality
Cell
Role of the Arabidopsis DNA glycosylase/lyase ROS1 in active DNA demethylation
Proc. Natl. Acad. Sci. U.S.A.
Restriction landmark genome scanning identifies culture-induced DNA methylation instability in the human embryonic stem cell epigenome
Hum. Mol. Genet.
Concurrent replication and methylation at mammalian origins of replication
Mol. Cell Biol.
DNA methylation and restriction
Annu. Rev. Biochem.
HpaII methyltransferase is mutagenic in Escherichia coli
J. Bact.
Age and tissues specificity of 5-methylcytosine content in DNA of spawning salmon
Biokchem
DNA methylation: evolution of a bacterial immune function into a regulator of gene expression and genome structure in higher eukaryotes
Phylos. Trans. R. Soc. Lond. B. Biol. Sci.
Differentiation of two mouse cell lines is associated with hypomethylation of their genomes
Mol. Cell. Biol.
Dnmt1 overexpression causes genomic hypermethylation, loss of imprinting, and embryonic lethality
Mol. Cell. Biol.
DNA methylation patterns and epigenetic memory
Genes Dev.
Intra-individual change over time in DNA methylation with family clustering
J. Am. Med. Assoc.
Folate deficiency causes uracil misincorporation into human DNA and chromosome breakage: implications for cancer and neuronal damage
Proc. Natl. Acad Sci. U.S.A.
Inter-individual variation of DNA methylation and its implications for large-scale epigenome mapping
Nucl. Acid.
Why does the human factor IX gene have a G + C content of 40%?
Am. J. Hum. Genet.
Genome-epigenome interactions in cancer
Hum. Mol. Genet.
Genome-wide hypomethylation in human glioblastomas associated with specific copy number alteration, methylenetetrahydrofolate reductase allele status, and increased proliferation
Cancer Res.
Sequence-dependent enhancement of hydrolytic deamination of cytosines in DNA by the restriction enzyme PspGI
Nucleic. Acids Res.
Genome-wide epigenetic alterations in cloned bovine foetuses
Biol. Reprod.
Reduced rates of gene loss, gene silencing, and gene mutation in Dnmt1-deficient ()embryonic stem cells
Mol. Cell. Biol.
High frequency mutagenesis by a DNA methyltransferase
Cell
The rate of hydrolytic deamination of 5-methylcytosine in double-stranded DNA
Nucleic Acids Res.
A mutant HpaII methyltransferase functions as a mutator enzyme
Nucleic Acids Res.
DNA hypomethylation leads to elevated mutation rates
Nature
DNA damage, cellular senescence and organismal ageing:causal or correlative?
Nucl. Acids Res.
Molecular basis of base substitution hotspots in Escherichia coli
Nature
The CpG dinucleotides and human genetic diseases
Hum. Genet.
Cytosine methylation and the fate of CpG dinucleotides in vertebrate genomes
Hum. Genet.
Influence of sex, smoking and age on human hprt mutation frequencies and spectra
Genetics
The age of cancer
Nature
Causes of death before birth
Nature
Evolution of gene sequence in response to chromosomal location
Genetics
In pursuit of the first recognized epigenetic signal-DNA methylation: a 1976 to 2008 synopsis
Epigenetics
Genome dynamics in aging mice
Genome Res.
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