Methylation is a term used in the chemical sciences to denote the attachment or substitution of a methyl group on various substrates. This term is commonly used in chemistry, biochemistry, and the biological sciences.
In biochemistry, methylation more specifically refers to the replacement of a hydrogen atom with the methyl group.
In biological systems, methylation is catalyzed by enzymes; such methylation can be involved in modification of heavy metals, regulation of gene expression, regulation of protein function, and RNA metabolism. Methylation of heavy metals can also occur outside of biological systems. Chemical methylation of tissue samples is also one method for reducing certain histological staining artifacts.
Methylation contributing to epigenetic inheritance can occur either through DNA methylation or protein methylation.
DNA methylation in vertebrates typically occurs at CpG sites (cytosine-phosphate-guanine sites; that is, where a cytosine is directly followed by a guanine in the DNA sequence); this methylation results in the conversion of the cytosine to 5-methylcytosine. The formation of Me-CpG is catalyzed by the enzyme DNA methyltransferase. CpG sites are uncommon in vertebrate genomes but are often found at higher density near vertebrate gene promoters where they are collectively referred to as CpG islands. The methylation state of these CpG sites can have a major impact on gene activity/expression.
Protein methylation typically takes place on arginine or lysine amino acid residues in the protein sequence. Arginine can be methylated once (monomethylated arginine) or twice, with either both methyl groups on one terminal nitrogen (asymmetric dimethylated arginine) or one on both nitrogens (symmetric dimethylated arginine) by peptidylarginine methyltransferases (PRMTs). Lysine can be methylated once, twice or three times by lysine methyltransferases. Protein methylation has been most well studied in the histones. The transfer of methyl groups from S-adenosyl methionine to histones is catalyzed by enzymes known as histone methyltransferases. Histones which are methylated on certain residues can act epigenetically to repress or activate "gene" expression. Protein methylation is one type of post-translational modification.
In early development (fertilization to 8-cell stage), the eukaryotic genome is demethylated. From the 8-cell stage to the morula, de novo methylation of the genome occurs, modifying and adding epigenetic information to the genome. By blastula stage, the methylation is complete. This process is referred to as "epigenetic reprogramming". The importance of methylation was shown in knockout mutants without DNA methyltransferase. All the resulting embryos died at the morula stage.
Methylation in postnatal development
Increasing evidence is revealing a role of methylation in the interaction of environmental factors with genetic expression. Differences in maternal care during the first 6 days of life in the rat induce differential methylation patterns in some promoter regions and thus influencing gene expression (Weaver IC, et al (Aug 2004; epub Jun 27 2004). "Epigenetic programming by maternal behavior.". Nature Neuroscience 7(8): 791-92.). Furthermore, even more dynamic processes such as interleukin signaling have been shown to be regulated by methylation (Bird A. (Mar 2003). "Il2 transcription unleashed by active DNA demethylation.". Nature Immunology 4(3): 208-9.).
Methylation and cancer
The pattern of methylation has recently become an important topic for research. Studies have found that in normal tissue, methylation of a gene is mainly localised to the coding region, which is CpG poor. In contrast, the promoter region of the gene is unmethylated, despite a high density of CpG islands in the region.
Neoplasia is characterized by "methylation imbalance" where genome-wide hypomethylation is accompanied by localized hypermethylation and an increase in expression of DNA methyltransferase (1). The overall methylation state in a cell might also be a precipitating factor in carcinogenesis as evidence suggests that genome-wide hypomethylation can lead to chromosome instability and increased mutation rates (3). The methylation state of some genes can be used as a biomarker for tumorigenesis. For instance, hypermethylation of the pi-class glutathione S-transferase gene (GSTP1) appears to be a promising diagnostic indicator of prostate cancer (2).
In cancer, the dynamics of genetic and epigenetic gene silencing are very different. Somatic genetic mutation leads to a block in the production of functional protein from the mutant allele. If a selective advantage is conferred to the cell, the cells expand clonally to give rise to a tumor in which all cells lack the capacity to produce protein. In contrast, epigenetically mediated gene silencing occurs gradually. It begins with a subtle decrease in transcription, fostering a decrease in protection of the CpG island from the spread of flanking heterochromatin and methylation into the island. This loss results in gradual increases of individual CpG sites, which vary between copies of the same gene in different cells (6).
Methylation and bacterial host defense
Additionally, adenosine or cytosine methylation is part of the restriction modification system of many bacteria. Bacterial DNAs are methylated periodically throughout the genome. A methylase is the enzyme that recognizes a specific sequence and methylates one of the bases in or near that sequence. Foreign DNAs (which are not methylated in this manner) that are introduced into the cell are degraded by sequence-specific restriction enzymes. Bacterial genomic DNA is not recognized by these restriction enzymes. The methylation of native DNA acts as a sort of primitive immune system, allowing the bacteria to protect themselves from infection by bacteriophage or phage. These restriction enzymes are the basis of restriction fragment length polymorphism (RFLP) testing. With this technique, geneticists use various bacterial restriction endonucleases (restriction enzymes) to split DNA at specific sites in order to detect DNA polymorphisms, useful for genetic fingerprinting and genetic engineering.
Methylation in chemistry
The term methylation in organic chemistry refers to the alkylation process used to describe the delivery of a CH3 group. This is commonly performed using electrophilic methyl sources - iodomethane, dimethyl sulfate, dimethyl carbonate, or less commonly with the more powerful (and more dangerous) methylating reagents of methyl triflate or methyl fluorosulfonate (magic methyl), which all react via SN2 nucleophilic substitution. For example a carboxylate may be methylated on oxygen to give a methyl ester, an alkoxide salt RO− may be likewise methylated to give an ether, ROCH3, or a ketone enolate may be methylated on carbon to produce a new ketone.
Alternatively, the methylation may involve use of nucleophilic methyl compounds such as methyllithium (CH3Li) or Grignard reagents (CH3MgX). For example, CH3Li will methylate acetone, adding across the carbonyl (C=O) to give the lithium alkoxide of tert-butanol:
- M. Nakayama, M. L. Gonzalgo, S. Yegnasubramanian, X. Lin, A. M. D. Marzo and W. G. Nelson (2004). "GSTP1 CpG island hypermethylation as a molecular biomarker for prostate cancer". Journal of Cellular Biochemistry 91 (3): 540-552. doi:10.1002/jcb.10740.
- Grewal, S.I.; Rice, J.C. (2004). "Regulation of heterochromatin by histone methylation and small RNAs". Current Opinion in Cell Biology 16 (3): 230-238.
- Jones, P.A.; Baylin, S.B. (2002). "The fundamental role of epigenetic events in cancer". Nature Reviews Genetics 3 (6): 415-428. doi:10.1038/nrg816.
- Nakayama, J.; Rice, J. C.; Strahl, B. D.; Allis, C.D.; Grewal, S.I.; (2001). "Role of histone H3 lysine 9 methylation in epigenetic control of heterochromatin assembly". Science 292 (5514): 110-113. doi:10.1126/science.1060118.
- March, J.; Advanced Organic Chemistry, 5th ed., Wiley, New York, 2001.
- Walsh, C.. "Chapter 5 - Protein Methylation", Posttranslational Modifications of Proteins. ISBN 0-9747077-3-2.
- Baylin, S.B.; Herman, J.G.; Graff, J.R.; Vertino, P.M.; Issa, J.P. (1998). "Alterations in DNA methylation: a fundamental aspect of neoplasia.". Advances in Cancer Research 72: 141-96. PMID 9338076.
- Chen, R.Z.; Pettersson, U.; Beard, C.; Jackson-grusby, L.; Jaenisch, R. (1998). "DNA hypomethylation leads to elevated mutation rates.". Nature 395 (6697): 89-93. doi:10.1038/25779.
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