Methylated cytosine

Small quantities of additional purines and pyrimidines occur in DNA and RNAs. Examples include 5-methyl-cytosine of bacterial and human DNA, 5-hydroxy-methylcytosine of bacterial and viral nucleic acids, and mono- and di-N-methylated adenine and guanine of... 

SCHEME 2.4 Competing reaction pathways in the benzylation of cytosine (R = H) and 1-methyl cytosine (R = Me) explored by computational DFT means. 

Mismatch Repair. Mispairs that break the normal base-pairing rules can arise spontaneously due to DNA biosynthetic errors, events associated with genetic recombination and the deamination of methylated cytosine (Modrich, 1987). With the latter, when cytosine deaminates to uracil, an endonuclease enzyme, /V-uracil-DNA glycosylase (Lindahl, 1979), excises the uracil residue before it can pair with adenine at the next replication. However, 5-methyl cytosine deaminates to form thymine and will not be excised by a glycosylase. As a result, thymine exits on one strand paired with guanine on the sister strand, that is, a mismatch. This will result in a spontaneous point mutation if left unrepaired. For this reason, methylated cytosines form spontaneous mutation hot-spots (Miller, 1985). The cell is able to repair mismatches by being able to distinguish between the DNA strand that exists before replication and a newly synthesized strand. 

Although chemically modifying DNA have distinctive implications for chromatin transitions and fiber structure in the presence of HI [250], in vivo these effects appear to work in concert with chromosomal proteins. 5 -Methylcytosines are specifically bound by members of the MBD (methyl-CpG-binding-domain) family, such as MeCP2 (Methyl-Cytosine binding Protein 2) and MBDl. These proteins have been shown to interact with HDACs and provide a casual link between DNA methylation, histone deacetylation and transcriptional repression [251-253]. 

Fig. 8. Proposed models that link histone methylation to DNA methylation (for details see Section 5.2). Methylated cytosines attract histone methyltransferases that contain a methyl-binding domain or a methyl-CpG binding protein (MeCP2) that recruits histone methylase activities these introduce methyl groups into the histone tails. The binding of chromodomain HPl proteins to H3 tails methylated at lysine 9 generates a secondary layer of repressive chromatin structure, (b) In a reverse scenario, methylated histone tails attract chromodomain-binding proteins, which in turn recruit Dmnts to methylate adjacent DNA sequences.

This massive amount of information should not be considered as insurmountable or only material to be marveled at but not understood. Much of the chemistry is already available to mine this information successfully. Much of it is understandable in somewhat simple terms, generally only after we have discovered the key to this simplicity. For instance, there is a marked decrease in the frequency of the dinucleotide CpG in some areas of the genome. The deficiency is believed to be due to the fact that most CpG nucleotides are methylated on the cytosine base, and spontaneous deamination of the methyl-cytosine residue creates T residues. Thus, CpG dinucleotide sequences mutate to TpG dinucleotides. But there still remain some questions. There are certain regions or islands where the CpG sequences exist in a nonmethylated form and where the frequency of CpG occurs within the expected or normal rate. Why These CpG islands are of particular interest because they are associated with the 5 ends of genes. 

Some time ago, an allyl-like radical was observed in irradiated crystals of 5 -dCMP [26]. This radical was thought to be a sugar radical, although no likely scheme was proposed for its formation. It now appears that this radical is formed on 5-methyl cytosine impurities in these crystals [27]. This radical forms by deprotonation at the methyl group of the cytosine cation, 5meCyt(Me—H) , and may have important consequences in the radiation chemistry of DNA since the ionization potential of 5-methyl cytosine is lower than that of either cytosine or thymine. 

The photochemical behavior of aqueous solutions of O-methyl cytosine (14, 2-methoxy-4-aminopyrimidine) was apparently different... 

FIGURE 25-27 Direct repair of alkylated bases by AlkB. The AlkB protein is an a-ketoglutarate-Fe2+-dependent dioxygenase. It catalyzes the oxidative demethylation of 1-methyladenine and 3-methyl-cytosine residues. 

DNA Repair Mechanisms Vertebrate and plant cells often methylate cytosine in DNA to form 5-methylcytosine (see Fig. 8-5a). In these same cells, a specialized repair system recognizes G-T mismatches and repairs them to G=C base pairs. How might this repair system be advantageous to the cell (Explain in terms of the presence of 5-methylcytosine in the DNA.)... 

From the comparison of ionization constants it was also deduced that both 1-methyl and 3-methyl cytosines exist predominantly in the lactam-amine forms 2 and 3, respectively.100,102 The conclusions based on a comparison of ionization constants have been confirmed by UV analysis (see below). 

For instance Shugar and Fox106 have shown that the spectrum of cytosine resembles that of 1-methylcytosine (form 2), not that of 2-methoxycytosine (form 1). Similarly, Kenner et al.100 have established that lactam-imine forms 6 were unimportant both for 1-methylcytosine (thus also for cytosine itself) and for its aminoacetyl derivatives. Katritzky and Waring60 have extended these studies and in particular showed that tautomers 3 and zwitterionic structures 10 were unimportant in the tautomeric equilibrium in both aqueous and dimethyl sulfoxide solutions. Although these last comparisons were made with only two partially methylated cytosines still capable of tautomerism, the reported conclusions agree with those reached later by Brown and Lyall96 on much wider evidence. 

CG doublets. The only modified base commonly found in eukaryotes is 5-methylcytosine,222/223 which upon deamination becomes thymine (Eq. 27-1). Most methylation occurs when C is followed by G. Usually 60-90% of all 5 -CG sequences (CpG sequences) in eukaryotic DNA are methylated. However, the fraction of methylated cytosine varies from almost zero for Drosophilia, Caenorhabditis, and Saccharomyces to as much as 30% in higher plants.224... 

A body of chemical information that proved vital to understanding DNA structure came from Erwin Chargaff s analyses of the nucleotide composition of duplex DNAs from various sources (table 25.1). Although the base compositions varied over a wide range, Chargaff found that within the DNA of each source that he examined, the amount of A was very nearly equal to the amount of T, and the amount of G was very nearly equal to the amount of C. The C is present as both unmodified C and, to a lesser extent, 5-methyl-cytosine, which results from postreplicative... 

Dekker and Elmore34 isolated 2-deoxy-5 -methylcytidine (VI, R = CH3) from enzymic hydrolyzates of wheat-germ deoxyribonucleic acid. (It had been shown previously32 that wheat germ is relatively rich in 5-methyl-cytosine.) By use of ion-exchange and partition chromatography, a small... 

The discovery of the presence of glucose in the nucleoside of 5-(hydroxy-methyl)cytosine was as unexpected as the original finding of the presence36 of 5-(hydroxymethyl)cytosine and the absence of cytosine.36 As Cohen suggests,84 other viruses contain cytosine they therefore lack the hydroxy -... 

It should be stated, however, that these experiments do not, of themselves, justify the authors conclusions. The isolation of 3-methylthymine surely excludes position 3 in thymidine as a point of union between the aglycon residue and the sugar residue. The isolation of a df-methylated cytosine does not even exclude position 3, since no experimental evidence what-... 

Finally, NGS can also be applied to epigenome analysis, currently one of the main areas of cancer research. By one of three different approaches, immunoprecipitation of methyl-cytosine after DNA fragmentation, capture of CpG islands and associated sequences, or complete genome sequencing after bisulfite treatment, all of them are directed to envision what promoter regions may be modified by methylation-based silencing (121,122). 

Clark SJ, Harrison J, Paul CL, Frommer M. High sensitivity mapping of methylated cytosines. Nucleic Acids Res 1994 22 2990-2997. 

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