“The DNA methylation is a process of addition of methyl group to the bases of DNA thereby inactivating a gene function.”
The epigenetic alterations are those which affects directly the gene expression.
For example, DNA methylation, histone modification, acetylation and chromatin remodelling are some of those alterations categorised into epigenetic alterations.
The main function of a gene- the functional part of the DNA, is to form a protein. Formation of DNA to protein occurs via the mRNA intermediate. A gene transcribed into the mRNA and translated into the chain of amino acid.
Now, this is the main function of genes in us, however, the amount of gene expressed in each cell or tissue is also very crucial.
The entire branch or sub-branch of science which deals with the study of gene expression is known as epigenetics.
An epigenetic alteration or we can say an epigenetic mutation is totally different from the genetic mutation.
In the genetic mutations, a change in the DNA sequence occurs due to the deletion, addition, translocation or inversion which influence gene function.
While in the epigenetic alteration sequence variation is not observed in a gene still, expression of a gene-altered. Thus the alterations are not directly associated with a change in the DNA sequence.
Alike the genetic alterations, epigenetic alterations might be either heritable or non-heritable.
What is DNA methylation?
DNA methylation is one of the epigenetic alterations which regulates or on or off gene expression by adding the methyl group to the gene or DNA strand.
Cytosine and CpG island, methyltransferase and DNA packaging play an important role in DNA methylation.
The DNMTs- DNA methyltransferase is the class of enzyme which regulates and catalyse the methylation reaction.
Those enzymes are divided into DNMT1, DNMT2, DNMT3A, DNMT3b and DNMTL.
The enzymes used in the catalytic process of DNA methylation can be divided into three broad categories.
The enzyme DNMT3a and DNMT3b are categorised into the inserter which incorporates methyl group to de novo.
This means that these enzymes insert methyl group at a totally new location and therefore it regulates gene expression.
Overactivity of DNMT3a and DNMT3b are reported in the hypermethylated DNA and cancer-like conditions.
Though both DNMT3a and DNMT3b are very much similar to the DNMT1, unlike the DNMT1, it methylates DNA bases not necessarily hemimethylated.
The DMNT1, DNMT3a and DNMT3b shares almost similar structure with larger regulatory N-terminal domain and catalytic C-terminus domain.
The DNMT1 enzyme is categorised into the maintenance category because it is used during the replication. It inserts the methyl group to the hemimethylated newly synthesised DNA.
The methyl group is inserted to the hemimethylated DNA of the newly synthesised DNA stranded.
The DNMT1 inserts methyl group at the same locations alike the original/parental DNA. It also repairs the errors in during the replication too.
Thus it is categorised into the maintenance enzyme.
The DNMT3L although does not have a catalytic domain, still, it enhances the activity of other members of this class.
Epigenetically reprogramming of the gene is mediated by the removal of the methyl group, hence a gene can be activated by removal of the methyl group.
The process of a removal methyl group from the DNA is known as demethylation.
Removal or methyl group or demethylation includes gene expression by facilitating transcription.
TET like enzymes mediated the process of DNA demethylation (TET- ten-eleven translocation).
The fifth base:
The major portion of the genome is made up of the non-coding DNA sequence and these sequences are compactly packed.
From the non-coding DNA, the major portion is GC rich DNA and are highly methylated.
The 5th carbon of the cytosine base is more prone to the methylation and thus often methylated, convert into the 5-methylcytosine (5-mC).
Approximately, 1.5% of the entire human genome consists of 5-methylcytosine henceforth it is also known as the “fifth base”.
Although, it is not naturally occurring in the DNA through replication.
5-mC is commonly present in DNA major groove and prevent the binding of enzymes for gene expression and hence regulates gene expression.
The fifth base or 5-mC commonly occurs in the GC dinucleotide rich region of the genome known as CpG island in somatic cells, however, the 5-mC is non-CpG regions in embryonic cells.
A post replication modification in which the addition of the methyl group mainly to the CpG island (CG dinucleotide rich region) deactivates gene activity.
An active gene can be repressed or deactivated by addition of the methyl group on its promoter region, by doing this, it prevents the binding of enzymes and other proteins involved in the gene expression.
Ultimately, the expression of a gene is regulated.
The promoter regions of a gene are non-coding CG rich region of DNA which are highly prone to DNA methylation.
DNA hypermethylation: Increase in methylation
DNA hypomethylation: decrease in methylation
Methods for detection of methylation:
Alike the DNA mutation or other epigenetic alterations, scientists are believed that the DNA methylation also plays an important role in the development of the disease.
A unique DNA methylation pattern of individuals makes them adaptive for the local environment, however, by studying DNA methylation scientists can determine many facts related to epigenetic alterations.
Some of the methods for studying DNA methylation are enlisted here:
DNA bisulphite PCR:
Bisulfite and sodium bisulfite are commonly used in the gene-specific methylation studies or we can say studying methylation associated with a particular gene.
Now the entire concept of using the bisulphite is quite amazing which is used to differentiate the cytosine from the methylated cytosine.
By using the bisulphite, the cytosine bases of a gene are converted into the uracil while the methylcytosine bases remain unchanged.
This will allow the researcher to differentiate the unmethylated area of the gene from the methylated area however, for doing this complete conversation of cytosine to uracil is required.
Called an MS-PCR, the template or amplicon prepared from this method is used in the downstream application such as bisulphite DNA sequencing and methylation-based- DNA microarray.
Methylated DNA quantification:
Another method used to detect the amount of methylation in the DNA methylation quantification. The hypomethylation caused by the deficiency of the methyl and involved in the development of cancer.
Some of the environmental factors are responsible for the global decrease in the methylation amount can be encountered by quantifying the methylated DNA.
Bisulfite DNA sequencing and methylation microarray are two other high-end applications used in the quantification of the methylated DNA or amount of methylation.
Besides this, Methylated DNA immunoprecipitation and DNA methyltransferase/ demethylase assays are two immuno-genetic techniques used to do so.
conclusively we can say, “DNA methylation is a kind of on/off switch, the addition of methyl group turns off the gene expression switch while the demethylation terns on the gene expression switch.”
We need more sophisticated set up to screen vast data of DNA methylation.
Moore LD, Le T, Fan G. DNA methylation and its basic function. Neuropsychopharmacology. 2013;38(1):23–38. doi:10.1038/npp.2012.112.