Editing Regulatory sequence (section)

===CpG island methylation and demethylation===

[[File:Cytosine and 5-methylcytosine.svg|thumb|262x262px|A methyl group is added on the carbon at the number 5 position of the ring to form 5-methylcytosine]]

[[5-methylcytosine|5-Methylcytosine]] (5-mC) is a [[methylation|methylated]] form of the [[DNA]] base [[cytosine]] (see figure).  5-mC is an [[Epigenetics|epigenetic]] marker found predominantly on cytosines within CpG dinucleotides, which consist of a cytosine is followed by a guanine reading in the 5' to 3' direction along the DNA strand ([[CpG sites]]).  About 28 million CpG dinucleotides occur in the human genome.<ref name="pmid26932361">{{cite journal | vauthors = Lövkvist C, Dodd IB, Sneppen K, Haerter JO | title = DNA methylation in human epigenomes depends on local topology of CpG sites | journal = Nucleic Acids Research | volume = 44 | issue = 11 | pages = 5123–32 | date = June 2016 | pmid = 26932361 | pmc = 4914085 | doi = 10.1093/nar/gkw124 }}</ref> In most tissues of mammals, on average, 70% to 80% of CpG cytosines are methylated (forming 5-methyl-CpG, or 5-mCpG).<ref name="pmid15177689">{{cite journal | vauthors = Jabbari K, Bernardi G | title = Cytosine methylation and CpG, TpG (CpA) and TpA frequencies | journal = Gene | volume = 333 | issue =  | pages = 143–9 | date = May 2004 | pmid = 15177689 | doi = 10.1016/j.gene.2004.02.043 }}</ref>  Methylated cytosines within CpG sequences often occur in groups, called [[CpG site#CpG islands|CpG islands]]. About 59% of promoter sequences have a CpG island while only about 6% of enhancer sequences have a CpG island.<ref name="pmid32338759">{{cite journal | vauthors = Steinhaus R, Gonzalez T, Seelow D, Robinson PN | title = Pervasive and CpG-dependent promoter-like characteristics of transcribed enhancers | journal = Nucleic Acids Research | volume = 48 | issue = 10 | pages = 5306–5317 | date = June 2020 | pmid = 32338759 | pmc = 7261191 | doi = 10.1093/nar/gkaa223 }}</ref> CpG islands constitute regulatory sequences, since if CpG islands are methylated in the promoter of a gene this can reduce or silence gene expression.<ref name="pmid11782440">{{cite journal | vauthors = Bird A | title = DNA methylation patterns and epigenetic memory | journal = Genes & Development | volume = 16 | issue = 1 | pages = 6–21 | date = January 2002 | pmid = 11782440 | doi = 10.1101/gad.947102 | doi-access = free }}</ref>

DNA methylation regulates gene expression through interaction with methyl binding domain (MBD) proteins, such as MeCP2, MBD1 and MBD2.  These MBD proteins bind most strongly to highly methylated [[CpG site#CpG islands|CpG islands]].<ref name=Du>{{cite journal | vauthors = Du Q, Luu PL, Stirzaker C, Clark SJ | title = Methyl-CpG-binding domain proteins: readers of the epigenome | journal = Epigenomics | volume = 7 | issue = 6 | pages = 1051–73 | date = 2015 | pmid = 25927341 | doi = 10.2217/epi.15.39 | doi-access = free }}</ref>  These MBD proteins have both a methyl-CpG-binding domain and a transcriptional repression domain.<ref name=Du />  They bind to methylated DNA and guide or direct protein complexes with chromatin remodeling and/or histone modifying activity to methylated CpG islands. MBD proteins generally repress local chromatin by means such as catalyzing the introduction of repressive histone marks or creating an overall repressive chromatin environment through [[nucleosome]] remodeling and chromatin reorganization.<ref name=Du />

[[Transcription factors]] are proteins that bind to specific DNA sequences in order to regulate the expression of a given gene.  The binding sequence for a transcription factor in DNA is usually about 10 or 11 nucleotides long.  There are approximately 1,400 different transcription factors encoded in the human genome, and they constitute about 6% of all human protein coding genes.<ref name="pmid19274049">{{cite journal | vauthors = Vaquerizas JM, Kummerfeld SK, Teichmann SA, Luscombe NM | title = A census of human transcription factors: function, expression and evolution | journal = Nature Reviews. Genetics | volume = 10 | issue = 4 | pages = 252–63 | date = April 2009 | pmid = 19274049 | doi = 10.1038/nrg2538 | s2cid = 3207586 }}</ref>  About 94% of transcription factor binding sites that are associated with signal-responsive genes occur in enhancers while only about 6% of such sites occur in promoters.<ref name="pmid29987030"/>

[[EGR1]] is a transcription factor important for regulation of methylation of CpG islands.  An EGR1 transcription factor binding site is frequently located in enhancer or promoter sequences.<ref name=SunZ>{{cite journal | vauthors = Sun Z, Xu X, He J, Murray A, Sun MA, Wei X, Wang X, McCoig E, Xie E, Jiang X, Li L, Zhu J, Chen J, Morozov A, Pickrell AM, Theus MH, Xie H | display-authors = 6 | title = EGR1 recruits TET1 to shape the brain methylome during development and upon neuronal activity | journal = Nature Communications | volume = 10 | issue = 1 | pages = 3892 | date = August 2019 | pmid = 31467272 | pmc = 6715719 | doi = 10.1038/s41467-019-11905-3 | bibcode = 2019NatCo..10.3892S }}</ref>  There are about 12,000 binding sites for EGR1 in the mammalian genome and about half of EGR1 binding sites are located in promoters and half in enhancers.<ref name=SunZ />  The binding of EGR1 to its target DNA binding site is insensitive to cytosine methylation in the DNA.<ref name=SunZ />

While only small amounts of EGR1 protein are detectable in cells that are un-stimulated, EGR1 translation into protein at one hour after stimulation is markedly elevated.<ref name=Kubosaki>{{cite journal | vauthors = Kubosaki A, Tomaru Y, Tagami M, Arner E, Miura H, Suzuki T, Suzuki M, Suzuki H, Hayashizaki Y | display-authors = 6 | title = Genome-wide investigation of in vivo EGR-1 binding sites in monocytic differentiation | journal = Genome Biology | volume = 10 | issue = 4 | pages = R41 | date = 2009 | pmid = 19374776 | pmc = 2688932 | doi = 10.1186/gb-2009-10-4-r41 | doi-access = free }}</ref>  Expression of EGR1 in various types of cells can be stimulated by growth factors, neurotransmitters, hormones, stress and injury.<ref name=Kubosaki />  In the brain, when neurons are activated, EGR1 proteins are upregulated, and they bind to (recruit) pre-existing TET1 enzymes, which are highly expressed in neurons.  [[TET enzymes]] can catalyze demethylation of 5-methylcytosine.  When EGR1 transcription factors bring TET1 enzymes to EGR1 binding sites in promoters, the TET enzymes can [[DNA demethylation|demethylate]] the methylated CpG islands at those promoters.  Upon demethylation, these promoters can then initiate transcription of their target genes.  Hundreds of genes in neurons are differentially expressed after neuron activation through EGR1 recruitment of TET1 to methylated regulatory sequences in their promoters.<ref name=SunZ />