Editing Epigenetics (section)

===DNA repair===
Damage to DNA is very common and is constantly being repaired. Epigenetic alterations can accompany DNA repair of oxidative damage or double-strand breaks. In human cells, oxidative DNA damage occurs about 10,000 times a day and DNA double-strand breaks occur about 10 to 50 times a cell cycle in somatic replicating cells (see [[DNA damage (naturally occurring)]]). The selective advantage of DNA repair is to allow the cell to survive in the face of DNA damage. The selective advantage of epigenetic alterations that occur with DNA repair is not clear.{{citation needed|date=March 2023}}

====Repair of oxidative DNA damage can alter epigenetic markers====
In the steady state (with endogenous damages occurring and being repaired), there are about 2,400 oxidatively damaged guanines that form [[8-oxo-2'-deoxyguanosine]] (8-OHdG) in the average mammalian cell DNA.<ref name="pmid21163908">{{cite journal |vauthors=Swenberg JA, Lu K, Moeller BC, Gao L, Upton PB, Nakamura J, Starr TB |title=Endogenous versus exogenous DNA adducts: their role in carcinogenesis, epidemiology, and risk assessment |journal=Toxicol Sci |volume=120 |issue= Suppl 1|pages=S130–45 |date=March 2011 |pmid=21163908 |pmc=3043087 |doi=10.1093/toxsci/kfq371 |url=}}</ref> 8-OHdG constitutes about 5% of the oxidative damages commonly present in DNA.<ref name=Hamilton>{{cite journal |vauthors=Hamilton ML, Guo Z, Fuller CD, Van Remmen H, Ward WF, Austad SN, Troyer DA, Thompson I, Richardson A |title=A reliable assessment of 8-oxo-2-deoxyguanosine levels in nuclear and mitochondrial DNA using the sodium iodide method to isolate DNA |journal=Nucleic Acids Res |volume=29 |issue=10 |pages=2117–26 |date=May 2001 |pmid=11353081 |pmc=55450 |doi=10.1093/nar/29.10.2117 |url=}}</ref> The oxidized guanines do not occur randomly among all guanines in DNA. There is a sequence preference for the guanine at a [[DNA methylation|methylated]] [[CpG site]] (a cytosine followed by guanine along its [[Directionality (molecular biology)|5' → 3' direction]] and where the cytosine is methylated (5-mCpG)).<ref name="pmid24571128">{{cite journal |vauthors=Ming X, Matter B, Song M, Veliath E, Shanley R, Jones R, Tretyakova N |title=Mapping structurally defined guanine oxidation products along DNA duplexes: influence of local sequence context and endogenous cytosine methylation |journal=J Am Chem Soc |volume=136 |issue=11 |pages=4223–35 |date=March 2014 |pmid=24571128 |pmc=3985951 |doi=10.1021/ja411636j |bibcode=2014JAChS.136.4223M |url=}}</ref> A 5-mCpG site has the lowest ionization potential for guanine oxidation.{{citation needed|date=March 2023}}

[[File:Initiation of DNA demethylation at a CpG site.svg|thumb|Initiation of [[DNA demethylation]] at a [[CpG site]]. In adult somatic cells DNA methylation typically occurs in the context of CpG dinucleotides ([[CpG sites]]), forming [[5-methylcytosine]]-pG, or 5mCpG. Reactive oxygen species (ROS) may attack guanine at the dinucleotide site, forming [[8-oxo-2'-deoxyguanosine|8-hydroxy-2'-deoxyguanosine]] (8-OHdG), and resulting in a 5mCp-8-OHdG dinucleotide site. The [[base excision repair]] enzyme [[oxoguanine glycosylase|OGG1]] targets 8-OHdG and binds to the lesion without immediate excision. OGG1, present at a 5mCp-8-OHdG site recruits [[Tet methylcytosine dioxygenase 1|TET1]] and TET1 oxidizes the 5mC adjacent to the 8-OHdG. This initiates demethylation of 5mC.<ref name=Zhou>{{cite journal |vauthors=Zhou X, Zhuang Z, Wang W, He L, Wu H, Cao Y, Pan F, Zhao J, Hu Z, Sekhar C, Guo Z |title=OGG1 is essential in oxidative stress-induced DNA demethylation |journal=Cell Signal |volume=28 |issue=9 |pages=1163–1171 |date=September 2016 |pmid=27251462 |doi=10.1016/j.cellsig.2016.05.021 |url=}}</ref>]]

Oxidized guanine has mispairing potential and is mutagenic.<ref name="pmid31993111">{{cite journal |vauthors=Poetsch AR |title=The genomics of oxidative DNA damage, repair, and resulting mutagenesis |journal=Comput Struct Biotechnol J |volume=18 |issue= |pages=207–219 |date=2020 |pmid=31993111 |pmc=6974700 |doi=10.1016/j.csbj.2019.12.013 |url=}}</ref> [[Oxoguanine glycosylase]] (OGG1) is the primary enzyme responsible for the excision of the oxidized guanine during DNA repair. OGG1 finds and binds to an 8-OHdG within a few seconds.<ref name="pmid33171795">{{cite journal |vauthors=D'Augustin O, Huet S, Campalans A, Radicella JP |title=Lost in the Crowd: How Does Human 8-Oxoguanine DNA Glycosylase 1 (OGG1) Find 8-Oxoguanine in the Genome? |journal=Int J Mol Sci |volume=21 |issue=21 |date=November 2020 |page=8360 |pmid=33171795 |pmc=7664663 |doi=10.3390/ijms21218360 |url=|doi-access=free }}</ref> However, OGG1 does not immediately excise 8-OHdG. In HeLa cells half maximum removal of 8-OHdG occurs in 30 minutes,<ref name="pmid15365186">{{cite journal |vauthors=Lan L, Nakajima S, Oohata Y, Takao M, Okano S, Masutani M, Wilson SH, Yasui A |title=In situ analysis of repair processes for oxidative DNA damage in mammalian cells |journal=Proc Natl Acad Sci U S A |volume=101 |issue=38 |pages=13738–43 |date=September 2004 |pmid=15365186 |pmc=518826 |doi=10.1073/pnas.0406048101 |bibcode=2004PNAS..10113738L |url=|doi-access=free }}</ref> and in irradiated mice, the 8-OHdGs induced in the mouse liver are removed with a half-life of 11 minutes.<ref name=Hamilton />

When OGG1 is present at an oxidized guanine within a methylated [[CpG site]] it recruits [[TET enzymes|TET1]] to the 8-OHdG lesion (see Figure). This allows TET1 to demethylate an adjacent methylated cytosine. Demethylation of cytosine is an epigenetic alteration.{{citation needed|date=March 2023}}

As an example, when human mammary epithelial cells were treated with H<sub>2</sub>O<sub>2</sub> for six hours, 8-OHdG increased about 3.5-fold in DNA and this caused about 80% demethylation of the 5-methylcytosines in the genome.<ref name=Zhou /> Demethylation of CpGs in a gene promoter by [[TET enzymes|TET enzyme]] activity increases transcription of the gene into messenger RNA.<ref name="pmid24108092">{{cite journal |vauthors=Maeder ML, Angstman JF, Richardson ME, Linder SJ, Cascio VM, Tsai SQ, Ho QH, Sander JD, Reyon D, Bernstein BE, Costello JF, Wilkinson MF, Joung JK |title=Targeted DNA demethylation and activation of endogenous genes using programmable TALE-TET1 fusion proteins |journal=Nat. Biotechnol. |volume=31 |issue=12 |pages=1137–42 |date=December 2013 |pmid=24108092 |pmc=3858462 |doi=10.1038/nbt.2726 }}</ref> In cells treated with H<sub>2</sub>O<sub>2</sub>, one particular gene was examined, [[Beta-secretase 1|''BACE1'']].<ref name=Zhou /> The methylation level of the ''BACE1'' [[CpG site#CpG islands|CpG island]] was reduced (an epigenetic alteration) and this allowed about 6.5 fold increase of expression of ''BACE1'' messenger RNA.{{citation needed|date=March 2023}}

While six-hour incubation with H<sub>2</sub>O<sub>2</sub> causes considerable demethylation of 5-mCpG sites, shorter times of H<sub>2</sub>O<sub>2</sub> incubation appear to promote other epigenetic alterations. Treatment of cells with H<sub>2</sub>O<sub>2</sub> for 30 minutes causes the mismatch repair protein heterodimer MSH2-MSH6 to recruit DNA methyltransferase 1 ([[DNMT1]]) to sites of some kinds of oxidative DNA damage.<ref name="pmid26186941">{{cite journal |vauthors=Ding N, Bonham EM, Hannon BE, Amick TR, Baylin SB, O'Hagan HM |title=Mismatch repair proteins recruit DNA methyltransferase 1 to sites of oxidative DNA damage |journal=J Mol Cell Biol |volume=8 |issue=3 |pages=244–54 |date=June 2016 |pmid=26186941 |pmc=4937888 |doi=10.1093/jmcb/mjv050 |url=}}</ref> This could cause increased methylation of cytosines (epigenetic alterations) at these locations.

Jiang et al.<ref name=Jiang>{{cite journal |vauthors=Jiang Z, Lai Y, Beaver JM, Tsegay PS, Zhao ML, Horton JK, Zamora M, Rein HL, Miralles F, Shaver M, Hutcheson JD, Agoulnik I, Wilson SH, Liu Y |title=Oxidative DNA Damage Modulates DNA Methylation Pattern in Human Breast Cancer 1 (BRCA1) Gene via the Crosstalk between DNA Polymerase β and a de novo DNA Methyltransferase |journal=Cells |volume=9 |issue=1 |date=January 2020 |page=225 |pmid=31963223 |pmc=7016758 |doi=10.3390/cells9010225 |url=|doi-access=free }}</ref> treated [[HEK 293 cells]] with agents causing oxidative DNA damage, ([[potassium bromate]] (KBrO3) or [[potassium chromate]] (K2CrO4)). [[Base excision repair]] (BER) of oxidative damage occurred with the DNA repair enzyme [[DNA polymerase|polymerase beta]] localizing to oxidized guanines. Polymerase beta is the main human polymerase in short-patch BER of oxidative DNA damage. Jiang et al.<ref name=Jiang /> also found that polymerase beta recruited the [[DNA methyltransferase]] protein DNMT3b to BER repair sites. They then evaluated the methylation pattern at the single nucleotide level in a small region of DNA including the [[promoter (genetics)|promoter]] region and the early transcription region of the [[BRCA1]] gene. Oxidative DNA damage from bromate modulated the DNA methylation pattern (caused epigenetic alterations) at CpG sites within the region of DNA studied. In untreated cells, CpGs located at −189, −134, −29, −19, +16, and +19 of the BRCA1 gene had methylated cytosines (where numbering is from the [[messenger RNA]] transcription start site, and negative numbers indicate nucleotides in the upstream [[Promoter (genetics)|promoter]] region). Bromate treatment-induced oxidation resulted in the loss of cytosine methylation at −189, −134, +16 and +19 while also leading to the formation of new methylation at the CpGs located at −80, −55, −21 and +8 after DNA repair was allowed.

====Homologous recombinational repair alters epigenetic markers====
At least four articles report the recruitment of [[DNA methyltransferase|DNA methyltransferase 1 (DNMT1)]] to sites of DNA double-strand breaks.<ref name="pmid15956212">{{cite journal |vauthors=Mortusewicz O, Schermelleh L, Walter J, Cardoso MC, Leonhardt H |title=Recruitment of DNA methyltransferase I to DNA repair sites |journal=Proc Natl Acad Sci U S A |volume=102 |issue=25 |pages=8905–9 |date=June 2005 |pmid=15956212 |pmc=1157029 |doi=10.1073/pnas.0501034102 |bibcode=2005PNAS..102.8905M |url=|doi-access=free }}</ref><ref name=Cuozzo>{{cite journal |vauthors=Cuozzo C, Porcellini A, Angrisano T, Morano A, Lee B, Di Pardo A, Messina S, Iuliano R, Fusco A, Santillo MR, Muller MT, Chiariotti L, Gottesman ME, Avvedimento EV |title=DNA damage, homology-directed repair, and DNA methylation |journal=PLOS Genet |volume=3 |issue=7 |pages=e110 |date=July 2007 |pmid=17616978 |pmc=1913100 |doi=10.1371/journal.pgen.0030110 |url= |doi-access=free }}</ref><ref name="pmid18704159">{{cite journal |vauthors=O'Hagan HM, Mohammad HP, Baylin SB |title=Double strand breaks can initiate gene silencing and SIRT1-dependent onset of DNA methylation in an exogenous promoter CpG island |journal=PLOS Genet |volume=4 |issue=8 |pages=e1000155 |date=August 2008 |pmid=18704159 |pmc=2491723 |doi=10.1371/journal.pgen.1000155 |url= |doi-access=free }}</ref><ref name="pmid20940144">{{cite journal |vauthors=Ha K, Lee GE, Palii SS, Brown KD, Takeda Y, Liu K, Bhalla KN, Robertson KD |title=Rapid and transient recruitment of DNMT1 to DNA double-strand breaks is mediated by its interaction with multiple components of the DNA damage response machinery |journal=Hum Mol Genet |volume=20 |issue=1 |pages=126–40 |date=January 2011 |pmid=20940144 |pmc=3000680 |doi=10.1093/hmg/ddq451 |url=}}</ref> During [[homologous recombination|homologous recombinational repair (HR)]] of the double-strand break, the involvement of DNMT1 causes the two repaired strands of DNA to have different levels of methylated cytosines. One strand becomes frequently methylated at about 21 [[CpG site]]s downstream of the repaired double-strand break. The other DNA strand loses methylation at about six CpG sites that were previously methylated downstream of the double-strand break, as well as losing methylation at about five CpG sites that were previously methylated upstream of the double-strand break. When the chromosome is replicated, this gives rise to one daughter chromosome that is heavily methylated downstream of the previous break site and one that is unmethylated in the region both upstream and downstream of the previous break site. With respect to the gene that was broken by the double-strand break, half of the progeny cells express that gene at a high level and in the other half of the progeny cells expression of that gene is repressed. When clones of these cells were maintained for three years, the new methylation patterns were maintained over that time period.<ref name="pmid27629060">{{cite journal |vauthors=Russo G, Landi R, Pezone A, Morano A, Zuchegna C, Romano A, Muller MT, Gottesman ME, Porcellini A, Avvedimento EV |title=DNA damage and Repair Modify DNA methylation and Chromatin Domain of the Targeted Locus: Mechanism of allele methylation polymorphism |journal=Sci Rep |volume=6 |issue= |pages=33222 |date=September 2016 |pmid=27629060 |pmc=5024116 |doi=10.1038/srep33222 |bibcode=2016NatSR...633222R |url=}}</ref>

In mice with a CRISPR-mediated homology-directed recombination insertion in their genome there were a large number of increased methylations of CpG sites within the double-strand break-associated insertion.<ref name="pmid33267773">{{cite journal |vauthors=Farris MH, Texter PA, Mora AA, Wiles MV, Mac Garrigle EF, Klaus SA, Rosfjord K |title=Detection of CRISPR-mediated genome modifications through altered methylation patterns of CpG islands |journal=BMC Genomics |volume=21 |issue=1 |pages=856 |date=December 2020 |pmid=33267773 |pmc=7709351 |doi=10.1186/s12864-020-07233-2 |url= |doi-access=free }}</ref>

====Non-homologous end joining can cause some epigenetic marker alterations====
[[Non-homologous end joining]] (NHEJ) repair of a double-strand break can cause a small number of demethylations of pre-existing cytosine DNA methylations downstream of the repaired double-strand break.<ref name=Cuozzo /> Further work by Allen et al.<ref name="pmid28423717">{{cite journal |vauthors=Allen B, Pezone A, Porcellini A, Muller MT, Masternak MM |title=Non-homologous end joining induced alterations in DNA methylation: A source of permanent epigenetic change |journal=Oncotarget |volume=8 |issue=25 |pages=40359–40372 |date=June 2017 |pmid=28423717 |pmc=5522286 |doi=10.18632/oncotarget.16122 |url=}}</ref> showed that NHEJ of a DNA double-strand break in a cell could give rise to some progeny cells having repressed expression of the gene harboring the initial double-strand break and some progeny having high expression of that gene due to epigenetic alterations associated with NHEJ repair. The frequency of epigenetic alterations causing repression of a gene after an NHEJ repair of a DNA double-strand break in that gene may be about 0.9%.<ref name="pmid18704159"/>