24. Gackowski D, Zarakowska E, Starczak M, Modrzejewska M, Olinski R (2015) Tissuespecific differences in DNA modifications (5-hydroxymethylcytosine, 5-formylcytosine,
5-carboxylcytosine and 5-hydroxymethyluracil) and their interrelationships. PLoS One 10
(12):e0144859. https://doi.org/10.1371/journal.pone.0144859
25. Heyn H, Esteller M (2015) An adenine code for DNA: a second life for N6-methyladenine.
Cell 161(4):710–713. https://doi.org/10.1016/j.cell.2015.04.021
26. Koziol MJ, Bradshaw CR, Allen GE, Costa ASH, Frezza C, Gurdon JB (2016) Identification
of methylated deoxyadenosines in vertebrates reveals diversity in DNA modifications.
Nat Struct Mol Biol 23(1):24–30. https://doi.org/10.1038/nsmb.3145
27. Achwal CW, Iyer CA, Chandra HS (1983) Immunochemical evidence for the presence of
5mC, 6mA and 7mG in human, Drosophila and mealybug DNA. FEBS Lett 158(2):353–358
28. Ratel D, Ravanat JL, Charles MP, Platet N, Breuillaud L, Lunardi J, Berger F, Wion D (2006)
Undetectable levels of N6-methyl adenine in mouse DNA: cloning and analysis of PRED28, a
gene coding for a putative mammalian DNA adenine methyltransferase. FEBS Lett 580
(13):3179–3184. https://doi.org/10.1016/j.febslet.2006.04.074
29. Wu TP, Wang T, Seetin MG, Lai Y, Zhu S, Lin K, Liu Y, Byrum SD, Mackintosh SG,
Zhong M, Tackett A, Wang G, Hon LS, Fang G, Swenberg JA, Xiao AZ (2016)
DNA methylation on N(6)-adenine in mammalian embryonic stem cells. Nature 532
(7599):329–333. https://doi.org/10.1038/nature17640
30. Xiao CL, Zhu S, He M, Chen D, Zhang Q, Chen Y, Yu G, Liu J, Xie SQ, Luo F, Liang Z,
Wang DP, Bo XC, Gu XF, Wang K, Yan GR (2018) N(6)-methyladenine DNA modification
in the human genome. Mol Cell 71(2):306–318.e307. https://doi.org/10.1016/j.molcel.2018.
06.015
31. Ehrlich M, Wilson GG, Kuo KC, Gehrke CW (1987) N4-methylcytosine as a minor base in
bacterial DNA. J Bacteriol 169(3):939–943
32. Timinskas A, Butkus V, Janulaitis A (1995) Sequence motifs characteristic for DNA
[cytosine-N4] and DNA [adenine-N6] methyltransferases. Classification of all DNA
methyltransferases. Gene 157(1–2):3–11
33. Yu M, Ji L, Neumann DA, Chung DH, Groom J, Westpheling J, He C, Schmitz RJ (2015)
Base-resolution detection of N4-methylcytosine in genomic DNA using 4mC-Tet-assistedbisulfite- sequencing. Nucleic Acids Res 43(21):e148. https://doi.org/10.1093/nar/gkv738
34. Sharma S, Kelly TK, Jones PA (2010) Epigenetics in cancer. Carcinogenesis 31(1):27–36.
https://doi.org/10.1093/carcin/bgp220
35. Chapman VL, Terranova R, Moggs JG, Kimber I, Dearman RJ (2016) Evaluation of
5-methylcytosine and 5-hydroxymethylcytosine as potential biomarkers for characterisation
of chemical allergens. Toxicology 340:17–26. https://doi.org/10.1016/j.tox.2015.12.003
36. Zhubi A, Chen Y, Dong E, Cook EH, Guidotti A, Grayson DR (2014) Increased binding of
MeCP2 to the GAD1 and RELN promoters may be mediated by an enrichment of 5-hmC in
autism spectrum disorder (ASD) cerebellum. Transl Psychiatry 4:e349
37. Kadowaki H, Nishitoh H, Urano F, Sadamitsu C, Matsuzawa A, Takeda K, Masutani H,
Yodoi J, Urano Y, Nagano T, Ichijo H (2005) Amyloid b induces neuronal cell death through
ROS-mediated ASK1 activation. Cell Death Differ 12(1):19–24
38. Gnyszka A, Jastrzebski Z, Flis S (2013) DNA methyltransferase inhibitors and their emerging
role in epigenetic therapy of cancer. Anticancer Res 33(8):2989–2996
39. Tsai HC, Li H, van Neste L, Cai Y, Robert C, Rassool FV, Shin JJ, Harbom KM, Beaty R,
Pappou E, Harris J, Yen RW, Ahuja N, Brock MV, Stearns V, Feller-Kopman D, Yarmus LB,
Lin YC, Welm AL, Issa JP, Minn I, Matsui W, Jang YY, Sharkis SJ, Baylin SB, Zahnow CA
(2012) Transient low doses of DNA-demethylating agents exert durable antitumor effects on
hematological and epithelial tumor cells. Cancer Cell 21(3):430–446. https://doi.org/10.1016/
j.ccr.2011.12.029
40. Flis S, Gnyszka A, Flis K (2014) DNA methyltransferase inhibitors improve the effect of
chemotherapeutic agents in SW48 and HT-29 colorectal cancer cells. PLoS One 9(3):e92305.
https://doi.org/10.1371/journal.pone.0092305
Experimental Methodologies for Detection and Mapping of Epigenetic DNA Marks
515
5-carboxylcytosine and 5-hydroxymethyluracil) and their interrelationships. PLoS One 10
(12):e0144859. https://doi.org/10.1371/journal.pone.0144859
25. Heyn H, Esteller M (2015) An adenine code for DNA: a second life for N6-methyladenine.
Cell 161(4):710–713. https://doi.org/10.1016/j.cell.2015.04.021
26. Koziol MJ, Bradshaw CR, Allen GE, Costa ASH, Frezza C, Gurdon JB (2016) Identification
of methylated deoxyadenosines in vertebrates reveals diversity in DNA modifications.
Nat Struct Mol Biol 23(1):24–30. https://doi.org/10.1038/nsmb.3145
27. Achwal CW, Iyer CA, Chandra HS (1983) Immunochemical evidence for the presence of
5mC, 6mA and 7mG in human, Drosophila and mealybug DNA. FEBS Lett 158(2):353–358
28. Ratel D, Ravanat JL, Charles MP, Platet N, Breuillaud L, Lunardi J, Berger F, Wion D (2006)
Undetectable levels of N6-methyl adenine in mouse DNA: cloning and analysis of PRED28, a
gene coding for a putative mammalian DNA adenine methyltransferase. FEBS Lett 580
(13):3179–3184. https://doi.org/10.1016/j.febslet.2006.04.074
29. Wu TP, Wang T, Seetin MG, Lai Y, Zhu S, Lin K, Liu Y, Byrum SD, Mackintosh SG,
Zhong M, Tackett A, Wang G, Hon LS, Fang G, Swenberg JA, Xiao AZ (2016)
DNA methylation on N(6)-adenine in mammalian embryonic stem cells. Nature 532
(7599):329–333. https://doi.org/10.1038/nature17640
30. Xiao CL, Zhu S, He M, Chen D, Zhang Q, Chen Y, Yu G, Liu J, Xie SQ, Luo F, Liang Z,
Wang DP, Bo XC, Gu XF, Wang K, Yan GR (2018) N(6)-methyladenine DNA modification
in the human genome. Mol Cell 71(2):306–318.e307. https://doi.org/10.1016/j.molcel.2018.
06.015
31. Ehrlich M, Wilson GG, Kuo KC, Gehrke CW (1987) N4-methylcytosine as a minor base in
bacterial DNA. J Bacteriol 169(3):939–943
32. Timinskas A, Butkus V, Janulaitis A (1995) Sequence motifs characteristic for DNA
[cytosine-N4] and DNA [adenine-N6] methyltransferases. Classification of all DNA
methyltransferases. Gene 157(1–2):3–11
33. Yu M, Ji L, Neumann DA, Chung DH, Groom J, Westpheling J, He C, Schmitz RJ (2015)
Base-resolution detection of N4-methylcytosine in genomic DNA using 4mC-Tet-assistedbisulfite- sequencing. Nucleic Acids Res 43(21):e148. https://doi.org/10.1093/nar/gkv738
34. Sharma S, Kelly TK, Jones PA (2010) Epigenetics in cancer. Carcinogenesis 31(1):27–36.
https://doi.org/10.1093/carcin/bgp220
35. Chapman VL, Terranova R, Moggs JG, Kimber I, Dearman RJ (2016) Evaluation of
5-methylcytosine and 5-hydroxymethylcytosine as potential biomarkers for characterisation
of chemical allergens. Toxicology 340:17–26. https://doi.org/10.1016/j.tox.2015.12.003
36. Zhubi A, Chen Y, Dong E, Cook EH, Guidotti A, Grayson DR (2014) Increased binding of
MeCP2 to the GAD1 and RELN promoters may be mediated by an enrichment of 5-hmC in
autism spectrum disorder (ASD) cerebellum. Transl Psychiatry 4:e349
37. Kadowaki H, Nishitoh H, Urano F, Sadamitsu C, Matsuzawa A, Takeda K, Masutani H,
Yodoi J, Urano Y, Nagano T, Ichijo H (2005) Amyloid b induces neuronal cell death through
ROS-mediated ASK1 activation. Cell Death Differ 12(1):19–24
38. Gnyszka A, Jastrzebski Z, Flis S (2013) DNA methyltransferase inhibitors and their emerging
role in epigenetic therapy of cancer. Anticancer Res 33(8):2989–2996
39. Tsai HC, Li H, van Neste L, Cai Y, Robert C, Rassool FV, Shin JJ, Harbom KM, Beaty R,
Pappou E, Harris J, Yen RW, Ahuja N, Brock MV, Stearns V, Feller-Kopman D, Yarmus LB,
Lin YC, Welm AL, Issa JP, Minn I, Matsui W, Jang YY, Sharkis SJ, Baylin SB, Zahnow CA
(2012) Transient low doses of DNA-demethylating agents exert durable antitumor effects on
hematological and epithelial tumor cells. Cancer Cell 21(3):430–446. https://doi.org/10.1016/
j.ccr.2011.12.029
40. Flis S, Gnyszka A, Flis K (2014) DNA methyltransferase inhibitors improve the effect of
chemotherapeutic agents in SW48 and HT-29 colorectal cancer cells. PLoS One 9(3):e92305.
https://doi.org/10.1371/journal.pone.0092305
Experimental Methodologies for Detection and Mapping of Epigenetic DNA Marks
515