Research Progress in DNA Methylation Sequencing Technology

doi:10.13560/j.cnki.biotech.bull.1985.2023-1166

Abstract

Abstract:

DNA methylation is an important epigenetic modification that plays a key role in the growth and development of animals and plants, as well as in the occurrence of diseases and the regulation of gene expression. Clinically, DNA methylation tumor markers can serve as biological markers for the diagnosis, prognosis, and treatment of tumors. Accurate detection of DNA methylation is of great significance for elucidating the life mechanisms of biological growth, development, and disease occurrence. DNA methylation sequencing technology is a powerful tool for studying DNA methylation and is widely used for mapping DNA methylation in the genome. In recent years, in order to better detect DNA methylation site information, scientists have proposed a series of high-throughput DNA methylation sequencing technologies, which have improved sequencing detection sensitivity, reduced sequencing costs and experimental expenses, and greatly promoted the development of epigenomics. This article reviews a series of DNA methylation sequencing technologies, focusing on the technical principles and application scenarios of three sequencing technologies: WGBS, TAPS, and EM-seq. It also introduces sequencing methods for locating DNA methylation at the single-cell level, and finally prospects the future development direction.

Key words: DNA methylation, WGBS, TAPS, EM-seq, single cell, multiomics

YUAN Ming-bo, YE Guo-hua, YANG Dan, SONG Dong-xue. Research Progress in DNA Methylation Sequencing Technology[J]. Biotechnology Bulletin, 2024, 40(5): 58-65.

Figures/Tables 5

References 53

[1]	Guo HS, Zhu P, Yan LY, et al. The DNA methylation landscape of human early embryos[J]. Nature, 2014, 511(7511): 606-610.
[2]	Yang QW, Ali M, Treviño LS, et al. Developmental reprogramming of myometrial stem cells by endocrine disruptor linking to risk of uterine fibroids[J]. Cell Mol Life Sci, 2023, 80(9): 274. doi: 10.1007/s00018-023-04919-0 pmid: 37650943
[3]	Klutstein M, Nejman D, Greenfield R, et al. DNA methylation in cancer and aging[J]. Cancer Research, 2016, 76(12):3446-3450. doi: 10.1158/0008-5472.CAN-15-3278 pmid: 27256564
[4]	Vasanthakumar A, Godley LA. 5-hydroxymethylcytosine in cancer: significance in diagnosis and therapy[J]. Cancer Genet, 2015, 208(5): 167-177.
[5]	Seale K, Horvath S, Teschendorff A, et al. Making sense of the ageing methylome[J]. Nat Rev Genet, 2022, 23(10): 585-605. doi: 10.1038/s41576-022-00477-6 pmid: 35501397
[6]	He L, Huang H, Bradai M, et al. DNA methylation-free Arabidopsis reveals crucial roles of DNA methylation in regulating gene expression and development[J]. Nat Commun, 2022, 13(1): 1335.
[7]	Ma YZ, Min L, Wang MJ, et al. Disrupted genome methylation in response to high temperature has distinct affects on microspore abortion and anther indehiscence[J]. Plant Cell, 2018, 30(7): 1387-1403.
[8]	Takahashi Y, Morales Valencia M, Yu Y, et al. Transgenerational inheritance of acquired epigenetic signatures at CpG islands in mice[J]. Cell, 2023, 186(4): 715-731.e19. doi: 10.1016/j.cell.2022.12.047 pmid: 36754048
[9]	Zhang HM, Lang ZB, Zhu JK. Dynamics and function of DNA methylation in plants[J]. Nat Rev Mol Cell Biol, 2018, 19(8): 489-506.
[10]	Frommer M, McDonald LE, Millar DS, et al. A genomic sequencing protocol that yields a positive display of 5-methylcytosine residues in individual DNA strands[J]. Proc Natl Acad Sci USA, 1992, 89(5): 1827-1831. doi: 10.1073/pnas.89.5.1827 pmid: 1542678
[11]	Meissner A, Gnirke A, Bell GW, et al. Reduced representation bisulfite sequencing for comparative high-resolution DNA methylation analysis[J]. Nucleic Acids Res, 2005, 33(18): 5868-5877. doi: 10.1093/nar/gki901 pmid: 16224102
[12]	Weber M, Davies JJ, Wittig D, et al. Chromosome-wide and promoter-specific analyses identify sites of differential DNA methylation in normal and transformed human cells[J]. Nat Genet, 2005, 37(8): 853-862. pmid: 16007088
[13]	Lister R, Pelizzola M, Dowen RH, et al. Human DNA methylomes at base resolution show widespread epigenomic differences[J]. Nature, 2009, 462(7271): 315-322.
[14]	Booth MJ, Branco MR, Ficz G, et al. Quantitative sequencing of 5-methylcytosine and 5-hydroxymethylcytosine at single-base resolution[J]. Science, 2012, 336(6083): 934-937. doi: 10.1126/science.1220671 pmid: 22539555
[15]	Yu M, Hon GC, Szulwach KE, et al. Base-resolution analysis of 5-hydroxymethylcytosine in the mammalian genome[J]. Cell, 2012, 149(6): 1368-1380. doi: 10.1016/j.cell.2012.04.027 pmid: 22608086
[16]	Schutsky EK, DeNizio JE, Hu P, et al. Nondestructive, base-resolution sequencing of 5-hydroxymethylcytosine using a DNA deaminase[J]. Nat Biotechnol, 2018. DOI: 10.1038/nbt.4204.
[17]	Liu YB, Siejka-Zielińska P, Velikova G, et al. Bisulfite-free direct detection of 5-methylcytosine and 5-hydroxymethylcytosine at base resolution[J]. Nat Biotechnol, 2019, 37(4): 424-429. doi: 10.1038/s41587-019-0041-2 pmid: 30804537
[18]	Vaisvila R, Chaithanya Ponnaluri VK, Sun ZY, et al. Enzymatic methyl sequencing detects DNA methylation at single-base resolution from picograms of DNA[J]. Genome Res, 2021, 31(7): 1280-1289. doi: 10.1101/gr.266551.120 pmid: 34140313
[19]	Gao XF, Song Y, Wu JL, et al. Iron-dependent epigenetic modulation promotes pathogenic T cell differentiation in lupus[J]. J Clin Invest, 2022, 132(9): e152345.
[20]	Mazid MA, Ward C, Luo ZW, et al. Rolling back human pluripotent stem cells to an eight-cell embryo-like stage[J]. Nature, 2022, 605(7909): 315-324.
[21]	Liang WW, Lu RJH, Jayasinghe RG, et al. Integrative multi-omic cancer profiling reveals DNA methylation patterns associated with therapeutic vulnerability and cell-of-origin[J]. Cancer Cell, 2023, 41(9): 1567-1585.e7.
[22]	Heyn H, Li N, Ferreira HJ, et al. Distinct DNA methylomes of newborns and centenarians[J]. Proc Natl Acad Sci USA, 2012, 109(26): 10522-10527. doi: 10.1073/pnas.1120658109 pmid: 22689993
[23]	Thürmann L, Klös M, Mackowiak SD, et al. Global hypomethylation in childhood asthma identified by genome-wide DNA-methylation sequencing preferentially affects enhancer regions[J]. Allergy, 2023, 78(6): 1489-1506.
[24]	Zheng GH, Hu SQ, Cheng SM, et al. Factor of DNA methylation 1 affects woodland strawberry plant stature and organ size via DNA methylation[J]. Plant Physiol, 2023, 191(1): 335-351.
[25]	Tahiliani M, Koh KP, Shen YH, et al. Conversion of 5-methylcytosine to 5-hydroxymethylcytosine in mammalian DNA by MLL partner TET1[J]. Science, 2009, 324(5929): 930-935. doi: 10.1126/science.1170116 pmid: 19372391
[26]	Ito S, Shen L, Dai Q, et al. Tet proteins can convert 5-methylcytosine to 5-formylcytosine and 5-carboxylcytosine[J]. Science, 2011, 333(6047): 1300-1303. doi: 10.1126/science.1210597 pmid: 21778364
[27]	Siejka-Zielińska P, Cheng JF, Jackson F, et al. Cell-free DNA TAPS provides multimodal information for early cancer detection[J]. Sci Adv, 2021, 7(36): eabh0534.
[28]	Liu YB, Cheng JF, Siejka-Zielińska P, et al. Accurate targeted long-read DNA methylation and hydroxymethylation sequencing with TAPS[J]. Genome Biol, 2020, 21(1): 54. doi: 10.1186/s13059-020-01969-6 pmid: 32127008
[29]	Wang T, Fowler JM, Liu L, et al. Direct enzymatic sequencing of 5-methylcytosine at single-base resolution[J]. Nat Chem Biol, 2023, 19(8): 1004-1012. doi: 10.1038/s41589-023-01318-1 pmid: 37322153
[30]	Hu LL, Lu JY, Cheng JD, et al. Structural insight into substrate preference for TET-mediated oxidation[J]. Nature, 2015, 527(7576): 118-122.
[31]	Xiong J, Chen KK, Xie NB, et al. Bisulfite-free and single-base resolution detection of epigenetic DNA modification of 5-methylcytosine by methyltransferase-directed labeling with APOBEC3A deamination sequencing[J]. Anal Chem, 2022, 94(44): 15489-15498.
[32]	Feng SH, Zhong ZH, Wang M, et al. Efficient and accurate determination of genome-wide DNA methylation patterns in Arabidopsis thaliana with enzymatic methyl sequencing[J]. Epigenetics Chromatin, 2020, 13(1): 42.
[33]	Trinidad EM, Vidal E, Coronado E, et al. Liquidhope: methylome and genomic profiling from very limited quantities of plasma-derived DNA[J]. Brief Bioinform, 2023, 24(1): bbac575.
[34]	Trinidad EM, Juan-Ribelles A, Pisano G, et al. Evaluation of circulating tumor DNA by electropherogram analysis and methylome profiling in high-risk neuroblastomas[J]. Front Oncol, 2023, 13: 1037342.
[35]	Guo P, Zheng HL, Li YH, et al. Hepatocellular carcinoma detection via targeted enzymatic methyl sequencing of plasma cell-free DNA[J]. Clin Epigenetics, 2023, 15(1): 2.
[36]	https://www.neb.cn/zh-cn/-/media/nebus/files/application-notes/technote_nebnext_enzymatic_methyl-seq.pdf?rev=015e017f782a4bc9b50b61f1b0f3c807
[37]	Williams CJ, Dai DW, Tran KA, et al. Dynamic DNA methylation turnover in gene bodies is associated with enhanced gene expression plasticity in plants[J]. Genome Biol, 2023, 24(1): 227. doi: 10.1186/s13059-023-03059-9 pmid: 37828516
[38]	Navin NE. Delineating cancer evolution with single-cell sequencing[J]. Sci Transl Med, 2015, 7(296): 296fs29.
[39]	Farlik M, Sheffield NC, Nuzzo A, et al. Single-cell DNA methylome sequencing and bioinformatic inference of epigenomic cell-state dynamics[J]. Cell Rep, 2015, 10(8): 1386-1397. doi: 10.1016/j.celrep.2015.02.001 pmid: 25732828
[40]	Angermueller C, Clark SJ, Lee HJ, et al. Parallel single-cell sequencing links transcriptional and epigenetic heterogeneity[J]. Nat Methods, 2016, 13(3): 229-232. doi: 10.1038/nmeth.3728 pmid: 26752769
[41]	Pott S. Simultaneous measurement of chromatin accessibility, DNA methylation, and nucleosome phasing in single cells[J]. eLife, 2017, 6: e23203.
[42]	Mulqueen RM, Pokholok D, Norberg SJ, et al. Highly scalable generation of DNA methylation profiles in single cells[J]. Nat Biotechnol, 2018, 36(5): 428-431. doi: 10.1038/nbt.4112 pmid: 29644997
[43]	Luo CY, Rivkin A, Zhou JT, et al. Robust single-cell DNA methylome profiling with snmC-seq2[J]. Nat Commun, 2018, 9(1): 3824. doi: 10.1038/s41467-018-06355-2 pmid: 30237449
[44]	Clark SJ, Argelaguet R, Kapourani CA, et al. scNMT-seq enables joint profiling of chromatin accessibility DNA methylation and transcription in single cells[J]. Nat Commun, 2018, 9(1): 781. doi: 10.1038/s41467-018-03149-4 pmid: 29472610
[45]	Bian SH, Hou Y, Zhou X, et al. Single-cell multiomics sequencing and analyses of human colorectal cancer[J]. Science, 2018, 362(6418): 1060-1063. doi: 10.1126/science.aao3791 pmid: 30498128
[46]	Guo F, Li L, Li JY, et al. Single-cell multi-omics sequencing of mouse early embryos and embryonic stem cells[J]. Cell Res, 2017, 27(8): 967-988. doi: 10.1038/cr.2017.82 pmid: 28621329
[47]	Gu C, Liu SL, Wu QH, et al. Integrative single-cell analysis of transcriptome, DNA methylome and chromatin accessibility in mouse oocytes[J]. Cell Res, 2019, 29(2): 110-123. doi: 10.1038/s41422-018-0125-4 pmid: 30560925
[48]	Han YN, Zheleznyakova GY, Marincevic-Zuniga Y, et al. Comparison of EM-seq and PBAT methylome library methods for low-input DNA[J]. Epigenetics, 2022, 17(10): 1195-1204.
[49]	Cao YL, Bai YL, Yuan TJ, et al. Single-cell bisulfite-free 5mC and 5hmC sequencing with high sensitivity and scalability[J]. Proc Natl Acad Sci U S A, 2023, 120(49): e2310367120.
[50]	Li J, Liang Y, Fan J, et al. DNA methylation subtypes guiding prognostic assessment and linking to responses the DNA methyltransferase inhibitor SGI-110 in urothelial carcinoma[J]. BMC Med, 2022, 20(1): 222. doi: 10.1186/s12916-022-02426-w pmid: 35843958
[51]	Ka-Yue Chow L, Lai-Shun Chung D, Tao LH, et al. Epigenomic landscape study reveals molecular subtypes and EBV-associated regulatory epigenome reprogramming in nasopharyngeal carcinoma[J]. EBioMedicine, 2022, 86: 104357.
[52]	Lee YH, Ren DA, Jeon B, et al. S-Adenosylmethionine: more than just a methyl donor[J]. Nat Prod Rep, 2023, 40(9): 1521-1549.
[53]	Wang T, Kohli RM. Discovery of an unnatural DNA modification derived from a natural secondary metabolite[J]. Cell Chem Biol, 2021, 28(1): 97-104.e4. doi: 10.1016/j.chembiol.2020.09.006 pmid: 33053370

方法 Method	DNA片段化 DNA framentation	酶氧化 Enzymatic oxidation	强酸强碱Strong acids and alkalis	脱氨方式 Deamination method	扩增方式 Amplification method	文库丰富度 Library richness	数据分析软件 Data analysis software
WGBS	亚硫酸氢盐打断/超声打断	不使用	使用	亚硫酸氢盐	PCR	低	Bismark & BWAMeth
TAPS	超声打断	TET1氧化	不使用	吡啶硼烷	PCR	高	asTair
EM-seq	超声打断	TET2氧化	不使用	APOBEC3A脱氨酶	PCR	低	Bismark & BWAMeth

方法 Method	DNA片段化 DNA framentation	酶氧化 Enzymatic oxidation	强酸强碱Strong acids and alkalis	脱氨方式 Deamination method	扩增方式 Amplification method	文库丰富度 Library richness	数据分析软件 Data analysis software
WGBS	亚硫酸氢盐打断/超声打断	不使用	使用	亚硫酸氢盐	PCR	低	Bismark & BWAMeth
TAPS	超声打断	TET1氧化	不使用	吡啶硼烷	PCR	高	asTair
EM-seq	超声打断	TET2氧化	不使用	APOBEC3A脱氨酶	PCR	低	Bismark & BWAMeth