Citation: Jun Xiong, Ke-Ke Chen, Neng-Bin Xie, Wei Chen, Wen-Xuan Shao, Tong-Tong Ji, Si-Yu Yu, Yu-Qi Feng, Bi-Feng Yuan. Demethylase-assisted site-specific detection of N¹-methyladenosine in RNA[J]. Chinese Chemical Letters, ;2024, 35(5): 108953. doi: 10.1016/j.cclet.2023.108953 shu

Demethylase-assisted site-specific detection of N¹-methyladenosine in RNA

Jun Xiong ^{a, 1} ,
Ke-Ke Chen ^{a, 1} ,
Neng-Bin Xie ^{a, 1} ,
Wei Chen ^{b, 1} ,
Wen-Xuan Shao ^a ,
Tong-Tong Ji ^a ,
Si-Yu Yu ^a ,
Yu-Qi Feng ^a ,
Bi-Feng Yuan ^{a, *,}

a.
College of Chemistry and Molecular Sciences, Research Center of Public Health, Renmin Hospital of Wuhan University, School of Public Health, Wuhan University, Wuhan 430071, China
b.
Department of Laboratory Medicine, Zhongnan Hospital of Wuhan University, Wuhan University, Wuhan 430071, China

bfyuan@whu.edu.cn

Received Date: 15 May 2023
Revised Date: 7 August 2023
Accepted Date: 21 August 2023
Available Online: 24 August 2023

Figures(4)

The dynamic RNA modifications have been viewed as new posttranscriptional regulator in modulating gene expression as well as in a broad range of physiological processes. N¹-methyladenosine (m¹A) is one of the most prevalent modifications existing in multiple types of RNAs. In-depth investigation of the functions of m¹A requires the site-specific assessment of m¹A stoichiometry in RNA. Herein, we established a demethylase-assisted method (DA-m¹A) for the site-specific detection and quantification of m¹A in RNA. N1-methyl group in m¹A could result in the stalling of reverse transcription at m¹A site, thus producing the truncated cDNA. E. coli AlkB is a demethylase that can demethylate m¹A to produce adenine in RNA, thus generating full-length cDNA from AlkB-treated RNA. Evaluation of the produced amounts of full-length cDNA by quantitative real-time PCR can achieve the site-specific detection and quantification of m¹A in RNA. With the DA-m¹A method, we examined and successfully confirmed the previously well-characterized m¹A sites in various types of RNAs with low false positive rate. In addition, we found that the level of m¹A was significantly decreased at the bromodomain containing 2 (BRD2) mRNA position 1674 and CST telomere replication complex component 1 (CTC1) mRNA position 5643 in human hepatocellular carcinoma tissues. The results suggest that these two m¹A sites in mRNA may be involved in liver tumorigenesis. Taken together, the DA-m¹A method is simple and enables the rapid, cost-effective, and site-specific detection and quantification of m¹A in RNA, which provides a valuable tool to decipher the functions of m¹A in human diseases.
- N¹-methyladenosine,
- RNA modification,
- AlkB demethylase,
- RNA epigenetics,
- Quantitative real-time PCR,
- Hepatocellular carcinoma
1. [1]
  D. Wiener, S. Schwartz, Nat. Rev. Genet. 22 (2021) 119–131.
2. [2]
  S. Moshitch-Moshkovitz, D. Dominissini, G. Rechavi, Cell 185 (2022) 764–776. doi: 10.1016/j.cell.2022.02.007
3. [3]
  Y. Zhao, X. Zuo, Q. Li, et al., Sci. China Chem. 64 (2021) 171–203. doi: 10.1007/s11426-020-9864-7
4. [4]
  P. Boccaletto, F. Stefaniak, A. Ray, et al., Nucleic Acids Res. 50 (2022) D231–D235. doi: 10.1093/nar/gkab1083
5. [5]
  M.Y. Chen, Z. Gui, K.K. Chen, et al., Chin. Chem. Lett. 33 (2022) 2086–2090. doi: 10.1016/j.cclet.2021.08.094
6. [6]
  X.J. You, S. Zhang, J.J. Chen, et al., Nucleic Acids Res. 50 (2022) 9858–9872. doi: 10.1093/nar/gkac770
7. [7]
  Y. Dai, C.B. Qi, Y. Feng, et al., Anal. Chem. 93 (2021) 6938–6946. doi: 10.1021/acs.analchem.0c04630
8. [8]
  M.Y. Cheng, X.J. You, J.H. Ding, et al., Chem. Sci. 12 (2021) 8149–8156. doi: 10.1039/D1SC01972D
9. [9]
  Y. Motorin, M. Helm, Wiley Interdiscip. Rev. RNA 13 (2022) e1691. doi: 10.1002/wrna.1691
10. [10]
  J.H. Ding, M.Y. Chen, N.B. Xie, et al., Biosens. Bioelectron. 219 (2023) 114821. doi: 10.1016/j.bios.2022.114821
11. [11]
  M.Y. Chen, C.B. Qi, X.M. Tang, et al., Chin. Chem. Lett. 33 (2022) 3772–3776. doi: 10.1016/j.cclet.2021.12.008
12. [12]
  T.R. Fischer, L. Meidner, M. Schwickert, et al., Nucleic Acids Res. 50 (2022) 4216–4245. doi: 10.1093/nar/gkac224
13. [13]
  F. Nie, P. Feng, X. Song, et al., Database 2020 (2020) baaa049. doi: 10.1093/database/baaa049
14. [14]
  H.L. Huang, H.Y. Weng, X.L. Deng, J.J. Chen, Annu. Rev. Cancer Biol. 4 (2020) 221–240. doi: 10.1146/annurev-cancerbio-030419-033357
15. [15]
  K. Chen, B.S. Zhao, C. He, Cell Chem. Biol. 23 (2016) 74–85. doi: 10.1016/j.chembiol.2015.11.007
16. [16]
  H. Shi, P. Chai, R. Jia, X. Fan, Mol. Cancer 19 (2020) 78. doi: 10.1186/s12943-020-01194-6
17. [17]
  M. Frye, B.T. Harada, M. Behm, C. He, Science 361 (2018) 1346–1349. doi: 10.1126/science.aau1646
18. [18]
  Y.Y. Chen, Z. Gui, D. Hu, et al., Chin. Chem. Lett. 34 (2023) 108522.
19. [19]
  J.J. Chen, X.J. You, L. Li, et al., Anal. Chem. 94 (2022) 8740–8747. doi: 10.1021/acs.analchem.2c01226
20. [20]
  S. Murakami, S.R. Jaffrey, Mol. Cell. 82 (2022) 2236–2251. doi: 10.1016/j.molcel.2022.05.029
21. [21]
  J. Li, H. Zhang, H. Wang, Comput. Struct. Biotechnol. J. 20 (2022) 6578–6585. doi: 10.1016/j.csbj.2022.11.045
22. [22]
  C. Zhang, G. Jia, Genom. Proteom. Bioinform. 16 (2018) 155–161. doi: 10.1016/j.gpb.2018.03.003
23. [23]
  X. Xiong, X. Li, C. Yi, Curr. Opin. Chem. Biol. 45 (2018) 179–186. doi: 10.1016/j.cbpa.2018.06.017
24. [24]
  S. Ozanick, A. Krecic, J. Andersland, J.T. Anderson, RNA 11 (2005) 1281–1290. doi: 10.1261/rna.5040605
25. [25]
  E. Vilardo, C. Nachbagauer, A. Buzet, et al., Nucleic Acids Res. 40 (2012) 11583–11593. doi: 10.1093/nar/gks910
26. [26]
  T. Chujo, T. Suzuki, RNA 18 (2012) 2269–2276. doi: 10.1261/rna.035600.112
27. [27]
  T. Waku, Y. Nakajima, W. Yokoyama, et al., J. Cell Sci. 129 (2016) 2382–2393.
28. [28]
  X. Li, X. Xiong, K. Wang, et al., Nat. Chem. Biol. 12 (2016) 311–316. doi: 10.1038/nchembio.2040
29. [29]
  D. Dominissini, S. Nachtergaele, S. Moshitch-Moshkovitz, et al., Nature 530 (2016) 441–446. doi: 10.1038/nature16998
30. [30]
  Y. Zhang, L. Lu, X. Li, Exp. Mol. Med. 54 (2022) 1601–1616. doi: 10.1038/s12276-022-00821-0
31. [31]
  H. Liu, Y. Wang, X. Zhou, RSC Chem. Biol. 3 (2022) 994–1007. doi: 10.1039/D2CB00087C
32. [32]
  A.S. Felix, A.L. Quillin, S. Mousavi, J.M. Heemstra, Acc. Chem. Res. 55 (2022) 2271–2279. doi: 10.1021/acs.accounts.2c00287
33. [33]
  D. Bartee, S. Thalalla Gamage, C.N. Link, J.L. Meier, Chem. Soc. Rev. 50 (2021) 9482–9502. doi: 10.1039/D1CS00214G
34. [34]
  J.D. Alfonzo, J.A. Brown, P.H. Byers, et al., Nat. Genet. 53 (2021) 1113–1116. doi: 10.1038/s41588-021-00903-1
35. [35]
  X.J. You, L. Li, T.T. Ji, et al., Chin. Chem. Lett. 34 (2023) 107181. doi: 10.1016/j.cclet.2022.01.074
36. [36]
  Q. Wang, J.H. Ding, J. Xiong, et al., Chin. Chem. Lett. 32 (2021) 3426–3430. doi: 10.1016/j.cclet.2021.05.020
37. [37]
  Q.F. Zhang, H.M. Xiao, J.T. Zhan, B.F. Yuan, Y.Q. Feng, Chin. Chem. Lett. 33 (2022) 4746–4749. doi: 10.1016/j.cclet.2022.01.004
38. [38]
  B. Chen, B.F. Yuan, Y.Q. Feng, Anal. Chem. 91 (2019) 743–756. doi: 10.1021/acs.analchem.8b04078
39. [39]
  X.M. Tang, T.T. Ye, X.J. You, et al., Chin. Chem. Lett. 34 (2023) 107531. doi: 10.1016/j.cclet.2022.05.045
40. [40]
  W.B. Tao, N.B. Xie, Q.Y. Cheng, Y.Q. Feng, B.F. Yuan, Chin. Chem. Lett. 34 (2023) 108243. doi: 10.1016/j.cclet.2023.108243
41. [41]
  S.C. Trewick, T.F. Henshaw, R.P. Hausinger, T. Lindahl, B. Sedgwick, Nature 419 (2002) 174–178. doi: 10.1038/nature00908
42. [42]
  P.O. Falnes, R.F. Johansen, E. Seeberg, Nature 419 (2002) 178–182. doi: 10.1038/nature01048
43. [43]
  M. Safra, A. Sas-Chen, R. Nir, et al., Nature 551 (2017) 251–255. doi: 10.1038/nature24456
44. [44]
  H. Zhou, S. Rauch, Q. Dai, et al., Nat. Methods 16 (2019) 1281–1288. doi: 10.1038/s41592-019-0550-4
45. [45]
  X. Li, X. Xiong, M. Zhang, et al., Mol. Cell. 68 (2017) 993–1005.e9. doi: 10.1016/j.molcel.2017.10.019
46. [46]
  J.H. Ding, C.J. Ma, M.Y. Chen, et al., Anal. Chem. 92 (2020) 2612–2619. doi: 10.1021/acs.analchem.9b04454
47. [47]
  I. Barbieri, T. Kouzarides, Nat. Rev. Cancer 20 (2020) 303–322. doi: 10.1038/s41568-020-0253-2
48. [48]
  J. Xiong, K.K. Chen, N.B. Xie, et al., Anal. Chem. 94 (2022) 15489–15498. doi: 10.1021/acs.analchem.2c03808
49. [49]
  N.B. Xie, M. Wang, T.T. Ji, et al., Chem. Sci. 13 (2022) 7046–7056. doi: 10.1039/D2SC01052F
50. [50]
  Y. Wang, J. Wang, X. Li, et al., Nat. Commun. 12 (2021) 6314. doi: 10.1038/s41467-021-26718-6
1. [1]
  Tian Feng , Yun-Ling Gao , Di Hu , Ke-Yu Yuan , Shu-Yi Gu , Yao-Hua Gu , Si-Yu Yu , Jun Xiong , Yu-Qi Feng , Jie Wang , Bi-Feng Yuan . Chronic sleep deprivation induces alterations in DNA and RNA modifications by liquid chromatography-mass spectrometry analysis. Chinese Chemical Letters, 2024, 35(8): 109259-. doi: 10.1016/j.cclet.2023.109259
2. [2]
  Keqiang Shi , Xiujuan Hong , Dongyan Xu , Tao Pan , Huiwen Wang , Hongru Feng , Cheng Guo , Yuanjiang Pan . Analysis of RNA modifications in peripheral white blood cells from breast cancer patients by mass spectrometry. Chinese Chemical Letters, 2025, 36(3): 110079-. doi: 10.1016/j.cclet.2024.110079
3. [3]
  Zhiyu Yu , Xiang Luo , Cheng Zhang , Xin Lu , Xiaohui Li , Pan Liao , Zhongqiu Liu , Rong Zhang , Shengtao Wang , Zhiqiang Yu , Guochao Liao . Mitochondria-targeted carrier-free nanoparticles based on dihydroartemisinin against hepatocellular carcinoma. Chinese Chemical Letters, 2024, 35(10): 109519-. doi: 10.1016/j.cclet.2024.109519
4. [4]
  Qiuye Wang , Yabing Sun , Liangxue Lai , Haijing Cui , Yonglong Ye , Ming Yang , Weihao Zhu , Bo Yuan , Quanliang Mao , Wenzhi Ren , Aiguo Wu . MMP-9-responsive probe for fluorescence-magnetic resonance dual-mode imaging of hepatocellular carcinoma models with different metastatic capacities. Chinese Chemical Letters, 2025, 36(4): 110212-. doi: 10.1016/j.cclet.2024.110212
5. [5]
  Cheng Guo , Xiaoxiao Zhang , Xiujuan Hong , Yiqiu Hu , Lingna Mao , Kezhi Jiang . Graphene as adsorbent for highly efficient extraction of modified nucleosides in urine prior to liquid chromatography-tandem mass spectrometry analysis. Chinese Chemical Letters, 2024, 35(4): 108867-. doi: 10.1016/j.cclet.2023.108867
6. [6]
  Jinyu Guo , Yandai Lin , Shaohua He , Yueqing Chen , Fenglu Li , Renjie Ruan , Gaoxing Pan , Hexin Nan , Jibin Song , Jin Zhang . Utilizing dual-responsive iridium(Ⅲ) complex for hepatocellular carcinoma: Integrating photoacoustic imaging with chemotherapy and photodynamic therapy. Chinese Chemical Letters, 2024, 35(9): 109537-. doi: 10.1016/j.cclet.2024.109537
7. [7]
  Yan Liu , Yang Wang , Jiayi Zhu , Xuxian Su , Xudong Lin , Liang Xu , Xiwen Xing . Employing pH-responsive RNA triplex to control CRISPR/Cas9-mediated gene manipulation in mammalian cells. Chinese Chemical Letters, 2024, 35(9): 109427-. doi: 10.1016/j.cclet.2023.109427
8. [8]
  Ziqin Li , Kai Hao , Longwei Xiang , Huayu Tian . Cationic covalent organic framework nanocarriers integrating both efficient gene silencing and real-time gene detection. Chinese Chemical Letters, 2025, 36(4): 109943-. doi: 10.1016/j.cclet.2024.109943
9. [9]
  Jia-Qi Feng , Xiang Tian , Rui-Ge Cao , Yong-Xiu Li , Wen-Long Liu , Rong Huang , Si-Yong Qin , Ai-Qing Zhang , Yin-Jia Cheng . An AIE-based theranostic nanoplatform for enhanced colorectal cancer therapy: Real-time tumor-tracking and chemical-enhanced photodynamic therapy. Chinese Chemical Letters, 2024, 35(12): 109657-. doi: 10.1016/j.cclet.2024.109657
10. [10]
  Ke Gong , Jinghan Liao , Jiangtao Lin , Quan Wang , Zhihua Wu , Liting Wang , Jiali Zhang , Yi Dong , Yourong Duan , Jianhua Chen . Mitochondria-targeted nanoparticles overcome chemoresistance via downregulating BACH1/CD47 axis in ovarian carcinoma. Chinese Chemical Letters, 2024, 35(5): 108888-. doi: 10.1016/j.cclet.2023.108888
11. [11]
  Lijun Mao , Shuo Li , Xin Zhang , Zhan-Ting Li , Da Ma . Cucurbit[n]uril-based nanostructure construction and modification. Chinese Chemical Letters, 2024, 35(8): 109363-. doi: 10.1016/j.cclet.2023.109363
12. [12]
  Jianhui Yin , Wenjing Huang , Changyong Guo , Chao Liu , Fei Gao , Honggang Hu . Tryptophan-specific peptide modification through metal-free photoinduced N-H alkylation employing N-aryl glycines. Chinese Chemical Letters, 2024, 35(6): 109244-. doi: 10.1016/j.cclet.2023.109244
13. [13]
  Chaochao Jin , Kai Li , Jiongpei Zhang , Zhihua Wang , Jiajing Tan . N,O-Bidentated difluoroboron complexes based on pyridine-ester enolates: Facile synthesis, post-complexation modification, optical properties, and applications. Chinese Chemical Letters, 2024, 35(9): 109532-. doi: 10.1016/j.cclet.2024.109532
14. [14]
  Quanyou Guo , Yue Yang , Tingting Hu , Hongqi Chu , Lijun Liao , Xuepeng Wang , Zhenzi Li , Liping Guo , Wei Zhou . Regulating local electron transfer environment of covalent triazine frameworks through F, N co-modification towards optimized oxygen reduction reaction. Chinese Chemical Letters, 2025, 36(1): 110235-. doi: 10.1016/j.cclet.2024.110235
15. [15]
  Runze Liu , Yankai Bian , Weili Dai . Qualitative and quantitative analysis of Brønsted and Lewis acid sites in zeolites: A combined probe-assisted 1H MAS NMR and NH3-TPD investigation. Chinese Journal of Structural Chemistry, 2024, 43(4): 100250-100250. doi: 10.1016/j.cjsc.2024.100250
16. [16]
  Wenbi Wu , Yinchu Dong , Haofan Liu , Xuebing Jiang , Li Li , Yi Zhang , Maling Gou . Modification of plasma protein for bioprinting via photopolymerization. Chinese Chemical Letters, 2024, 35(8): 109260-. doi: 10.1016/j.cclet.2023.109260
17. [17]
  Chenghe Yang , Yi Lü , Rui Liu . The Rise to Fame of Digital PCR. University Chemistry, 2025, 40(4): 340-345. doi: 10.12461/PKU.DXHX202406111
18. [18]
  Jiechen Liu , Xiaoguang Li , Ruiyang Xia , Yuqi Wang , Fenghe Zhang , Yongzhi Pang , Qing Li . Efficient suppression of oral squamous cell carcinoma through spatial dimension conversion drug delivery systems-enabled immunomodulatory-photodynamic therapy. Chinese Chemical Letters, 2024, 35(12): 109619-. doi: 10.1016/j.cclet.2024.109619
19. [19]
  Hai-Yang Song , Jun Jiang , Yu-Hang Song , Min-Hang Zhou , Chao Wu , Xiang Chen , Wei-Min He . Supporting-electrolyte-free electrochemical [2 + 2 + 1] annulation of benzo[d]isothiazole 1,1-dioxides, N-arylglycines and paraformaldehyde. Chinese Chemical Letters, 2024, 35(6): 109246-. doi: 10.1016/j.cclet.2023.109246
20. [20]
  Liangfeng Yang , Liang Zeng , Yanping Zhu , Qiuan Wang , Jinheng Li . Copper-catalyzed photoredox 1,4-amidocyanation of 1,3-enynes with N-amidopyridin-1-ium salts and TMSCN: Facile access to α-amido allenyl nitriles. Chinese Chemical Letters, 2024, 35(11): 109685-. doi: 10.1016/j.cclet.2024.109685

Get Citation

PDF

XML

Metrics

PDF Downloads(1)
Abstract views(279)
HTML views(2)

通讯作者: 陈斌, bchen63@163.com

1.
沈阳化工大学材料科学与工程学院沈阳 110142

Demethylase-assisted site-specific detection of N1-methyladenosine in RNA

Metrics

通讯作者: 陈斌, bchen63@163.com

Demethylase-assisted site-specific detection of N¹-methyladenosine in RNA