Bisulfite-free, Base-resolution Analysis of 5-formylcytosine at the Genome Scale

Overview

Journal Nat Methods

Specialties Biomedical Engineering
Pathology

Date 2015 Sep 8

PMID 26344045

Citations 46

Authors

Bo Xia

Dali Han

Xingyu Lu

Zhaozhu Sun

Ankun Zhou

Qiangzong Yin

Hu Zeng

Menghao Liu

Xiang Jiang

Wei Xie

Chuan He

Chengqi Yi

Affiliations

Soon will be listed here.

Abstract

Active DNA demethylation in mammals involves oxidation of 5-methylcytosine (5mC) to 5-hydroxymethylcytosine (5hmC), 5-formylcytosine (5fC) and 5-carboxylcytosine (5caC). However, genome-wide detection of 5fC at single-base resolution remains challenging. Here we present fC-CET, a bisulfite-free method for whole-genome analysis of 5fC based on selective chemical labeling of 5fC and subsequent C-to-T transition during PCR. Base-resolution 5fC maps showed limited overlap with 5hmC, with 5fC-marked regions more active than 5hmC-marked ones.

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References

Rivera C, Ren B . Mapping human epigenomes. Cell. 2013; 155(1):39-55. PMC: 3838898. DOI: 10.1016/j.cell.2013.09.011. View

Song C, He C . Potential functional roles of DNA demethylation intermediates. Trends Biochem Sci. 2013; 38(10):480-4. PMC: 4013277. DOI: 10.1016/j.tibs.2013.07.003. View

Iurlaro M, Ficz G, Oxley D, Raiber E, Bachman M, Booth M . A screen for hydroxymethylcytosine and formylcytosine binding proteins suggests functions in transcription and chromatin regulation. Genome Biol. 2013; 14(10):R119. PMC: 4014808. DOI: 10.1186/gb-2013-14-10-r119. View

Booth M, Marsico G, Bachman M, Beraldi D, Balasubramanian S . Quantitative sequencing of 5-formylcytosine in DNA at single-base resolution. Nat Chem. 2014; 6(5):435-40. PMC: 4188980. DOI: 10.1038/nchem.1893. View

Raiber E, Beraldi D, Ficz G, Burgess H, Branco M, Murat P . Genome-wide distribution of 5-formylcytosine in embryonic stem cells is associated with transcription and depends on thymine DNA glycosylase. Genome Biol. 2012; 13(8):R69. PMC: 3491369. DOI: 10.1186/gb-2012-13-8-r69. View

Hon G, Song C, Du T, Jin F, Selvaraj S, Lee A . 5mC oxidation by Tet2 modulates enhancer activity and timing of transcriptome reprogramming during differentiation. Mol Cell. 2014; 56(2):286-297. PMC: 4319980. DOI: 10.1016/j.molcel.2014.08.026. View

Bachman M, Uribe-Lewis S, Yang X, Williams M, Murrell A, Balasubramanian S . 5-Hydroxymethylcytosine is a predominantly stable DNA modification. Nat Chem. 2014; 6(12):1049-55. PMC: 4382525. DOI: 10.1038/nchem.2064. View

Wu H, Wu X, Shen L, Zhang Y . Single-base resolution analysis of active DNA demethylation using methylase-assisted bisulfite sequencing. Nat Biotechnol. 2014; 32(12):1231-40. PMC: 4269366. DOI: 10.1038/nbt.3073. View

Peng X, Wu J, Brunmeir R, Kim S, Zhang Q, Ding C . TELP, a sensitive and versatile library construction method for next-generation sequencing. Nucleic Acids Res. 2014; 43(6):e35. PMC: 4381050. DOI: 10.1093/nar/gku818. View

10.

Wang D, Hara R, Singh G, Sancar A, Lippard S . Nucleotide excision repair from site-specifically platinum-modified nucleosomes. Biochemistry. 2003; 42(22):6747-53. DOI: 10.1021/bi034264k. View

11.

Zhang Y, Liu T, Meyer C, Eeckhoute J, Johnson D, Bernstein B . Model-based analysis of ChIP-Seq (MACS). Genome Biol. 2008; 9(9):R137. PMC: 2592715. DOI: 10.1186/gb-2008-9-9-r137. View

12.

Tahiliani M, Koh K, Shen Y, Pastor W, Bandukwala H, Brudno Y . Conversion of 5-methylcytosine to 5-hydroxymethylcytosine in mammalian DNA by MLL partner TET1. Science. 2009; 324(5929):930-5. PMC: 2715015. DOI: 10.1126/science.1170116. View

13.

Obeid S, Blatter N, Kranaster R, Schnur A, Diederichs K, Welte W . Replication through an abasic DNA lesion: structural basis for adenine selectivity. EMBO J. 2010; 29(10):1738-47. PMC: 2876968. DOI: 10.1038/emboj.2010.64. View

14.

Heinz S, Benner C, Spann N, Bertolino E, Lin Y, Laslo P . Simple combinations of lineage-determining transcription factors prime cis-regulatory elements required for macrophage and B cell identities. Mol Cell. 2010; 38(4):576-89. PMC: 2898526. DOI: 10.1016/j.molcel.2010.05.004. View

15.

Song C, Szulwach K, Fu Y, Dai Q, Yi C, Li X . Selective chemical labeling reveals the genome-wide distribution of 5-hydroxymethylcytosine. Nat Biotechnol. 2010; 29(1):68-72. PMC: 3107705. DOI: 10.1038/nbt.1732. View

16.

Williams K, Christensen J, Pedersen M, Johansen J, Cloos P, Rappsilber J . TET1 and hydroxymethylcytosine in transcription and DNA methylation fidelity. Nature. 2011; 473(7347):343-8. PMC: 3408592. DOI: 10.1038/nature10066. View

17.

Krueger F, Andrews S . Bismark: a flexible aligner and methylation caller for Bisulfite-Seq applications. Bioinformatics. 2011; 27(11):1571-2. PMC: 3102221. DOI: 10.1093/bioinformatics/btr167. View

18.

Pfaffeneder T, Hackner B, Truss M, Munzel M, Muller M, Deiml C . The discovery of 5-formylcytosine in embryonic stem cell DNA. Angew Chem Int Ed Engl. 2011; 50(31):7008-12. DOI: 10.1002/anie.201103899. View

19.

Ito S, Shen L, Dai Q, Wu S, Collins L, Swenberg J . Tet proteins can convert 5-methylcytosine to 5-formylcytosine and 5-carboxylcytosine. Science. 2011; 333(6047):1300-3. PMC: 3495246. DOI: 10.1126/science.1210597. View

20.

He Y, Li B, Li Z, Liu P, Wang Y, Tang Q . Tet-mediated formation of 5-carboxylcytosine and its excision by TDG in mammalian DNA. Science. 2011; 333(6047):1303-7. PMC: 3462231. DOI: 10.1126/science.1210944. View