Global profiling of 6mA sites at single-nucleotide resolution in the genome of Arabidopsis thaliana using PacBio sequencing Nextomics program

In April 2018, Nextomics joins hands with research group of Xiaofeng Gu, biotechnology research institute, Chinese academy of agricultural sciences, and research group of Hao Yu, department of biological sciences and Temasek life sciences laboratory, National University of Singapore, to publish a paper titled as “DNA N⁶-Adenine Methylation in Arabidopsis thaliana” in the journal of Development Cell. This research reported that global profiling of 6mA sites at single-nucleotide resolution in the genome of A. thaliana at different developmental stages and ecotypes using single-molecule real-time sequencing.

This is the first research article of distributing eukaryote methylation mapped by PacBio SMRT.

Highlights

• DNA methylation on N⁶-adenine (6mA) widely occurs in the Arabidopsisgenome;
• 6mA is more enriched on gene bodies than intergenic regions;
• 6mA is a dynamic DNA modification during Arabidopsisdevelopment;
• 6mA is associated with actively expressed genes in

Results

6mA widely occurs in the Arabidopsis genome

Dot blot analysis of gDNA extracted from 3- to 21-day-old Col wild-type plants revealed a gradual increase in the 6mA signal intensity from vegetative to reproductive stage (Fig.1).

Fig. 1 6mA Occurs in Arabidopsis Genomic DNA

Genome-Wide Mapping of 6mA in Arabidopsis sequenced by PacBio SMRT

Compare the dynamic and distribution of 6mA in D9 and D21, and combined with the transcriptome data researchers revealed the function of 6mA during the development of Arabidopsis (Fig.2).

Fig.2 Circos plots of 6mA DNA methylation profiles of 9- and 21-day-old Col wild-type Arabidopsis.

6mA is more enriched on gene bodies than intergenic regions

6mA distribution in Col genomic regions divided into gene bodies, promoters, 5’intergenic, 3’intergenic, and other intergenic regions. The results showed that 32% of 6mA sites were located within gene bodies, and protein-coding genes contained more than half of the 6mA sites in all methylated genes (Fig. 3).

Fig. 3 Distribution Pattern of 6mA Methylation in Genomic DNA of 9-Day-Old Col Plants

6mA is a dynamic DNA modification during Arabidopsis development

Genome-wide mapping of 6mA in gDNA from 21-day-old Col wild-type plants at the reproductive stage by SMRT sequencing identified 184,633 6mA sites, among which 9,909 sites were overlapped with those in 9-day-old plants (Fig. 4), which implied that 6mA DNA methylation positively correlates with the transition from vegetative to reproductive growth.

Fig.4 Venn diagram comparison of the numbers of 6mA sites in D9 and D21.

6mA associated with actively expressed genes in Arabidopsis

Analysis of 6mA methylomes and RNA sequencing data demonstrates that 6mA frequency positively correlates with the gene expression level and the transition from vegetative to reproductive growth in Arabidopsis (Fig. 5).

Fig. 5 Correlation between 6mA and Gene Expression in Col Plants.

This research uncover 6mA as a DNA mark associated with actively expressed genes in Arabidopsis, suggesting that 6mA serves as a hitherto unknown epigenetic mark in land plants. Nextomics took part in the program and provided technical support.

Reference

Liang et al., DNA N6-Adenine Methylation in Arabidopsis thaliana, Developmental Cell (2018), https://doi.org/10.1016/j.devcel.2018.03.012

Other eukaryote methylation published papers

[1]Greer, E.L. et al. DNA methylation on N6-adenine in C. elegans. Cell 161, 868–878 (2015).

[2]Wu, T.P. et al. DNA methylation on N(6)-adenine in mammalian embryonic stem cells. Nature 532, 329–333 (2016).

[3]Mondo, S.J. et al. Widespread adenine N6-methylation of active genes in fungi. Nature Genetics (2017).