Integrated Transcriptome Analysis and Single-Base Resolution Methylomes of Watermelon (Citrullus lanatus) Reveal Epigenome Modifications in Response to Osmotic Stress.

Fangming Zhu,Manwen Yan,Mingyan Li,Fei Qiao,Xuefei Jiang

doi:10.3389/fpls.2021.769712

Abstract

DNA methylation plays an important role against adverse environment by reshaping transcriptional profile in plants. To better understand the molecular mechanisms of watermelon response to osmotic stress, the suspension cultured watermelon cells were treated with 100mM mannitol, and then a methylated cytosines map was generated by whole genome bisulfite sequencing (WGBS). Combined with transcriptome sequencing, the effects of osmotic stress on differentially methylated expressed genes (DMEGs) were assessed. It was found that genes related to plant hormone synthesis, signal transduction, osmoregulatory substance-related and reactive oxygen species scavenging-related enzyme could rapidly respond to osmotic stress. The overall methylation level of watermelon decreased after osmotic stress treatment, and demethylation occurred in CG, CHG, and CHH contexts. Moreover, differentially methylated expressed genes (DMEGs) were significantly enriched in RNA transport, starch and sucrose metabolism, plant hormone signal transduction and biosynthesis of secondary metabolites, especially in biosynthesis of osmolytes synthase genes. Interestingly, demethylation of a key enzyme gene Cla014489 in biosynthesis of inositol upregulated its expression and promoted accumulation of inositol, which could alleviate the inhibition of cell growth caused by osmotic stress. Meanwhile, a recombinant plasmid pET28a-Cla014489 was constructed and transferred into Escherichia coli BL21 for prokaryotic expression and the expression of ClMIPS protein could improve the tolerance of E. coli to osmotic stress. The effect of methylation level on the expression properties of inositol and its related genes was further confirmed by application of DNA methylation inhibitor 5-azacytidine. These results provide a preliminary insight into the altered methylation levels of watermelon cells in response to osmotic stress and suggest a new mechanism that how watermelon cells adapt to osmotic stress.

Highlights

With the continuous exploration of molecular mechanisms, it has been discovered that some biological phenomena do not exactly conform to genetic inheritance laws
A lower number of differentially expressed genes (DEGs) indicates that the gene expression patterns of watermelon suspension cells at 2 h and 4 h were similar after osmotic stress
The results showed that the expression levels of most genes related to cell growth changed dynamically, with 13 genes related to Auxin, 4 genes related to Cytokinin and 3 genes related to GA undergoing significant up-regulation of expression (Figure 2), suggesting that cell proliferation may be accelerating in response to adversity in the early stages

Summary

Introduction

With the continuous exploration of molecular mechanisms, it has been discovered that some biological phenomena do not exactly conform to genetic inheritance laws. DNA methylation patterns and levels are diverse across species and can typically occur at symmetric CG and CHG sites and asymmetric CHH sites where H represents A, C, or T (Cao and Jacobsen, 2002; Henderson and Jacobsen, 2007). They can all be established by Domains Rearranged Methyltrans-ferase 2 (DRM2) via the RNA-directed DNA methylation pathway and maintained by different methylation transferases (Law and Jacobsen, 2010; Stroud et al, 2014). It was found that most DNA methylation modified genes in plants undergo transcriptional changes in response to adversity stress, suggesting that complex DNA methylation patterns are directly related to stress response and affect gene expression (Lu et al, 2017; Xu et al, 2018a)

Methods

Results

Conclusion