5-azacytidine enhances the aluminum tolerance of grapes by reducing the DNA methylation level

5-Azacitidine (5-Azac) is a base analog that replaces methylated cytosine in DNA, thereby reducing the methylation level of the genome. However, the mechanism by which exogenous 5-Azac induces aluminum accumulation in grapes through methylation remains unclear.

Spraying 'Shuijing' grape seedlings with a 5-Azac treatment solution under aluminum (Al) stress, agronomic traits of grapes were observed, and the activities of SOD, POD, and MDA, as well as the content of H₂O₂ and Al, and the rate of oxygen free radical production, were measured. Furthermore, whole genome bisulfite sequencing (WGBS) was employed to generate high-coverage cytosine methylation profiles of the grape genome.

After treatment with 5-Azac, 'Shuijing' grape seedlings under Al stress showed decreased methylation rates across all chromosomes and a reduced overall DNA methylation level. Specifically, methylation levels at CG, CHG, and CHH sites in promoters increased, enabling transcription factors to activate these promoters and induce upregulation of hypo-differentially methylated regions (DMRs) in the genome, thus mitigating Al stress. Gene Ontology enrichment analysis of DMRs demonstrated that 5-Azac treatment effectively reduced the Al toxicity in grape plants under Al stress by upregulating related genes and effectively reducing the Al stress-induced DNA methylation level of the grape genome.

Treatment with 5-Azac results in a decrease in the degree of DNA methylation in the grape genome. Analyzing the methylation characteristics has provided insights into the molecular mechanism underlying the response of grapes to Al stress.