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BACKGROUND AND AIMS: DNA methylation plays a crucial role in plant stress response, particularly under abiotic stress. Suaeda aralocaspica (S. aralocaspica), a halophyte with C4 photosynthetic pathway within a single polarized cell, exhibits unique adaptations to environmental challenges. This study investigates cytosine methylation patterns in S. aralocaspica under dark (D), light recovery (LR), and NaCl stress (100 and 500 mM, N100 and N500), focusing on the methylation-mediated regulation of stress-responsive genes, including different phosphoenolpyruvate carboxylase genes (SaPEPCs). METHODS: The whole-genome bisulfite sequencing was used to investigate DNA methylation under different stress treatments. Gene expression and Western blot analyses in photosynthesis and salt response pathways were conducted using a DNA methylation inhibitor (5-azacytidine, 5-azaC) and a methylating agent (methyl trifluoromethanesulfonate, MTFMS) to assess the functional link between methylation and gene regulation. KEY RESULTS: Global methylation levels in S. aralocaspica decreased under stress, with non-CG methylation showing the most significant changes. The majority of 81854 and 14653 differentially methylated regions (DMRs) under N500 and darkness, respectively, were hypo-methylated. Dynamic DNA methylation directly regulates the expression of genes involved in both photosynthetic and salt stress responses. In most cases, mRNA expression of these genes increased when their promoters were hypo-methylated. The functional significance of these methylation patterns was further confirmed through DNA methylation inhibitor (5-azaC) or DNA methylating agent (MTFMS). These treatments revealed that hypo-methylation may enhance the gene expression, as observed in key genes such as SaPEPC1, SaPEPC2, PPDK, CPK3, CPK4. Between the two SaPEPC genes, SaPEPC2 showed hypo-methylated DMRs specifically under N500 treatment, by contrast, SaPEPC1 exhibited a hypo-DMR under darkness. Combined with our previous study, our results suggest a functional divergence among these paralogs, and SaPEPC1 may be the key gene in carbon fixation. CONCLUSIONS: This study revealed a stress-induced genome-wide DNA hypo-methylation in S. aralocaspica. SaPEPC1 and SaPEPC2 exhibited distinct methylation responses to darkness and salt stress, respectively. Through methylation alteration experiments, we established the associations between DNA methylation and relevant gene expression, which should improve our understanding of epigenetic regulation in halophyte stress adaptation.
Yang et al. (Tue,) studied this question.