Cytosine DNA methylation is an epigenetic regulatory system used by plants to control gene expression. Methylation pattern always changes after abiotic stresses, pathogens and pest infections or after a treatment with salicylic acid (SA). The latter is a key player in plant development and defense against insect herbivores, pathogens, and abiotic stresses. The roles of SA on the methylation patterns and the plant development were performed in 4 pearl millet (Pennisetum glaucum) varieties. Seedlings of 4 early-flowering photosensitive genotypes (PMS3, PMI8, PMG, and PMT2) were grown on MS medium supplemented with null or different doses of SA. Root growth was used as a parameter to evaluate the effects of SA at early stage development. DNA from these seedlings was extracted and Methylation-Sensitive Amplified Polymorphism (MSAP) was measured to assess the effects of SA on methylome. The methylation analysis revealed that SA treatment decreased the methylation, while inhibiting the root growth for all varieties tested, except in PMG at 0.5 mM, indicating a dose and a genotype response-dependence. The methylation level was positively correlated with the root growth. This suggests that SA influences both the methylome by demethylation activities and the root growth by interfering with the root development-responsive genes. The demethylation process, induced by the REPRESSOR OF SILCENCING 1 (ROS1) may activate R genes, or GH3.5 and downregulate the hormonal pathway under root development. These findings showed the pearl millet metabolism prioritized and promoted the defense pathways over vegetative development during stress.

The cytosine DNA methylation and demethylation have a role in regulating plant responses to the environment by affecting the promoter regions of most plant defense-related genes through the CpG islands or the CCGG motifs. Salicylic acid, a defense and signaling plant hormone, is seen playing crucial role in the variation of the methylome. In this study, the effects of salicylic acid and feeding of the millet headminer (Heliocheilus albipunctella de Joannis) on pearl millet DNA methylome changes were evaluated through MSAP epigenotyping during panicle development. The results showed that millet headminer feeding increased the level of genomic methylation while application of salicylic acid caused DNA demethylation occurring mostly at external cytosine and accompanied by a decrease of the number of larvae per panicle. This suggests that hemimethylation (external cytosine methylation) has key role in regulating defense responses and conferring tolerance to pearl millet through salicylic acid application.

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