Overexpression of NtROS2a Gene Encoding Cytosine DNA Demethylation Enhanced Drought Tolerance in Transgenic Plants

DNA methylation is a common and rapid evolving epigenetic mechanism among higher eukaryotic organisms including cytosine residues in CG and CNG sequences. NtROS2a gene isolated from tobacco plants (Nicotiana tabacum cv Xanthi nc) contains DNA glycosylase domain with significant similarities to Arabidopsis ROS1 and DME at the C-terminus. Purified NtROS2a protein gene is 5,879 bp long and is designated as Nicotiana tabacum Repressor Of Silencing (NtROS: accession numbers; AB281588). qRT-PCR analysis initially revealed the adaptive response of the lines overexpressing the gene in tobacco to various abiotic stresses. To study the transcript levels of epigenetically regulated genes in tobacco, we conducted microarray analysis with Agilent Tobacco 4 x 44K oligo chip by using overexpression line OX1, RNAi plant line RNAi 13, and wild type plant WT. It has been reported that salt- and H₂O₂- stress tolerance of these transgenic lines are enhanced with various phenotypic characters. Differentially expressed genes (DEGs) related to metabolism, nutrient supply, and various stressed were up-regulated by approximately 1.5- to 80- fold. In this study, the morphology and stress tolerance of transgenic rice overexpressing NtROS2a were determined. Transgenic plants exhibited less and shorter lateral shoots. Under various treatments, rice overexpressing NtROS2a showed alleviation of damage symptoms with higher survival rate. After drought and re-watering treatment, transgenic rice seedlings restored their normal growth. However, wild type plants could not be rescued. These findings indicate that overexpression of NtROS2a gene in rice seedlings can increase their tolerance to drought stresses.