Overexpression of a microRNA-targeted NAC transcription factor improves drought and salt tolerance in Rice via ABA-mediated pathways

The NAC (NAM, AFAT, and CUC) transcription factors play critical roles in rice (Oryza sativa) development and stress regulation. Overexpressing a microRNA (miR164b)-resistant OsNAC2 mutant gene, which generates transcripts that cannot be targeted by miR164b, improves rice plant architecture and yield; however, the performance of these mOsNAC2-overexpressing lines, named ZUOErN3 and ZUOErN4, under abiotic stress conditions such as drought have not yet been fully characterized.In this study, we showed that the germination of ZUOErN3 and ZUOErN4 seeds was delayed in comparison with the wild-type (WT) seeds, although the final germination rates of all lines were over 95%. The quantification of the endogenous ABA levels revealed that the germinating mOsNAC2-overexpressing seeds had elevated ABA levels, which resulted in their slower germination. The mOsNAC2-overexpressing plants were significantly more drought tolerance than the WT plants, with the survival rate increasing from 11.2% in the WT to nearly 70% in ZUOErN3 and ZUOErN4 plants after a drought treatment. Salt (NaCl) tolerance was also increased in the ZUOErN3 and ZUOErN4 plants due to significantly increased ABA levels. A reverse transcription quantitative PCR (RT-qPCR) analysis showed a significant increase in the expression of the ABA biosynthesis genes OsNCED1 and OsNCED3 in the mOsNAC2-overexpressing lines, and the expression levels of the stress-responsive genes OsP5CS1, OsLEA3, and OsRab16 were significantly increased in these plants. Moreover, OsNAC2 directly interacted with the promoters of OsLEA3 and OsNCED3 in yeast one-hybrid assays.Taken together, our results show that OsNAC2 plays a positive regulatory role in drought and salt tolerance in rice through ABA-mediated pathways.