Enhancing Plant Stress Tolerance: The Role of LcWRKY40 Gene in Drought and Alkaline Salt Resistance in Tobacco and Yeast

<i>Leymus chinensis</i>, a halophytic perennial grass belonging to the Poaceae family, thrives in saline-alkali grasslands and harbors a rich repository of resistance-related genetic resources. This study focused on deciphering the stress-responsive mechanisms of <i>L. chinensis</i> by conducting transcriptomic sequencing under NaHCO₃ stress, which resulted in the annotation of a segment corresponding to the 51WRKY gene. The alkali-induced gene <i>LcWRKY40</i> (QIG37591) was identified by phylogenetic analysis. Real-time quantitative PCR analysis was performed on <i>L. chinensis</i> plants subjected to PEG6000 and alkaline salt (NaHCO₃) stress, and the results indicated that the <i>LcWRKY40</i> gene was upregulated in both the leaves and roots. The localization of the LcWRKY40 protein was confirmed by the use of green fluorescent protein (GFP) fusion technology in transformed rice protoplast cells. The GAL4-driven transformation of the <i>LcWRKY40</i> gene in INVScI yeast cells, which exhibited enhanced tolerance upon overexpression of the <i>LcWRKY40</i> gene under mannitol and alkaline salt (NaHCO₃) stress conditions. Under drought stress using mannitol, the fresh weight of Nicotiana tabacum overexpressing the <i>LcWRKY40</i> gene was significantly higher than that of wild-type(WT) tobacco. Through drought and salt alkali stress, we found that overexpressed tobacco at different stages always outperformed the wild type in terms of fresh weight, SOD, MDA, and Fv/Fm. This study provides preliminary insights into the involvement of the <i>LcWRKY40</i> gene in responding to drought and alkaline salt stresses, highlighting its role in enhancing plant resistance to drought and saline-alkaline conditions. These findings lay the foundation for future molecular breeding strategies aimed at improving grass resistance from different aspects.