A WRKY protein, MfWRKY40, of resurrection plant Myrothamnus flabellifolia plays a positive role in regulating tolerance to drought and salinity stresses of Arabidopsis

Authors: HUANG, ZHUO - Sichuan Agricultural University; WANG, JIATONG - Sichuan Agricultural University; LI, YUAN - Sichuan Agricultural University; SONG, LI - Sichuan Agricultural University; CHEN, DUO'ER - Sichuan Agricultural University; LIU, LING - Sichuan Agricultural University; Jiang, Cai-Zhong

Submitted to: International Journal of Molecular Sciences
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: 7/21/2022
Publication Date: 7/24/2022
Citation: Huang, Z., Wang, J., Li, Y., Song, L., Chen, D., Liu, L., Jiang, C. 2022. A WRKY protein, MfWRKY40, of resurrection plant Myrothamnus flabellifolia plays a positive role in regulating tolerance to drought and salinity stresses of Arabidopsis. International Journal of Molecular Sciences. 23(15). Article 8145. https://doi.org/10.3390/ijms23158145.
DOI: https://doi.org/10.3390/ijms23158145

Interpretive Summary: Throughout the life cycle, plants are subjected to various environmental stresses, such as drought, high salt, extreme temperatures, etc. Water and salt stresses are among the two major forms of abiotic stress that have serious impacts on plant growth. To adapt such unfavorable environmental conditions, plants have evolved complex mechanisms to respond to these abiotic stresses at phenotypic (height, leaf size, root growth, etc.), physiological and biochemical (metabolism, enzymes, hormones, etc.), and molecular (protein expression, gene expression, etc.) levels. Transcription factors are specialized proteins that bind to DNA-regulatory sequences usually localized in the 5’-upstream region of target genes, to activate or suppress gene transcription. Numerous studies indicated that transcription factor families such as bZIP, bHLH, WRKY, MYB, AP2/ERF are involved in plants' response to abiotic stresses. WRKY, one of the largest transcription factor families in plants, is important for plant growth, development, and resistance to external stresses, especially drought and salt. For example, transgenic Arabidopsis overexpressing TaWRKY46 can increase drought tolerance in ABA-dependent and ABA-independent ways. Moreover, overexpression of cotton GhWRKY34 and GhWRKY39-1 in Arabidopsis and tobacco can significantly improve the salt tolerance of transgenic plants. Myrothamnus flabellifolia Welw is the only woody resurrection plant. Various aspects of drought tolerance of M. flabellifolia have been extensively studied, including its morphological structure, physiological and biochemical characteristics as well as its physiological and metabolic pathways during rehydration, but research on its drought tolerance mechanism is limited. Ma et al. examined the transcriptome of M. flabellifolia and found 295 TF are responsive to dehydration. MfWRKY40 was evidently up-regulated in early dehydration stage, suggesting that it may participates in response to water deficit. In this study, we cloned the MfWRKY40 and overexpressed it in Arabidopsis to investigate its potential roles in tolerance to drought and salt. MfWRKY40 is localized in nucleus and homologous to WRKY40 of Arabidopsis. It has a conserved WRKY motif but lacks a typical zinc finger motif in the WRKY domain. We overexpressed MfWRKY40 in Arabidopsis and found that transgenic lines exhibited better tolerance to both drought and salt stresses. Introduction of MfWRKY40 in Arabidopsis promoted primary root length elongation, and reduced water loss rate and stomata aperture (width/length) under stress, which may improve Arabidopsis the better water uptake and retention abilities. MfWRKY40 also facilitated osmotic adjustment under drought and salt stresses by accumulating more osmolytes, such as proline, soluble sugar and soluble protein. Additionally, the antioxidation ability was also significantly enhanced reflected by higher chlorophyll content, less malondialdehyde and reactive oxygen species accumulation, as well as higher antioxidation enzyme activities. All these results indicated that MfWRKY40 might positively regulate tolerance to drought and salinity stresses. Further investigation on relationship of the missing zinc finger motif of MfWRKY40 and its regulatory role is necessary to obtain better understanding of the mechanism underlying the excellent drought tolerance of M. flabellifolia.

Technical Abstract: WRKY transcription factors (TFs), one of the largest transcription factor families in plants, play an important role in abiotic stress responses. The resurrection plant Myrothamnus flabellifolia has a strong tolerance to dehydration, but there are only a few functional studies on WRKY proteins related to abiotic stress response have been conducted. In this study, we identified an early dehydration-induced gene MfWRKY40 of M. flabellifolia. MfWRKY40 is localized in nucleus and homologous to WRKY40 of Arabidopsis. It has a conserved WRKY motif but lacks a typical zinc finger motif in the WRKY domain. We overexpressed MfWRKY40 in Arabidopsis and found that transgenic lines exhibited better tolerance to both drought and salt stresses. Introduction of MfWRKY40 in Arabidopsis promoted primary root length elongation, and reduced water loss rate and stomata aperture (width/length) under stress, which may improve Arabidopsis the better water uptake and retention abilities. MfWRKY40 also facilitated osmotic adjustment under drought and salt stresses by accumulating more osmolytes, such as proline, soluble sugar and soluble protein. Additionally, the antioxidation ability was also significantly enhanced reflected by higher chlorophyll content, less malondialdehyde and reactive oxygen species accumulation, as well as higher antioxidation enzyme activities. All these results indicated that MfWRKY40 might positively regulate tolerance to drought and salinity stresses. Further investigation on relationship of the missing zinc finger motif of MfWRKY40 and its regulatory role is necessary to obtain better understanding of the mechanism underlying the excellent drought tolerance of M. flabellifolia.