Molecular Characterization of U-box E3 Ubiquitin Ligases (TaPUB2 and TaPUB3) Involved in the Positive Regulation of Drought Stress Response in Arabidopsis

Abstract

Plant U-box E3 ubiquitin ligase (PUB) is involved in various environmental stress conditions. However, the molecular mechanism of U-box proteins in response to abiotic stress in wheat remains unknown. In this study, two U-box E3 ligase genes (TaPUB2 and TaPUB3), which are highly expressed in response to adverse abiotic stresses, were isolated from common wheat, and their cellular functions were characterized under drought stress. Transient expression assay revealed that TaPUB2 was localized in the cytoplasm and Golgi apparatus, whereas TaPUB3 was expressed only in the Golgi apparatus in wheat protoplasts. Additionally, TaPUB2 and TaPUB3 underwent self-ubiquitination. Moreover, TaPUB2/TaPUB3 heterodimer was identified in yeast and the cytoplasm of wheat protoplasts using a pull-down assay and bimolecular fluorescence complementation analysis. Heterogeneous overexpression of TaPUB2 and TaPUB3 conferred tolerance to drought stress. Taken together, these results implied that the heterodimeric form of U-box E3 ubiquitin ligases (TaPUB2/TaPUB3) responded to abiotic stress and roles as a positive regulator of drought stress tolerance.

Wheat (Triticum aestivum. L) Plant U‐box E3 ligases TaPUB2 and TaPUB3 enhance ABA response and salt stress resistance in Arabidopsis

“…In this study, we explored the functions of two wheat PUBs (TaPUB2 and TaPUB3) under salinity stress and ABA conditions. Although overexpression of these genes improved drought tolerance in transgenic A. thaliana compared with the WT in our recent study [12], their function under ABA and salinity stress The expression patterns of various ABA-responsive genes (AtABF2, AtABF4, AtABI4 and AtABI5) were investigated using qRT-PCR. The difference in the expression of each gene during ABA treatment (5 h) was compared with that in the normal treatment (0 h).…”

“…In this study, we explored the functions of two wheat PUBs (TaPUB2 and TaPUB3) under salinity stress and ABA conditions. Although overexpression of these genes improved drought tolerance in transgenic A. thaliana compared with the WT in our recent study [12], their function under ABA and salinity stress conditions is unknown. Therefore, we designed an experiment to determine how these genes respond to ABA and salinity stress, which are two of the major abiotic stresses that affect crop growth, development and yield.…”

“…We recently discovered that TaPUB2 and TaPUB3 are more involved in enhancing drought tolerance in transgenic Arabidopsis thaliana than that in WT plants. We hypothesized that TaPUB2 and TaPUB3 may be involved in regulating salinity stress in an ABA‐dependent or ABA‐independent manner, as they were involved in regulating drought stress in our previous study [12]. To accomplish these goals, we created transgenic A. thaliana plants that overexpressed TaPUB2 and TaPUB3 and performed various physiological experiments to confirm our hypothesis.…”

“…The 35S: TaPUB2 and 35S: TaPUB3 constructs were used for Agrobacterium (GV3101)‐mediated gene transfer and transformed into A. thaliana using the floral dipping method [24]. The transforming information for 35S: TaPUB2 and 35S: TaPUB3 has been previously described [12]. Seeds of A. thaliana (WT and TaPUB2‐ and TaPUB3‐overexpression lines) were grown on Murashige–Skoog (MS) media and soil in a glasshouse and growth chamber under controlled conditions of 22 °C, 16/8 day/night light cycle and 60% relative humidity.…”

“…Many PUB proteins have been found to play significant regulatory roles in plant growth and physiological activities. Most functions of PUB proteins in response to biotic or abiotic stress conditions have been investigated [11][12][13]. Furthermore, PUB proteins with plant developmental functions, such as self-incompatibility [14] and seed germination [15], have been defined.…”