Overexpression of the Wheat (Triticum aestivum L.) TaPEPKR2 Gene Enhances Heat and Dehydration Tolerance in Both Wheat and Arabidopsis

Abstract

Wheat (Triticum aestivum L.) yield and quality are adversely affected by heat, drought, or the combination of these two stresses in many regions of the world. A phosphoenolpyruvate carboxylase kinase-related kinase gene, TaPEPKR2, was identified from our previous heat stress-responsive transcriptome analysis of heat susceptible and tolerant wheat cultivars. Based on the wheat cultivar Chinese Spring genome sequence, TaPEPKR2 was mapped to chromosome 5B. Expression analysis revealed that TaPEPKR2 was induced by heat and polyethylene glycol treatment. To analyze the function of TaPEPKR2 in wheat, we transformed it into the wheat cultivar Liaochun10, and observed that the transgenic lines exhibited enhanced heat and dehydration stress tolerance. To examine whether TaPEPKR2 exhibits the same function in dicotyledonous plants, we transformed it into Arabidopsis, and found that its overexpression functionally enhanced tolerance to heat and dehydration stresses. Our results imply that TaPEPKR2 plays an important role in both heat and dehydration stress tolerance, and could be utilized as a candidate gene in transgenic breeding.

Keywords: Arabidopsis; PEP carboxylase kinase-related kinase; TaPEPKR2; dehydration stress; heat stress; wheat.

FIGURE 1

Sequence alignment of TaPEPKR2 and orthologous genes from Arabidopsis, rice, and maize. Regions corresponding to the S_TKc domain are indicated by the SMART program.

FIGURE 2

Relative expression of TaPEPKR2 under heat (A) and dehydration (B) stress conditions, as determined by RT-qPCR. Data represent the mean of three replicates ± SD.

FIGURE 3

Heat tolerance assay of TaPEPKR2 transgenic wheat seedlings. (A) Phenotypes of 10-day-old LC10 and three TaPEPKR2 transgenic lines before heat treatment (left). 5-day-old LC10 and three TaPEPKR2 transgenic wheat lines were treated at 45°C for 18 h, then recovered at 22°C for 5 days. Images were taken post-recovery (right). (B) Ion leakage assay of the transgenic seedlings in panel (A) after heat treatment. The symbol “^∗” indicates significance at P < 0.05.

FIGURE 4

Dehydration stress tolerance assay of TaPEPKR2 transgenic wheat seedlings. (A) Phenotypes of 10-day-old LC10 and transgenic lines overexpressing TaPEPKR2 under control and 20% PEG conditions. (B) Total root length of 10-day-old LC10 and transgenic seedlings under stress conditions. The symbol “^∗” indicates significance at P < 0.05.

FIGURE 5

Effect of TaPEPKR2 overexpression on heat tolerance in transgenic Arabidopsis. (A) Real-time PCR of transgenic Arabidopsis lines overexpressing TaPEPKR2. L1–L7 are seven individual TaPEPKR2 transgenic lines. Relative TaPEPKR2 expression in transgenic line L1 was set as the control. Data are the means of three replicates ± SD. (B) The heat treatment regime. (C) Seedlings grown for 5.5 days at 22°C were used as the control. (D) Phenotypes of WT, L1, L3, and L5 after heat treatment and recovery at 22°C for 7 days. (E) Survival rates of three TaPEPKR2 transgenic lines (L1, L3, and L5) and WT after heat treatment. Data are the mean of three replicates ± SD (n = 60 for each experiment). (F) Ion leakage assay of the seedlings in panel (D) at 42°C for 1 h. Data are the means ± SD of three replicates; ^∗∗P < 0.01 (Student’s t-test).

FIGURE 6

TaPEPKR2 transgenic plants show improved dehydration tolerance. (A) WT and TaPEPKR2 transgenic seedlings grown in soil were subjected to water deprivation for 25 days, then re-watered for ∼1 week. (B) Survival rates of WT and TaPEPKR2 transgenic plants after dehydration treatment. Data are the mean ± SD of three replicates (n = 30 for each experiment); ^∗∗P < 0.01 (Student’s t-test). (C) Relative water loss rate of WT and TaPEPKR2 transgenic plants. Data are the mean ± SD of three replicates; ^∗P < 0.05, ^∗∗P < 0.01 (Student’s t-test). (D) Leaf wilting of detached leaves from WT and transgenic plants after 5 h.