In plants, chloride channels (CLC) are involved in a series of specific functions, such as regulation of nutrient transport and stress tolerance. Members of the wheat Triticum aestivum L. CLC (TaCLC) gene family have been proposed to encode anion channels/transporters that may be related to nitrogen transportation. To better understand their roles, TaCLC family was screened and 23 TaCLC gene sequences were identified using a Hidden Markov Model in conjunction with wheat genome database. Gene structure, chromosome location, conserved motif, and expression pattern of the resulting family members were then analyzed. Phylogenetic analysis showed that the TaCLC family can be divided into two subclasses (I and II) and seven clusters (-a, -c1, -c2, -e, -f1, -f2, and -g2). Using a wheat RNA-seq database, the expression pattern of TaCLC family members was determined to be an inducible expression type. In addition, seven genes from seven different clusters were selected for quantitative real-time PCR (qRT-PCR) analysis under low nitrogen stress or salt stress conditions, respectively. The results indicated that the gene expression levels of this family were up-regulated under low nitrogen stress and salt stress, except the genes of TaCLC-c2 cluster which were from subfamily -c. The yeast complementary experiments illustrated that TaCLC-a-6AS-1, TaCLC-c1-3AS, and TaCLC-e-3AL all had anion transport functions for NO3− or Cl−, and compensated the hypersensitivity of yeast GEF1 mutant strain YJR040w (Δgef1) in restoring anion-sensitive phenotype. This study establishes a theoretical foundation for further functional characterization of TaCLC genes and provides an initial reference for better understanding nitrate nitrogen transportation in wheat.

Potassium (K) is an essential nutrient for plant physiological processes. Members of the HAK/KUP/KT gene family act as potassium transporters, and the family plays an important role in potassium uptake and utilization in plants. In this study, the TaHAK13 gene was cloned from wheat and its function characterized. Real-time quantitative PCR (RT-qPCR) revealed that TaHAK13 expression was induced by environmental stress and up-regulated under drought (PEG6000), low potassium (LK), and salt (NaCl) stress. GUS staining indicated that TaHAK13 was mainly expressed in the leaf veins, stems, and root tips in Arabidopsis thaliana, and expression varied with developmental stage. TaHAK13 mediated K+ absorption when heterologously expressed in yeast CY162 strains, and its activity was slightly stronger than that of a TaHAK1 positive control. Subcellular localization analysis illustrated that TaHAK13 was located to the plasma membrane. When c(K+) ≤0.01 mM, the root length and fresh weight of TaHAK13 transgenic lines (athak5/TaHAK13, Col/TaHAK13) were significantly higher than those of non-transgenic lines (athak5, Col). Non-invasive micro-test technology (NMT) indicated that the net K influx of the transgenic lines was also higher than that of the non-transgenic lines. This suggests that TaHAK13 promotes K+ absorption, especially in low potassium media. Membrane-based yeast two-hybrid (MbY2H) and luciferase complementation assays (LCA) showed that TaHAK13 interacted with TaNPF5.10 and TaNPF6.3. Our findings have helped to clarify the biological functions of TaHAK13 and established a theoretical framework to dissect its function in wheat.