ABSTRACT
Amaranthus tricolor is an important vegetable, and its quality is affected by salt stress. Cation/proton antiporters (CPA) contribute to plant development and tolerance to salt stress. In this study, 35 CPA genes were identified from a genome database for A. tricolor, including 9 NHX, 5 KEA, and 21 CPA2 genes. Furthermore, in A. tricolor, the expression levels of most AtrNHX genes were higher at a low salinity level (50 or 100 mM NaCl) than in the control or 200 mM NaCl treatment. Levels of most AtrNHX genes were elevated in the stem. Moreover, AtrNHX8 was homologous to AtNHX4, which is involved in the regulation of sodium homeostasis and salt stress response. After AtrNHX8 overexpression in Arabidopsis thaliana, seed germination was better, and the flowering time was earlier than that of wild-type plants. Additionally, the overexpression of AtrNHX8 in A. thaliana improved salt tolerance. These results reveal the roles of AtrNHX genes under salt stress and provide valuable information on this gene family in amaranth.
ABSTRACT
Amaranth species are C4 plants that are rich in betalains, and they are tolerant to salinity stress. A small family of plant-specific TCP transcription factors are involved in the response to salt stress. However, it has not been investigated whether amaranth TCP1 is involved in salt stress. We elucidated that the growth and physiology of amaranth were affected by salt concentrations of 50-200 mmol·L-1 NaCl. The data showed that shoot and root growth was inhibited at 200 mmol·L-1, while it was promoted at 50 mmol·L-1. Meanwhile, the plants also showed physiological responses, which indicated salt-induced injuries and adaptation to the salt stress. Moreover, AtrTCP1 promoted Arabidopsis seed germination. The germination rate of wild-type (WT) and 35S::AtrTCP1-GUS Arabidopsis seeds reached around 92% by the seventh day and 94.5% by the second day under normal conditions, respectively. With 150 mmol·L-1 NaCl treatment, the germination rate of the WT and 35S::AtrTCP1-GUS plant seeds was 27.0% by the seventh day and 93.0% by the fourth day, respectively. Under salt stress, the transformed 35S::AtrTCP1 plants bloomed when they grew 21.8 leaves after 16.2 days of treatment, which was earlier than the WT plants. The transformed Arabidopsis plants flowered early to resist salt stress. These results reveal amaranth's growth and physiological responses to salt stress, and provide valuable information on the AtrTCP1 gene.
Subject(s)
Amaranthus , Arabidopsis , Gene Expression Regulation, Plant , Germination , Plant Proteins , Salt Stress , Gene Expression Regulation, Plant/drug effects , Amaranthus/drug effects , Amaranthus/genetics , Amaranthus/growth & development , Plant Proteins/genetics , Plant Proteins/metabolism , Germination/drug effects , Germination/genetics , Arabidopsis/genetics , Arabidopsis/drug effects , Arabidopsis/growth & development , Arabidopsis/physiology , Transcription Factors/genetics , Transcription Factors/metabolism , Plants, Genetically Modified , Plant Roots/growth & development , Plant Roots/drug effects , Plant Roots/genetics , Seeds/drug effects , Seeds/growth & development , Seeds/genetics , Salt Tolerance/genetics , Sodium Chloride/pharmacologyABSTRACT
Introduction: WRKY TFs (WRKY transcription factors) contribute to the synthesis of secondary metabolites in plants. Betalains are natural pigments that do not coexist with anthocyanins within the same plant. Amaranthus tricolor ('Suxian No.1') is an important leaf vegetable rich in betalains. However, the WRKY family members in amaranth and their roles in betalain synthesis and metabolism are still unclear. Methods: To elucidate the molecular characteristics of the amaranth WRKY gene family and its role in betalain synthesis, WRKY gene family members were screened and identified using amaranth transcriptome data, and their physicochemical properties, conserved domains, phylogenetic relationships, and conserved motifs were analyzed using bioinformatics methods. Results: In total, 72 WRKY family members were identified from the amaranth transcriptome. Three WRKY genes involved in betalain synthesis were screened in the phylogenetic analysis of WRKY TFs. RT-qPCR showed that the expression levels of these three genes in red amaranth 'Suxian No.1' were higher than those in green amaranth 'Suxian No.2' and also showed that the expression level of AtrWRKY42 gene short-spliced transcript AtrWRKY42-2 in Amaranth 'Suxian No.1' was higher than that of the complete sequence AtrWRKY42-1, so the short-spliced transcript AtrWRKY42-2 was mainly expressed in 'Suxian No.2' amaranth. Moreover, the total expression levels of AtrWRKY42-1 and AtrWRKY42-2 were down-regulated after GA3 treatment, so AtrWRKY42-2 was identified as a candidate gene. Therefore, the short splice variant AtrWRKY42-2 cDNA sequence, gDNA sequence, and promoter sequence of AtrWRKY42 were cloned, and the PRI 101-AN-AtrWRKY42-2-EGFP vector was constructed to evaluate subcellular localization, revealing that AtrWRKY42-2 is located in the nucleus. The overexpression vector pRI 101-AN-AtrWRKY42-2-EGFP and VIGS (virus-induced gene silencing) vector pTRV2-AtrWRKY42-2 were transferred into leaves of 'Suxian No.1' by an Agrobacterium-mediated method. The results showed that AtrWRKY42-2 overexpression could promote the expression of AtrCYP76AD1 and increase betalain synthesis. A yeast one-hybrid assay demonstrated that AtrWRKY42-2 could bind to the AtrCYP76AD1 promoter to regulate betalain synthesis. Discussion: This study lays a foundation for further exploring the function of AtrWRKY42-2 in betalain metabolism.
