Scientia Agricultura Sinica ›› 2026, Vol. 59 ›› Issue (20): 4433-4446.doi: 10.3864/j.issn.0578-1752.2026.20.001

• CROP GENETICS & BREEDING·GERMPLASM RESOURCES·MOLECULAR GENETICS • Previous Articles     Next Articles

Functional Characterization of the ERF Transcription Factor TaERF1-A in Salt Tolerance in Triticum aestivum L.

YU Yang1,3(), LI MengTing2(), LIU Dan1, XU QingFen1, WAN ZiYu1, WANG TingTing3, FENG Gang1, WANG JianHe1, WANG CongLei1(), LIANG Dan1()   

  1. 1 Key Laboratory of Crop Genetic Breeding, Tianjin Academy of Agricultural Sciences, Tianjin 300192
    2 College of Food Science and Biotechnology, Tianjin Agricultural University, Tianjin 300192
    3 Northwest A&F University/National Key Laboratory of Crop Stress Resistance and High-Efficiency Production, Yangling 712100, Shaanxi
  • Received:2026-02-06 Accepted:2026-04-02 Online:2026-10-21 Published:2026-10-21
  • Contact: WANG CongLei, LIANG Dan

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Abstract:

【Objective】Ethylene response factor (ERF) transcription factors play important roles in regulating plant responses to abiotic stresses. This study aimed to investigate the function of the wheat ERF transcription factor gene TaERF1-A in salt tolerance, providing valuable genetic resources and a theoretical basis for breeding salt-tolerant wheat varieties.【Method】The expression pattern of TaERF1-A under salt stress was analyzed by qRT-PCR. Cis-acting regulatory elements in the promoter region of TaERF1-A were predicted using PlantCARE. Subcellular localization of the TaERF1-A protein was determined by transient expression in tobacco leaves. A TaERF1-A overexpression construct and a wheat gene-silencing vector were generated, and transgenic wheat lines with TaERF1-A overexpression or transient silencing were obtained via Agrobacterium-mediated transformation. Salt tolerance and related physiological indices of the transgenic plants were evaluated. Potential interacting proteins of TaERF1-A were screened using a yeast two-hybrid (Y2H) system, and the interactions were further validated by bimolecular fluorescence complementation (BiFC) and luciferase complementation assay (LCA). A dual-luciferase (LUC) assay was used to examine the effects of TaERF1-A and its interacting proteins on the promoters of stress-related genes.【Result】The TaERF1-A promoter contains various cis-regulatory elements associated with abiotic stress responses, and the encoded protein is localized in the nucleus. Under salt stress, TaERF1-A expression was significantly upregulated. Silencing of TaERF1-A in wheat resulted in increased malondialdehyde (MDA) content and decreased proline content as well as reduced activities of antioxidant enzymes (POD, CAT, and SOD) under salt stress. In addition, the expression of stress-responsive genes (TaDREB1 and TaERF3) and antioxidant enzyme genes (TaPOD, TaCAT, and TaSOD) was suppressed. In contrast, overexpression of TaERF1-A led to reduced MDA levels, increased proline accumulation, enhanced activities of POD, CAT, and SOD, and upregulated expression of TaDREB1, TaERF3, TaPOD, TaCAT, and TaSOD, indicating that TaERF1-A positively regulates salt tolerance in wheat. Y2H, BiFC, and LCA assays confirmed the interaction between TaERF1-A and TaNAC48 in yeast and tobacco cells. Moreover, the expression of TaNAC48 was induced by salt stress, and co-expression of TaNAC48 with TaERF1-A enhanced the activation of stress-responsive gene promoters. 【Conclusion】TaERF1-A acts as a positive regulator in the salt stress response of wheat. Overexpression and silencing of TaERF1-A respectively enhanced and reduced salt tolerance in transgenic wheat. TaERF1-A may cooperate with TaNAC48 to regulate the expression of stress-related genes, thereby improving salt tolerance in wheat.

Key words: wheat, TaERF1-A, gene silencing, overexpression, salt tolerance, protein interaction

Fig. 1

Characterization of the TaERF1-A gene A: Phylogenetic analysis of TaERF1-A. Ta: Triticum aestivum, Td: Triticum durum, Aet: Aegilops tauschii, Bd: Brachypodium distachyon, Zm: Zea mays, Pv: Panicum virgatum, Nu: Nelumbo nucifera, Vv: Vitis vinifera, Cc: Citrus clementina, At: Arabidopsis thaliana, Pt: Populus trichocarpa; B: Subcellular localization of TaERF1-A; C: Cis-acting elements in the TaERF1-A promoter (1500 bp upstream of the ATG)"

Fig. 2

Salt tolerance analysis of TaERF1-A-silenced wheat plants A: Expression of TaERF1-A in wheat following 200 mmol·L⁻¹ NaCl stress at different time points; B: Leaf pathogenesis map of BSMV-VIGS; C: Silencing efficiency of TaERF1-A in wheat; D: Phenotypic analysis of TaERF1-A-silenced wheat plants under salt stress. **: Extremely significant differences (P<0.01). The same as below"

Fig. 3

Changes in physiological traits of TaERF1-A-silenced wheat lines following salt stress A: Peroxidase (POD) activity; B: Catalase (CAT) activity; C: Superoxide dismutase (SOD) activity; D: Malondialdehyde (MDA) content; E: Proline content. *: Significant differences (P<0.05). The same as below"

Fig. 4

Analysis of the transcription levels of stress-responsive genes in TaERF1-A-silenced wheat plants"

Fig. 5

Analysis of salt tolerance in TaERF1-A-overexpressing wheat A: qRT-PCR validation of TaERF1-A expression in transgenic wheat lines; B: Salt tolerance phenotypic analysis of transgenic wheat seedlings"

Fig. 6

Changes in physiological traits of TaERF1-A- overexpressing wheat lines under salt stress A: Peroxidase (POD) activity; B: Catalase (CAT) activity; C: Superoxide dismutase (SOD) activity; C: Malondialdehyde (MDA) content; D: Proline content"

Fig. 7

Analysis of transcript levels of stress-responsive genes in salt-stressed wheat overexpression lines"

Fig. 8

Screening and validation of TaERF1-A-interacting proteins A: Screening of interacting proteins. Colonies in the red boxes were sequenced and identified as TaNAC48; B: Verification of the interaction between TaERF1-A and TaNAC48 in tobacco cells; C: Validation of the interaction between TaERF1-A and TaNAC48 using a LCA"

Table 1

TaERF1-6A interacting proteins annotation information"

基因Gene 候选蛋白Putative protein 登记号Accession
TaNAC48 NAC结构域蛋白NAC domain-containing protein 48 XM_044463398.1
TaVDAC 线粒体外膜孔道蛋白Mitochondrial outer membrane porin XM_044532654.1

Fig. 9

Response of wheat TaNAC48 to salt stress"

Fig. 10

Effect of the cooperative interaction between TaNAC48 and TaERF1-A on the activation of stress-responsive gene promoters A: Schematic diagrams of the reporter and effector constructs used in the dual-luciferase assay; B: Dual-luciferase reporter assay. Different letters indicate statistically significant differences at 0.01% significance level"

Fig. 11

Sequence variation of TaERF1-A in wheat"

Table 2

Distribution of two allelic variants of TaERF1-A among different wheat cultivars"

单倍型
Haplotype		 国内品种
Domestic cultivars		 国外品种
Foreign cultivars		 品种数目
Cultivar number
HapⅠ 周8425B、川麦104、济麦22、宁春4号
Zhou 8425B, Chuanmai 104, Jimai 22, Ningchun 4		 Alchemy, Julius, Landmark, Mace, Stanley 9
HapⅡ
 中国春、郑麦366、西农6028、矮抗58、小偃6号、北京8号、扬麦33、川麦48、扬麦158、邯6172、藏1817、济麦44、郑麦16、济麦47、郑麦22、克丰11、科农9024、石4185、苏麦3号
Chinese Spring, Zhengmai 366, Xinong 6028, Aikang 58, Xiaoyan 6, Beijing 8, Yangmai 33, Chuanmai 48, Yangmai 158, Han 6172, Zang 1817, Jimai 44, Zhengmai 16, Jimai 47, Zhengmai 22, Kefeng11, Kenong 9024, Shi 4185, Sumai 3		 ArinalrFor, Claire, Jagger,
Kariega, Lancer, Norin61, Robigus, Sy Mattis		 27
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