Transcription Factor CaERF3 Positively Regulates Pepper (Capsicum annuum L.) Salt Tolerance by Maintaining Galactose and Amino Sugar Homeostasis

Abstract

Salt stress severely restricts pepper growth and productivity. Although AP2/ERF transcription factors are widely involved in plant responses to environmental stress, their functions in salt tolerance in pepper remain largely unknown. In this study, transcriptome-based screening identified CaERF3 (Capana07g002313) as a primary-candidate salt-responsive ERF transcription factor. Functional analyses of CaERF3 knockout (KOERF3) and overexpression (OEERF3) lines demonstrated that CaERF3 positively regulated both seedling growth and salt tolerance. Under salt stress, OEERF3 plants exhibited higher SOD, POD, and CAT activities, lower TBARS accumulation, and a more favorable Na+/K+ balance, whereas KOERF3 plants showed the opposite phenotype. Integrated metabolomic and transcriptomic analyses indicated that CaERF3-mediated salt tolerance was associated with amino sugar and nucleotide sugar metabolism and galactose metabolism. The knockout of CaERF3 reduced the expression of the galactinol synthase genes CaGolS1-1 and CaGolS1-2, accompanied by decreased galactinol and stachyose accumulation, suggesting impaired raffinose-family oligosaccharide biosynthesis. In addition, genes associated with Golgi function, vesicle trafficking, and cell wall modification (CaLAC, CaVSR, CaUNC-1, and CaCHIEP3-1) were coordinately downregulated. Furthermore, CaERF3 functioned as a nuclear transcriptional activator and directly activated the endochitinase gene CaCHIEP3-1 by binding the GCC-box in its promoter. These findings establish CaERF3 as a positive regulator of salt tolerance in pepper and provide new insights into the transcriptional regulation of carbohydrate metabolism and glycan biosynthesis during salt stress.