Comprehensive characterization of the genes in AP2/ERF family and their involvement in salt–alkali stress response during Nelumbo nucifera seed germination

Abstract

Nelumbo nucifera Gaertn. is an economically and ecologically important aquatic plant, but its growth and productivity are severely constrained by soil salinization and alkalization. AP2/ERF transcription factors are key regulators of plant abiotic stress responses; however, their roles in salt–alkali tolerance in N. nucifera remain largely unclear. In this study, we performed a genome-wide identification and characterization of the AP2/ERF gene family in N. nucifera, followed by phylogenetic, structural, and physicochemical analyses. A total of 101 AP2/ERF genes were identified and classified into five subfamilies, showing both evolutionary conservation and species-specific divergence compared with Arabidopsis thaliana. Physiological analyses during seed germination under salt–alkali stress revealed significant changes in malondialdehyde content, proline accumulation, and antioxidant enzyme activities, suggesting activation of oxidative stress defense and osmotic adjustment mechanisms. Transcriptome profiling of seedlings treated with 150 mM salt–alkali solution for 5 and 10 days identified 7,350 differentially expressed genes, including 29 AP2/ERF members responsive to stress. Among them, 13 genes, including AP2-9, ERF23, ERF15, ERF31, ERF34, and DREB21, were consistently upregulated under both treatments, indicating their potential roles in stress adaptation. qRT-PCR validation further confirmed the sustained upregulation of key genes AP2-9, ERF23, ERF34, and DREB21, consistent with transcriptome data. Overall, this study provides the first comprehensive overview of the AP2/ERF gene family in N. nucifera and identifies candidate regulators involved in salt–alkali stress responses, offering valuable insights into the molecular mechanisms of stress adaptation and potential genetic resources for breeding salt–alkali tolerant aquatic plants.