Fig. 4

CmHRE2-like directly activates CmACS6 genes in chrysanthemum. A Heterologous expression of CmACS6 promoter in Arabidopsis thaliana in response to waterlogging stress. B Schematic of predicted ERF binding sites in the CmACS6 promoter. C Yeast one-hybrid assay demonstrating the binding of CmHRE2-like to CmACS6 promoter. D EMSA assay showing direct binding of CmHRE2-like protein to the CmACS6 promoter in vitro. E ChIP-qPCR analysis confirming CmHRE2-like directly binding to the CmACS6 promoter. F Luciferase reporter analysis indicating CmHRE2-like activates the expression of CmACS6 in tobacco. G Dual-luciferase assay demonstrating CmHRE2-like stimulates CmACS6 promoter activity in chrysanthemum protoplasts. Error bars represent standard deviation (SD, n = 3); Different lowercase letters indicate significant differences at P < 0.05 (Student's t-test and ANOVA, Turkey's correction)