CmCYC2-like transcription factors may interact with each other or bind to the promoter to regulate floral symmetry development in Chrysanthemum morifolium.

The complex capitulum of Chrysanthemum morifolium is often comprised of bilaterally symmetrical ray florets and radially symmetrical disc florets. The TCP transcription factor clade CYCLOIDEA2 (CYC2) appears to play a vital role in determining floral symmetry and in regulating floral organ development in Asteraceae. Our previous study identified six CmCYC2 genes from chrysanthemum and showed that CmCYC2c participated in the regulation of ray floret identity. However, the functions of other CmCYC2 genes and the underlying molecular mechanism of CmCYC2-mediated floral development regulation in chrysanthemums have not been elucidated. In this study, we analysed the function of CmCYC2 genes by ectopic expression of CmCYC2 in Arabidopsis. Then, we examined the protein-protein interaction using yeast two-hybrid (Y2H) and bimolecular fluorescence complementation (BiFC) assays. Finally, we analysed the protein-DNA interaction using yeast one-hybrid (Y1H) and dual-luciferase reporter assays. We found that ectopic expression of CmCYC2 genes in the Arabidopsis tcp1 mutant changed its floral symmetry and flowering time. Y2H and BiFC assays confirmed three pairs of interactions between CmCYC2 proteins, that is, CmCYC2b-CmCYC2d, CmCYC2b-CmCYC2e and CmCYC2c-CmCYC2d, suggesting that heterodimeric complexes may form between CmCYC2 proteins to increase their functional specificity. The results of Y1H and dual-luciferase reporter assays indicate that CmCYC2c can bind to the promoter of ClCYC2f. Our findings provided clues that CmCYC2-like transcription factors may interact with each other or bind to the promoter to regulate floral symmetry development in C. morifolium. KEY MESSAGE: CmCYC2-like transcription factors may interact with each other or bind to the promoter to regulate floral symmetry development in Chrysanthemum morifolium.