CmAP2 interacts with CmRAX2 and promotes branching by affecting the cytokinin pathway in chrysanthemum

Branching is a crucial ornamental trait in chrysanthemum (Chrysanthemum morifolium), directly determining its cultivation methods, ornamental quality, and production costs. The AP2 transcription factor family governs diverse aspects of growth and development in plant; however, the role of these genes in plant branching remains unclear. In this study, we identified a novel AP2 protein, CmAP2, which interacts with CmRAX2-a chrysanthemum homolog of RAX2 that functions as a positive regulator of branching. This interaction was confirmed through yeast two-hybrid (Y2H), bimolecular fluorescence complementation (BiFC), dual-luciferase complementation assay (LCA), and co-immunoprecipitation (Co-IP) experiments. Furthermore, heterologous overexpression of CmAP2 in Arabidopsis thaliana significantly enhanced branching and induced a late-flowering phenotype. Consistently, in chrysanthemum, overexpression of CmAP2 promoted axillary bud outgrowth, increased axillary bud number, and enhanced plant height. Moreover, CmRAX2 and CmAP2 were rapidly induced by decapitation, and the expression trends were similarly. RNA-seq analysis further revealed significant differential expression of genes associated with cytokinin biosynthesis and signaling pathways in transgenic lines. Collectively, our findings demonstrate that CmAP2 interacts with CmRAX2 and positively regulates branching while concomitantly altering the expression of cytokinin-related genes.