Molecular role of cytokinin in bud activation and outgrowth in apple branching based on transcriptomic analysis

Axillary bud activation and outgrowth were dependent on local cytokinin, and that bud activation preceded the activation of cell cycle and cell growth genes in apple branching. Cytokinin is often applied to apple trees to produce more shoot branches in apple seedlings. The molecular response of apple to the application of cytokinin, and the relationship between bud activation and cell cycle in apple branching, however, are poorly understood. In this study, RNA sequencing was used to characterize differential expression genes in axillary buds of 1-year grafted "Fuji" apple at 4 and 96 h after cytokinin application. And comparative gene expression analyses were performed in buds of decapitated shoots and buds of the treatment of biosynthetic inhibitor of cytokinin (Lovastatin) on decapitated shoots. Results indicated that decapitation and cytokinin increased ZR content in buds and internodes at 4-8 h, and induced bud elongation at 96 h after treatment, relative to buds in shoots receiving the Lovastatin treatment. RNA-seq analysis indicated that differential expression genes in auxin and cytokinin signal transduction were significantly enriched at 4 h, and DNA replication was enriched at 96 h. Cytokinin-responsive type-A response regulator, auxin polar transport, and axillary meristem-related genes were up-regulated at 4 h in the cytokinin and decapitation treatments, while qRT-PCR analysis showed that cell cycle and cell growth genes were up-regulated after 8 h. Collectively, the data indicated that bud activation and outgrowth might be dependent on local cytokinin synthesis in axillary buds or stems, and that bud activation preceded the activation of cell cycle genes during the outgrowth of ABs in apple shoots.