Genomic identification of the NF-Y gene family in apple and functional analysis of MdNF-YB18 involved in flowering transition

Apple is a crucial economic product extensively cultivated worldwide. Its production and quality are closely related to the floral transition, which is regulated by intricate molecular and environmental factors. Nuclear factor Y (NF-Y) is a transcription factor that is involved in regulating plant growth and development, with certain NF-Ys play significant roles in regulating flowering. However, there is little information available regarding NF-Ys and their role in apple flowering development. In the present study, 51 NF-Y proteins were identified and classified into three subfamilies, including 11 MdNF-YAs, 26 MdNF-YBs, and 14 MdNF-YCs, according to their structural and phylogenetic features. Further functional analysis focused on MdNF-YB18. Overexpression of MdNF-YB18 in Arabidopsis resulted in earlier flowering compared to the wild-type plants. Subcellular localization confirmed MdNF-YB18 was located in the nuclear. Interaction between MdNFY-B18 and MdNF-YC3/7 was demonstrated through yeast two-hybrid (Y2H) and bimolecular fluorescence complementation (BiFC) assays. Yeast one-hybrid (Y1H) and the dual-luciferase reporter assays showed MdNF-YB18 could bind the promoter of MdFT1 and activate its expression. Moreover, this activation was enhanced with the addition of MdNF-YC3 and MdNF-YC7. Additionally, MdNF-YB18 also could interact with MdCOLs (CONSTANS Like). This study lays the foundation for exploring the functional traits of MdNF-Y proteins, highlighting the crucial role of MdNF-YB18 in activating MdFT1 in Malus.