The Molecular Mechanisms of m6A Methylation Modification in Regulating Bolting and Flowering in Radish

N6-methyladenosine (m6A) is a crucial epigenetic modification in mRNA and the most abundant methylation in eukaryotes. However, research on m6A modification in radish (Raphanus sativus) is not as extensive as in model plants. In this study, we analyzed m6A modification during the vegetative and reproductive growth stages of radish using m6A-seq and RNA-seq to explore its potential role in bolting and flowering. The results showed that m6A peaks significantly increased during the reproductive growth stage compared to the vegetative stage, with more m6A modification sites in the stop codon, 3' UTR, and promoter regions. Overexpression of the RsALKBH10B gene led to a global reduction in m6A modification and resulted in an early bolting and flowering phenotype, with transcription levels of key flowering factors RsSOC1 and RsFT increasing by 10-40 fold. Correlation analysis between differential m6A modification and differentially expressed genes indicated that genes such as RsSUF4, RsAGL13, and RsCDF5 might regulate bolting and flowering. Overexpression of RsSUF4 delayed bolting and flowering and decreased overall m6A modification levels; in positive transgenic plants, higher m6A levels of RsSUF4 were associated with lower transcription levels. These findings suggest that m6A methylation may regulate gene expression related to bolting and flowering in radish by affecting mRNA stability, ultimately leading to bolting and flowering. Overall design: In this study, we collected radish leaf material XHT at 30 days after sowing (nutrient growth stage, VG) and 120 days after sowing (reproductive growth stage, F) for m6A methylome sequencing, mining of differential genes, and functional validation of genes.