Integrated multi-omics reveals Li-miR828z-LiMYB114 regulatory module controlling anthocyanin biosynthesis during flower color development in Lagerstroemia indica

The formation of flower color in crape myrtle (Lagerstroemia indica) is driven by spatiotemporal regulation of anthocyanin biosynthesis, yet its molecular mechanisms remain unclear. This study employed multi-omics approaches including metabolomics, transcriptomics, and small RNA sequencing to reveal the regulatory mechanisms of flower color changes during petal development. Results showed anthocyanin content significantly increased during petal development, particularly at stage S3 where it increased 2.68-fold, coinciding with the transition from pale pink to bright red. Metabolomic analysis revealed a positive correlation between anthocyanin accumulation and total flavonoid content as petal development progressed, with the anthocyanin-to-total flavonoid ratio increasing from 0.30 at stage S1 to 0.55 at stage S4. This further supports the dominant role of anthocyanins in flower coloration. Transcriptome analysis identified 11,131 differentially expressed genes that exhibited stage-specific expression patterns, with significant enrichment in anthocyanin biosynthesis pathways at stage S3. Weighted gene co-expression network analysis (WGCNA) identified a regulatory module containing 41 transcription factors and 4 structural genes highly associated with anthocyanin accumulation. Notably, we discovered and validated a novel Li-miR828z-LiMYB114 regulatory module using small RNA sequencing. Li-miR828z directly inhibits LiMYB114 expression by binding to its target sequence, while LiMYB114 functions as a transcriptional activator directly binding to promoters of key anthocyanin biosynthesis genes (LiC4H, LiF3'H, and LiANS). This module exhibits precise temporal regulation: low Li-miR828z and high LiMYB114 expression in early development promote anthocyanin synthesis, while increased Li-miR828z later prevents excessive pigment accumulation. Our findings provide key insights into flower pigmentation and genetic targets for engineering diverse crape myrtle flower colors.