Integrated multi-omics analysis reveals the regulatory mechanism of cryptochrome 1- mediated anthocyanin biosynthesis in Populus trichocarpa

BACKGROUND: Cryptochrome 1 (CRY1) is a blue/ultraviolet-A light photoreceptor that governs diverse photoresponses and flavonoid secondary metabolism in plants. Although CRY1 functions are well established in herbaceous plants, its regulatory networks and specific roles in anthocyanin and proanthocyanidins (PAs) accumulation remain poorly understood in woody species. This study aimed to elucidate the molecular mechanism underlying CRY1-mediated biosynthesis of anthocyanins and PAs in poplar. RESULTS: Four cryptochrome genes were identified in the Populus trichocarpa (P. trichocarpa) genome. PtrCRY1a/1b were evolutionarily conserved, exhibit the highest expression levels in leaves, and their proteins were localized to the nucleus and cytoplasm. PtrCRY1a/1b gene functions and regulatory networks were comprehensively characterized using phenotypic analysis, targeted metabolomics, transcriptome sequencing and integrated multi-omics approaches. PtrCRY1a/1b overexpression caused dwarf phenotype and intense red pigmentation in leaves and stems, driven by massive anthocyanin accumulation. Metabolomic analysis identified cyanidin-3-O-glucoside, delphinidin-3-O-galactoside and delphinidin-3-O-glucoside as the key anthocyanin metabolites in overexpression lines, while proanthocyanidins B1 and B3 were the main components responsible for the reduced PAs accumulation. Transcriptome data showed significant enrichment of flavonoid biosynthesis-related genes. Integrative multi-omics analysis obtained 29 differentially expressed genes and 20 candidate transcription factors (TFs) associated with anthocyanin and PAs metabolism, and we constructed a correlation-based putative regulatory network. Thirteen of these TFs showed inverse expression correlations with anthocyanin and PAs accumulation, suggesting their potential dual regulatory roles in this process. CONCLUSIONS: In conclusion, via integrated multi-omics analyses, our work reveals a putative antagonistic regulatory pattern of PtrCRY1, where PtrCRY1a/1b overexpression enhances anthocyanin accumulation and suppresses PAs biosynthesis in P. trichocarpa. These findings advance our understanding of light-modulated secondary metabolism in woody plants. The high-anthocyanin severely dwarfed overexpression lines generated in this study have potential high ornamental value, and provide candidate genetic resources for color improvement and secondary metabolism breeding in forest trees.