Multi-Omics Reveals Light-Quality-Dependent Phytohormone and Transcription Factor Networks Regulating Flavonoid Biosynthesis in Ludisia discolor

Background/Objectives: Ludisia discolor, an endangered medicinal orchid, is a vital source of bioactive flavonoids which requires in vitro tissue culture for propagation and metabolite production. While light quality influences metabolic processes, the mechanisms connecting light conditions, phytohormone signaling, and flavonoid biosynthesis remain unclear. This study investigates how specific light qualities trigger secondary metabolism to improve tissue culture and conservation strategies. Methods: L. discolor was cultivated under strictly regulated LED environments (blue, red, yellow, and green). An integrated multi-omics approach, combining transcriptomic sequencing and targeted metabolomic profiling, was employed to analyze leaves, correlating plant hormone changes with flavonoid metabolite levels. Results: LED light qualities significantly altered flavonoid and phytohormone profiles, yielding 80 unique flavonoids. Blue and red light effectively promoted flavonoid accumulation, whereas yellow light suppressed it. Transcriptomics, validated by qRT-PCR, revealed distinct expression patterns in key structural genes (e.g., 4CL, PAL, CYP73A, FLS, CCoAOMT, C12RT1). Ten transcription factors (including MYB93, bZIP36, bHLH4, and bZIP44) with hormone-responsive cis-elements were co-expressed with 16 structural genes. Notably, blue light induced reactive oxygen species (ROS) signaling, activating phytohormone production (IAA, GA, ABA). These hormones subsequently stimulated transcription factors, increasing the biosynthesis of compounds like neohesperidin and hesperetin. Conclusions: We propose a novel regulatory model where light-induced ROS and phytohormone cascades activate specific transcription factors, enhancing structural gene expression in the flavonoid pathway. These findings elucidate the molecular mechanisms of light-driven secondary metabolism, providing valuable insights for the sustainable agriculture and ex situ conservation of endangered medicinal orchids.