Effect of Hypoxia on Shikonin Biosynthesis of Arnebia euchroma Roots and Its Transcriptome Analysis

Shikonin derivatives from Arnebia euchroma (Royle) Johnst. roots possess significant economic value as pharmaceutical agents, natural dyes, and cosmetic additives. To investigate hypoxia's impact on shikonin biosynthesis and its mechanisms, we subjected A. euchroma roots to hypoxic stress (1% O₂). Hypoxic roots remained white with minimal shikonin accumulation. Upon reoxygenation (≈20% O₂), rapid shikonin synthesis occurred within 4 h. qRT-PCR revealed suppressed expression of shikonin pathway genes (CYP76B74, CYP76B100, AePGT, etc.) under hypoxia. Transcriptome analysis of roots at 0 h (hypoxia), 4 h, and 48 h (reoxygenation) showed differential gene enrichment in carbon metabolism, phenylpropanoid biosynthesis, and glycolysis. Aerobic respiration genes were upregulated post-reoxygenation, while anaerobic respiration genes declined. Critically, genes regulating upstream shikonin pathways (MVA/MEP-PP) were inhibited under hypoxia. These results demonstrate hypoxia's transcriptional repression of shikonin biosynthesis and identify oxygen-dependent metabolic switches limiting production. These genes are involved in regulating the upstream MVA/MEP-PP pathway of shikonin biosynthesis, further confirming the inhibitory effect of hypoxia on shikonin biosynthesis.