Yuan Yuan ^1* Weijie Tang ^2* Jia-Yuan Cao ^2* Ke Zhong ^2* Ze-Jun Mo ^2* Ying Zhou ^2* Yu-Xin Pang ^2*

• ¹ School of Life Sciences and Biopharmaceutics, Guangdong Pharmaceutical University, Guangzhou, China
• ² College of Pharmaceutical Sciences, Guiyang College of Traditional Chinese Medicine, Guiyang, China

Blumea balsamifera, a half-woody plant belonging to the Asteraceae family, is valued as both a medicinal and industrial crop primarily for its phytochemical component, (-)-borneol. Nitrogen (N) is essential for regulating the growth of B. balsamifera and the biosynthesis of (-)-borneol; however, the molecular mechanisms by which N influences these processes remain inadequately understood. This study aimed to elucidate the effects of N on growth and (-)-borneol synthesis at the molecular level. A field experiment was conducted in which B. balsamifera plants were fertilized with three different nitrogen regimes: 0 kg N ha -1 (control, CK), 150 kg N ha -1 (N1 treatment), and 300 kg N ha -1 (N2 treatment).Physiological and biochemical assessments were performed to evaluate the growth and metabolic responses of the plants under these varying N conditions. Additionally, transcriptome sequencing of leaves of B. balsamifera was conducted to elucidate the underlying molecular mechanisms involved. The results indicated that both the N1 and N2 treatments significantly promoted the growth of B. balsamifera, with the 150 kg N ha -1 treatment (N1) resulting in the most favorable effects. The leaves harvested in October, November, and December presented the greatest accumulation of (-)-borneol, with corresponding yields of 782 mg plant -1 , 1102 mg plant -1 , and 1774 mg plant -1 , respectively, which were significantly different from those observed in the CK and N2 treatments. Comparative transcriptome analysis revealed a total of 6,714 differentially expressed genes (DEGs). Notably, several DEGs associated with auxin signaling and N metabolism were upregulated in the N1 and N2 treatments. In contrast, many DEGs related to carbohydrate metabolism, terpenoid backbone biosynthesis, monoterpenoid biosynthesis, and flavonoid biosynthesis were significantly upregulated in the CK treatment. Moreover, potential transcription factors (TFs) that may link N nutrition with the synthesis of medicinal components were identified. The CK treatment appeared to promote (-)-borneol synthesis by inducing the terpenoid synthase gene (BbTPS) and MYB TFs in the leaves of B. balsamifera. Our findings provide a comprehensive understanding of the relationship between N nutrition and (-)-borneol yield in B. balsamifera, offering valuable insights for future cultivation practices.