Integrative metabolome and transcriptome analysis reveals GbKCS and GbMYB involved in the biosynthesis of ginkgolic acids

Ginkgolic acids is a sort of phenolic lipid compound predominantly enriched in the episperm of Ginkgo biloba. Ginkgolic acids exhibits cytotoxicity and allergenic effects, providing a defense against a variety of biotic stresses from insects, fungus, and bacteria. Ginkgolic acids specifically accumulates in the episperm of Ginkgo biloba, providing the seeds with a unique defence mechanism against biotic stresses and ensuring their survival. However, the regulatory mechanism of ginkgolic acids (GA) biosynthesis remains unclear. In this study, we performed combined metabolome and transcriptome analysis to identify key genes and metabolites involved in ginkgolic acids biosynthesis. The ginkgolic acids, including 6-[(8Z)-Pentadecenyl]-salicylic acid (GA 15:1), 2-hydroxy-6-pentadecylsalicylic acid (GA 15:0) and 5-(8-Pentadecenyl)-1, 3-benzenediol (BL 15:1), as well as their precursors, palmitic acid (FA 16:0) and palmitoleic acid (FA 16:1) were detected and showed regular changes in content during episperm development. From co-expression profiles of the differentially expressed genes, three enzyme-encoding genes including one ketoacyl-CoA synthase gene (GbKCS5) and two long-chain acyl-CoA synthase genes (GbAcsl4 and GbAcsl7) positively associated to GA 15:1 were identified as key candidates. Furthermore, coexpression analysis demonstrates that these genes might be negatively and positively regulated by the bHLH and MYB transcription factors, respectively. In addition, the endogenous jasmonic acid (JA) level in Ginkgo episperm were strongly associated with candidate ginkgolic-acid-related genes and metabolites. And JA may be an important regulator involved in ginkgolic acids biosynthesis. Exogenous JA treatment improved the expression of the GbMYB14, GbKCS5, GbAcsl4 and GbAcsl7 genes, while inhibited most Gb-bHLH genes, which was proposed to be significantly increased ginkgolic acids content. Our findings indicate that application of JA in the Ginkgo episperm can alter the expression levels of genes associated with ginkgolic acids and increase the production of ginkgolic acids. This provides a new insight for research on the regulation and utilization of ginkgolic acids.