Gibberellic acid overproduction in Fusarium fujikuroi using regulatory modification and transcription analysis.

Gibberellic acid (GA), one of the natural diterpenoids produced by Fusarium fujikuroi, serves as an important phytohormone in agriculture for promoting plant growth. Presently, the metabolic engineering strategies for increasing the production of GA are progressing slowly, which seriously restricted the advancing of the cost-effective industrial production of GA. In this study, an industrial strain with high-yield GA of F. fujikuroi was constructed by metabolic modification, coupling with transcriptome analysis and promoter engineering. The over-expression of AreA and Lae1, two positive factors in the regulatory network, generated an initial producing strain with GA production of 2.78 g L. Compared with a large abundance of transcript enrichments in the GA synthetic gene cluster discovered by the comparative transcriptome analysis, geranylgeranyl pyrophosphate synthase 2 (Ggs2), and cytochrome P450-3 genes, two key genes that respectively participated in the initial and final step of biosynthesis, were identified to be downregulated when the highest GA productivity was obtained. Employing with a nitrogen-responsive bidirectional promoter, the two rate-limiting genes were dynamically upregulated, and therefore, the production of GA was increased to 3.02 g L. Furthermore, the top 20 upregulated genes were characterized in GA over-production, and their distributions in chromosomes suggested potential genomic regions with a high transcriptional level for further strain development. The construction of a GA high-yield-producing strain was successfully achieved, and insights into the enriched functional transcripts provided novel strain development targets of F. fujikuroi, offering an efficient microbial development platform for industrial GA production. KEY POINTS: • Global regulatory modification was achieved in F. fujikuroi for GA overproduction. • Comparative transcriptome analysis revealed bottlenecks in GA specific-pathway. • A dynamically nitrogen-regulated bidirectional promoter was cloned and employed.