Deciphering triterpenoid saponin biosynthesis by leveraging transcriptome response to methyl jasmonate elicitation in Saponaria vaccaria.

Methyl jasmonate (MeJA) is a known elicitor of plant specialized metabolism, including triterpenoid saponins. Saponaria vaccaria is an annual herb used in traditional Chinese medicine, containing large quantities of oleanane-type triterpenoid saponins with anticancer properties and structural similarities to the vaccine adjuvant QS-21. Leveraging the MeJA-elicited saponin biosynthesis, we identify multiple enzymes catalyzing the oxidation and glycosylation of triterpenoids in S. vaccaria. This exploration is aided by Pacbio full-length transcriptome sequencing and gene expression analysis. A cellulose synthase-like enzyme can not only glucuronidate triterpenoid aglycones but also alter the product profile of a cytochrome P450 monooxygenase via preference for the aldehyde intermediate. Furthermore, the discovery of a UDP-glucose 4,6-dehydratase and a UDP-4-keto-6-deoxy-glucose reductase reveals the biosynthetic pathway for the rare nucleotide sugar UDP-D-fucose, a likely sugar donor for fucosylation of plant natural products. Our work enables the production and optimization of high-value saponins in microorganisms and plants through synthetic biology approaches.

Identification and characterization of Schizophyllum commune type I metacaspases.

The role of programmed cell death in filamentous fungi is not well-understood, but is important due to the role of fungi in opportunistic infections. Plants, fungi and protozoa do not have caspase genes, but instead express the homologous proteins denoted metacaspases. To better understand the role of metacaspases in fungi we present an analysis of the sequences and activities of all five Type I metacaspases from Schizophyllum commune (ScMC), a mushroom-forming basiodmycete that undergoes sexual reproduction. The five Type I metacaspases of S. commune can be divided into two groups based on sequence similarity. Enzymes both with and without the N-terminal prodomain are active, but here we report on the constructs without the prodomains (Δpro). All five ScMCΔpro proteins show the highest enzymatic activity between pH 7 and 8 and require calcium for optimal activity. Optimal Ca2+ concentrations for ScMC1Δpro and ScMC2Δpro are 50 mM, while ScMC3, ScMC4Δpro and ScMC5Δpro activity is optimal around 5 mM calcium. All five S. commune metacaspases have similar substrate specificity. They are most active with Arg in the P1 position and inactive with Asp in the P1 position.