Genome of Tripterygium wilfordii and identification of cytochrome P450 involved in triptolide biosynthesis

Abstract

Triptolide is a trace natural product of Tripterygium wilfordii. It has antitumor activities, particularly against pancreatic cancer cells. Identification of genes and elucidation of the biosynthetic pathway leading to triptolide are the prerequisite for heterologous bioproduction. Here, we report a reference-grade genome of T. wilfordii with a contig N50 of 4.36 Mb. We show that copy numbers of triptolide biosynthetic pathway genes are impacted by a recent whole-genome triplication event. We further integrate genomic, transcriptomic, and metabolomic data to map a gene-to-metabolite network. This leads to the identification of a cytochrome P450 (CYP728B70) that can catalyze oxidation of a methyl to the acid moiety of dehydroabietic acid in triptolide biosynthesis. We think the genomic resource and the candidate genes reported here set the foundation to fully reveal triptolide biosynthetic pathway and consequently the heterologous bioproduction.

The genome analysis ofTripterygium wilfordiireveals TwCYP712K1 andTwCYP712K2responsible for oxidation of friedelin in celastrol biosynthesis pathway

“…In this study, we provided a high-quality reference genome of T. wilfordii with a 340.12 Mb genome assembly and 3.09 Mb contig N50, and successfully anchored 91.02% sequences into 23 pseudochromosomes ( Table 1). The quality of our genome assembly was close to the recently reported T. wilfordii genome (348.38 Mb total contigs and 4.36 Mb contig N50) (Tu et al, 2020).…”

The chromosome-level reference genome assembly for Panax notoginseng and insights into ginsenoside biosynthesis

“…Gene mining of high-quality genomic and transcriptomic data can provide resources for further exploration of plant growth and secondary metabolism mechanisms ( Tu et al., 2020 ). As early as 2017, two P. notoginseng reference genomes were published ( Zhang et al., 2017a ; Chen et al., 2017 ); however, the quality of these genomes was insufficient because of the limited sequencing capacity at that time.…”

Nature's Chemists: The Discovery and Engineering of Phytochemical Biosynthesis

“…This feature has often been used to identify genes involved in the biosynthesis of various natural products. Cytochrome P450 (CYP728B70) catalyzing the oxidation step in triptolide biosynthesis, an abietane-type diterpenoid from Tripterygium wilfordii , was identified through an integrative gene prioritization approach (Tu et al, 2020 ). The prioritized candidate genes were highly expressed in MeJA-induced cells and/or in the root bark (main accumulation site of the diterpene), as well as, exhibited similar expression patterns to already characterized enzymes in the pathway; copalyl diphosphate synthase and miltiradiene synthase (Tu et al, 2020 ).…”

“…This approach may be preferred over in vitro assays if enzyme purification is challenging or higher throughput is required. If the substrate of interest is not available in yeast, an exogenously supplied substrate can be used to build the desired secondary metabolite (Tu et al, 2020 ). Another option is to engineer yeast strains through genome engineering (e.g., CRISPR or homologous recombination) to accumulate specific precursors that are not naturally present in yeast (Luo et al, 2019 ; Munakata et al, 2019 ), or to shutdown endogenous competitive branches that deplete the precursor of interest (Moses et al, 2014 ).…”