RNA-Seq analysis reveals the important co-expressed genes associated with polyphyllin biosynthesis during the developmental stages of Paris polyphylla.

BACKGROUND: Plants synthesize metabolites to adapt to a continuously changing environment. Metabolite biosynthesis often occurs in response to the tissue-specific combinatorial developmental cues that are transcriptionally regulated. Polyphyllins are the major bioactive components in Paris species that demonstrate hemostatic, anti-inflammatory and antitumor effects and have considerable market demands. However, the mechanisms underlying polyphyllin biosynthesis and regulation during plant development have not been fully elucidated. RESULTS: Tissue samples of P. polyphylla var. yunnanensis during the four dominant developmental stages were collected and investigated using high-performance liquid chromatography and RNA sequencing. Polyphyllin concentrations in the different tissues were found to be highly dynamic across developmental stages. Specifically, decreasing trends in polyphyllin concentration were observed in the aerial vegetative tissues, whereas an increasing trend was observed in the rhizomes. Consistent with the aforementioned polyphyllin concentration trends, different patterns of spatiotemporal gene expression in the vegetative tissues were found to be closely related with polyphyllin biosynthesis. Additionally, molecular dissection of the pathway components revealed 137 candidate genes involved in the upstream pathway of polyphyllin backbone biosynthesis. Furthermore, gene co-expression network analysis revealed 74 transcription factor genes and one transporter gene associated with polyphyllin biosynthesis and allocation. CONCLUSIONS: Our findings outline the framework for understanding the biosynthesis and accumulation of polyphyllins during plant development and contribute to future research in elucidating the molecular mechanism underlying polyphyllin regulation and accumulation in P. polyphylla.

Engineering a mevalonate pathway in Halomonas bluephagenesis for the production of lycopene.

Introduction: Red-colored lycopene has received remarkable attention in medicine because of its antioxidant properties for reducing the risks of many human cancers. However, the extraction of lycopene from natural hosts is limited. Moreover, the chemically synthesized lycopene raises safety concerns due to residual chemical reagents. Halomonas bluephagenesis is a versatile chassis for the production of fine chemicals because of its open growth property without sterilization. Methods: A heterologous mevalonate (MVA) pathway was introduced into H. bluephagenesis strain TD1.0 to engineer a bacterial host for lycopene production. A pTer7 plasmid mediating the expression of six MVA pathway genes under the control of a phage PMmp1 and an Escherichia coli Ptrc promoters and a pTer3 plasmid providing lycopene biosynthesis downstream genes derived from Streptomyces avermitilis were constructed and transformed into TD1.0. The production of lycopene in the engineered H. bluephagenesis was evaluated. Optimization of engineered bacteria was performed to increase lycopene yield. Results: The engineered TD1.0/pTer7-pTer3 produced lycopene at a maximum yield of 0.20 mg/g dried cell weight (DCW). Replacing downstream genes with those from S. lividans elevated the lycopene production to 0.70 mg/g DCW in the TD1.0/pTer7-pTer5 strain. Optimizing the PMmp1 promoter in plasmid pTer7 with a relatively weak Ptrc even increased the lycopene production to 1.22 mg/g DCW. However, the change in the Ptrc promoter in pTer7 with PMmp1 did not improve the yield of lycopene. Conclusion: We first engineered an H. bluephagenesis for the lycopene production. The co-optimization of downstream genes and promoters governing MVA pathway gene expressions can synergistically enhance the microbial overproduction of lycopene.