Exploring the Interplay between Metabolic Pathways and Taxane Production in Elicited Taxus baccata Cell Suspensions

Abstract

Taxus cell cultures are a reliable biotechnological source of the anticancer drug paclitaxel. However, the interplay between taxane production and other metabolic pathways during elicitation remains poorly understood. In this study, we combined untargeted metabolomics and elicited Taxus baccata cell cultures to investigate variations in taxane-associated metabolism under the influence of 1 µM coronatine (COR) and 150 µM salicylic acid (SA). Our results demonstrated pleiotropic effects induced by both COR and SA elicitors, leading to differential changes in cell growth, taxane content, and secondary metabolism. Metabolite annotation revealed significant effects on N-containing compounds, phenylpropanoids, and terpenoids. Multivariate analysis showed that the metabolomic profiles of control and COR-treated samples are closer to each other than to SA-elicited samples at different time points (8, 16, and 24 days). The highest level of paclitaxel content was detected on day 8 under SA elicitation, exhibiting a negative correlation with the biomarkers kauralexin A2 and taxusin. Our study provides valuable insights into the intricate metabolic changes associated with paclitaxel production, aiding its potential optimization through untargeted metabolomics and an evaluation of COR/SA elicitor effects.

Keywords: biomarkers; coronatine; metabolomics; paclitaxel; salicylic acid; taxanes.

Figure 1

Time courses of biomass production and viability of T. baccata cell suspensions cultured for 24 days in the production medium in control conditions, or with the addition of 1 µM COR (coronatine) or 150 µM SA (salicylic acid). (a) FW in g/L. (b) DW expressed in g/L. (c) Percentage of cell viability. In all cases, the initial inoculum was of 300 g/L of fresh cells. Data represent average values from three separate experiments ± SD. Values marked by an asterisk (*) are significantly different (p ≤ 0.05) according to Tukey’s honestly significant difference test.

Figure 2

Taxane content under different treatments. (a) Total taxanes (expressed as mg/L) produced in Taxus baccata cell cultures throughout 24 days of culture in control conditions, with the addition of coronatine (COR) or salicylic acid (SA); the values are means ± SD (n = 6). (b) Taxane distribution is expressed as a percentage of individual taxane content.

Figure 3

(a) Principal component analysis (PCA) score plot illustrating the temporal tendencies of each treatment. Each arrow represents a specific behaviour of the samples. (b) Orthogonal partial least squares discriminant analysis (OPLS-DA) score plot depicting the differentiation between treatments. OPLS1 shows variation related to the classes or treatments covered by the model while OPLS2 represents the remaining variation orthogonal to the predictive component.

Figure 4

Plots of secondary metabolite biosynthesis in Taxus baccata cell suspensions maintained in the taxane production medium and elicited with COR 1 µM or SA 150 µM on days 8 (a), 16 (b), and 24 (c) of culturing. The large dots represent the average (mean) of all log fold changes (FCs) for metabolites and the small dots represent the individual log FCs for each metabolite within each family. NCCs: nitrogen-containing secondary compounds; phenylpropanoids and derivatives; S-containing: sulfur-containing secondary compounds.

Figure 5

The S-plot illustrates the relationship between the magnitude of change (p[1]) and the reliability of the measure (p(corr)[1]) for each variable (compound) in the model. (a) This plot was obtained from the Control vs. COR OPLS-DA model. (b) This plot was obtained from the Control vs. SA OPLS-DA model. The red rectangle highlights compounds that meet the selected magnitude (≥|0.1|) and reliability (≥|0.75|) criteria. Green solid circles represent the compounds.