The effects of Fusarium graminearum cell extracts and culture filtrates on the production of paclitaxel and 10-deacetylbaccatin III in suspension cell cultures of Taxus baccata L

Abstract

Background: The genus Taxus (yew) is the sole producer of the paclitaxel with anticancer properties. The rising demand for plant-derived medicines has led to the overexploitation of various species and ecosystem degradation, which is further worsened by climate change. Taxus baccata L. cell culture represents a promising commercial approach for the production of taxanes. A variety of elicitors and signaling molecules have been utilized to enhance production taxanes with results significantly affected by several factors, including the specificity of the stimulant, its concentration, the timing of inoculation, and the duration of treatment. To date, no studies have revealed that the elicitors derived from Fusarium graminearum enhance 10-deacetylbaccatin III (10-DABIII) and paclitaxel production in T. baccata suspension cell cultures. Added to that, we evaluated the impact of these fungal elicitors on cell viability, growth, phenylalanine ammonia-lyase (PAL) activity, and the production levels of paclitaxel and 10-DABIII.

Results: We investigated the effects of autoclaved cell extracts (ACE) and autoclaved culture filtrates (ACF) derived from F. graminearum on the suspension cell cultures of T. baccata. Our results indicated that the highest paclitaxel production, 9.438 µg/g dry weight, was achieved with a 10% autoclaved culture filtrate treatment. Furthermore, the autoclaved cell extracts significantly enhanced the levels of 10-DABIII, resulting in a remarkable 7.38-fold increase at a 5% concentration compared to the control on day 21. Treatment of the cell cultures of T. baccata with ACE and ACF decreased cell viability by 31% and 23%, respectively, compared to an 18% reduction observed in the control group after 21 days. The fungal elicitors initially induced the activity of PAL in the cell cultures, followed by a subsequent decline in this enzymatic activity.

Conclusions: Our study offers valuable insights for biotechnological applications in pharmaceutical and agricultural industries. Moreover, this research contributes to a better understanding of elicitor-mediated improvements in paclitaxel biosynthesis, paving the way for sustainable and efficient production in T. baccata cell cultures.

Keywords: Fusarium graminearum; Taxus baccata; 10-DABIII; Cell extracts; Culture filtrates; Paclitaxel; Suspension cell culture.

Fig. 1

The schematic highlights the interaction between fungal pathogens and plants, emphasizing the defense strategies employed by plants to combat these pathogens and the connection to the production of paclitaxel and other taxanes

Fig. 2

Callus cultures of T. baccata. A Explants cultured on Gamborg medium (B5). B The calli appeared after 30 days on callus induction media

Fig. 3

The experimental design used for elicitation of T. baccata cell culture. Elicitation was performed at different time points (7, 14 and 21 days). Different concentrations (2.5, 5, 10% v/v) of autoclaved culture filtrates (ACF) and autoclaved cell extracts (ACE) were used individually for treatment of T. baccata cell culture. Four biological replicates (n = 4) for each group of treatments were used (each flask represents a separate biological replicate)

Fig. 4

A The sub-cultured calli. B The calli used for the preparation of cell suspension. C Cell suspension prepared for elicitation

Fig. 5

The growth curve and biomass production of T. baccata cell suspensions cultured for 28 days in the production medium in control conditions. A Cells dry weight (g/100 mL). B Cells fresh weight (g/100 mL). The values are presented as mean ± standard error (SE) of 4 replications (n = 4)

Fig. 6

Percentage of cell viability of T. baccata under different treatments. Different concentrations of autoclaved culture filtrates (ACF) and autoclaved cell extracts (ACE) were used as elicitors. The data expressed as the average mean ± standard deviation of 4 replications (n = 4) The values followed by different letters are significantly difference (P < 0.05) according to Duncan’s multiple-range tests

Fig. 7

Time course biomass production of T. baccata cell suspensions cultured for 21 days in the treated and control conditions. A Growth expressed as gram of fresh weight (FW) per 100 milliliters. B Biomass expressed as gram of dry weight (DW) per 100 milliliters

Fig. 8

Effect of utilizing culture filtrate and cell extract of F. graminearum, on PAL activity in T. baccata cells. Different concentrations of autoclaved culture filtrates (ACF) and autoclaved cell extracts (ACE) were used as elicitors. The values are presented as mean ± standard error (SE) of 4 replications (n = 4). The values followed by different letters are significantly difference (P < 0.05) according to Duncan’s multiple-range tests

Fig. 9

Effects of the elicitors on taxanes production in T. baccata cell culture treated with fungal elicitors in different time points is illustrated. A The content of paclitaxel (µg/g DW). B The content of 10-deacetyl baccatin III (10-DAB III) (µg/g DW). The values are presented as mean ± standard error (SE). The values followed by different letters are significantly difference (P < 0.05) according to Duncan’s multiple-range tests

Fig. 10

Heatmap of the multiple data sets analyzed for each treatment. The distribution and intensity of the measured data, including PAL activities, concentrations of paclitaxel and 10-DABIII, as well as viability and dry weight, across different treatments are presented