Application of Priming Strategy for Enhanced Paclitaxel Biosynthesis in Taxus × Media Hairy Root Cultures

Sykłowska-Baranek, Katarzyna; Sygitowicz, Grażyna; Maciejak-Jastrzębska, Agata; Pietrosiuk, Agnieszka; Szakiel, Anna

doi:10.3390/cells11132062

Cited by 9 publications

(8 citation statements)

References 41 publications

Supporting

Mentioning

Contrasting

Order By: Relevance

“…Additionally, since hairy roots are widely employed in many plants, especially species recalcitrant to stable genetic transformation, such as Rehmannia glutinosa [ 23 ], Panax ginseng [ 24 ], Cicer arietinum [ 25 ], Trachyspermum ammi [ 26 ], Rubia yunnanensis [ 27 ], Taxus spp. [ 28 ], Trigonella foenum-graecum [ 29 ], the eR&T is expected to be modified for application in these plants. For example, in vitro cultures of plants, including hairy roots, have been widely used in biotechnology to obtain high-value plant-derived products with a variety of biological and pharmacological activities [ 30 , 31 , 32 ].…”

Section: Discussionmentioning

confidence: 99%

See 1 more Smart Citation

An Efficient Agrobacterium rhizogenes-Mediated Hairy Root Transformation Method in a Soybean Root Biology Study

Huang

et al. 2022

IJMS

View full text Add to dashboard Cite

The stable genetic transformation of soybean is time-consuming and inefficient. As a simple and practical alternative method, hairy root transformation mediated by Agrobacterium rhizogenes is widely applied in studying root-specific processes, nodulation, biochemical and molecular functions of genes of interest, gene editing efficiency of CRISPR/Cas9, and biological reactors and producers. Therefore, many laboratories have developed unique protocols to obtain hairy roots in composite plants composed of transgenic roots and wild-type shoots. However, these protocols still suffer from the shortcomings of low efficiency and time, space, and cost consumption. To address this issue, we developed a new protocol efficient regeneration and transformation of hairy roots (eR&T) in soybean, by integrating and optimizing the main current methods to achieve high efficiency in both hairy root regeneration and transformation within a shorter period and using less space. By this eR&T method, we obtained 100% regeneration of hairy roots for all explants, with an average 63.7% of transformation frequency, which promoted the simultaneous and comparative analysis of the function of several genes. The eR&T was experimentally verified Promoter:GUS reporters, protein subcellular localization, and CRISPR/Cas9 gene editing experiments. Employing this approach, we identified several novel potential regulators of nodulation, and nucleoporins of the Nup107-160 sub-complex, which showed development-dependent and tissue-dependent expression patterns, indicating their important roles in nodulation in soybean. Thus, the new eR&T method is an efficient and economical approach for investigating not only root and nodule biology, but also gene function.

show abstract

Section: Discussionmentioning

confidence: 99%

“…Despite great progress in biotechnological approaches to paclitaxel production, the productivity of Taxus spp. in vitro cultures remains a challenge [ 28 ]. Paclitaxel has become a widely used anticancer drug for the treatment of various malignancies, including breast and ovarian cancers and Kaposi’s sarcoma [ 33 , 34 ].…”

Section: Discussionmentioning

confidence: 99%

An Efficient Agrobacterium rhizogenes-Mediated Hairy Root Transformation Method in a Soybean Root Biology Study

Huang

et al. 2022

IJMS

View full text Add to dashboard Cite

show abstract

“… Sykłowska-Baranek et al. (2022) recently described an efficient strategy that clearly increased PTX production of T. x media hairy roots.…”

Section: Progress In the Genetic Transformation Of Taxus ...mentioning

confidence: 99%

Genetic approaches in improving biotechnological production of taxanes: An update

et al. 2023

View full text Add to dashboard Cite

Paclitaxel (PTX) and its derivatives are diterpene alkaloids widely used as chemotherapeutic agents in the treatment of various types of cancer. Due to the scarcity of PTX in nature, its production in cell cultures and plant organs is a major challenge for plant biotechnology. Although significant advances have been made in this field through the development of metabolic engineering and synthetic biology techniques, production levels remain insufficient to meet the current market demand for these powerful anticancer drugs. A key stumbling block is the difficulty of genetically transforming the gymnosperm Taxus spp. This review focuses on the progress made in improving taxane production through genetic engineering techniques. These include the overexpression of limiting genes in the taxane biosynthetic pathway and transcription factors involved in its regulation in Taxus spp. cell cultures and transformed roots, as well as the development and optimization of transformation techniques. Attempts to produce taxanes in heterologous organisms such as bacteria and yeasts are also described. Although promising results have been reported, the transfer of the entire PTX metabolic route has not been possible to date, and taxane biosynthesis is still restricted to Taxus cells and some endophytic fungi. The development of a synthetic organism other than Taxus cells capable of biotechnologically producing PTX will probably have to wait until the complete elucidation of its metabolic pathway.

show abstract

“…Root culture experiments were carried out as in Syklowska-Baranek et al (2022) with slight variations. Briefly, transgenic L9 and wild type (untransformed) L4 roots were cultivated on a sterile rounded stainless steel grid in 250 mL Erlenmeyer flasks containing 30 mL of hormone-free modified liquid DCR medium (Gupta and Durzan, 1985;Syklowska-Baranek et al, 2009) and routinely subcultured every 4 weeks.…”

Section: Root Cultures and Elicitation Treatmentsmentioning

confidence: 99%

Overexpression of BAPT and DBTNBT genes in Taxus baccata in vitro cultures to enhance the biotechnological production of paclitaxel

Perez‐Matas,

Hidalgo‐Martinez,

Moyano

et al. 2023

Plant Biotechnology Journal

View full text Add to dashboard Cite

SummaryPaclitaxel is one of the most effective anticancer drugs ever developed. Although the most sustainable approach to its production is provided by plant cell cultures, the yield is limited by bottleneck enzymes in the taxane biosynthetic pathway: baccatin‐aminophenylpropanoyl‐13‐O‐transferase (BAPT) and 3′‐N‐debenzoyltaxol N‐benzoyltransferase (DBTNBT). With the aim of enhancing paclitaxel production by overcoming this bottleneck, we obtained distinct lines of Taxus baccata in vitro roots, each independently overexpressing either of the two flux‐limiting genes, BAPT or DBTNBT, through a Rhizobium rhizogenes A4‐mediated transformation. Due to the slow growth rate of the transgenic Taxus roots, they were dedifferentiated to obtain callus lines and establish cell suspensions. The transgenic cells were cultured in a two‐stage system and stimulated for taxane production by a dual elicitation treatment with 1 μm coronatine plus 50 mm of randomly methylated‐β‐cyclodextrins. A high overexpression of BAPT (59.72‐fold higher at 48 h) and DBTNBT (61.93‐fold higher at 72 h) genes was observed in the transgenic cell cultures, as well as an improved taxane production. Compared to the wild type line (71.01 mg/L), the DBTNBT line produced more than four times higher amounts of paclitaxel (310 mg/L), while the content of this taxane was almost doubled in the BAPT line (135 mg/L). A transcriptional profiling of taxane biosynthetic genes revealed that GGPPS, TXS and DBAT genes were the most reactive to DBTNBT overexpression and the dual elicitation, their expression increasing gradually and constantly. The same genes exhibited a pattern of isolated peaks of expression in the elicited BAPT‐overexpressing line.

show abstract

Application of Priming Strategy for Enhanced Paclitaxel Biosynthesis in Taxus × Media Hairy Root Cultures

Cited by 9 publications

References 41 publications

An Efficient Agrobacterium rhizogenes-Mediated Hairy Root Transformation Method in a Soybean Root Biology Study

An Efficient Agrobacterium rhizogenes-Mediated Hairy Root Transformation Method in a Soybean Root Biology Study

Genetic approaches in improving biotechnological production of taxanes: An update

Overexpression of BAPT and DBTNBT genes in Taxus baccata in vitro cultures to enhance the biotechnological production of paclitaxel

Contact Info

Product

Resources

About