Paclitaxel production and PAL activity in hairy root cultures of Taxus x media var. Hicksii carrying a taxadiene synthase transgene elicited with nitric oxide and methyl jasmonate

K Sykłowska-Baranek,M Bonfill,M R Naliwajski,M Grech-Baran,A Pietrosiuk

doi:10.1007/s11738-015-1949-x

K Sykłowska-Baranek, M Bonfill + Show 3 more

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https://doi.org/10.1007/s11738-015-1949-x

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Abstract

The role of sodium nitroprusside (SNP, 10 μM), a nitric oxide (NO) donor, and/or methyl jasmonate (MJ, 100 μM) spiked with l-phenylalanine (PHEN, 100 μM) and additional sucrose (S; 30 g l−1), in taxane production and phenyl ammonia lyase (PAL) activity in cultures of two Taxus media x var. Hicksii transgenic root lines (ATMA and ATM) carrying the taxadiene synthase transgene was investigated. SNP addition, when applied together with MJ and/or PHEN, resulted in paclitaxel production only in ATMA cultures. The application of the NO donor gave the highest paclitaxel content (7.56 mg l−1) in the combination of SNP+S+MJ+PHEN, after 2 weeks of treatment in the ATMA root line. In ATM cultures, taxane production was not affected by SNP. In both ATMA and ATM lines the highest total (intra+extracellular) paclitaxel yield was determined when elicited with MJ+PHEN, and amounted to 10.78 mg l−1 at 1 week and 1.63 mg l−1 at 2 weeks of treatment, in cultures of ATMA and ATM lines, respectively. The excretion of paclitaxel was observed only in ATMA cultures, with the highest level (2.34 mg l−1) obtained after elicitation with S+MJ+PHEN. The comparison of PAL activity in the two root lines revealed that this enzyme was almost 3-times more active in ATM than ATMA roots. An increase in both PAL activity and paclitaxel production was only observed in ATMA cultures growing in medium supplemented with S+MJ+PHEN.

Highlights

Paclitaxel is the active agent of TaxolÒ (Bristol-Myers Squibb), one of the most effective anticancer plantderived drugs
The growth of two transgenic root lines carrying the txs transgene was investigated, comparing control cultures with cultures supplemented at day 28 with a precursor (PHE), additional sucrose (S) and elicitor/s methyl jasmonate (MJ) and/or SNP
In control cultures of ATMA and ATM roots, the highest fresh biomass increase was noted at the end of the Taxane production and phenyl ammonia lyase (PAL) activity

Summary

Introduction

Paclitaxel is the active agent of TaxolÒ (Bristol-Myers Squibb), one of the most effective anticancer plantderived drugs. Paclitaxel is a first line treatment chosen for ovary, breast and lung cancers, but is used in combination therapies (Fauzee 2011). Paclitaxel has shown great promise as a locally delivered antirestenotic agent and seems to be an effective compound for the treatment of Alzheimer’s and other neurodegenerative diseases (Nims et al 2006; Li et al 2003; Fu et al 2009). A very promising approach for the production of paclitaxel and related taxanes, without forest harvesting, is provided by plant biotechnology approaches, and taxane production has been recently successfully scaled up (Frense 2007). Medium supplementation with elicitor/s and/ or precursors has proved to be the most effective

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