The Taxus genome provides insights into paclitaxel biosynthesis.

The ancient gymnosperm genus Taxus is the exclusive source of the anticancer drug paclitaxel, yet no reference genome sequences are available for comprehensively elucidating the paclitaxel biosynthesis pathway. We have completed a chromosome-level genome of Taxus chinensis var. mairei with a total length of 10.23 gigabases. Taxus shared an ancestral whole-genome duplication with the coniferophyte lineage and underwent distinct transposon evolution. We discovered a unique physical and functional grouping of CYP725As (cytochrome P450) in the Taxus genome for paclitaxel biosynthesis. We also identified a gene cluster for taxadiene biosynthesis, which was formed mainly by gene duplications. This study will facilitate the elucidation of paclitaxel biosynthesis and unleash the biotechnological potential of Taxus.

Recent Research Progress in Taxol Biosynthetic Pathway and Acylation Reactions Mediated by Taxus Acyltransferases.

Taxol is one of the most effective anticancer drugs in the world that is widely used in the treatments of breast, lung and ovarian cancer. The elucidation of the taxol biosynthetic pathway is the key to solve the problem of taxol supply. So far, the taxol biosynthetic pathway has been reported to require an estimated 20 steps of enzymatic reactions, and sixteen enzymes involved in the taxol pathway have been well characterized, including a novel taxane-10ß-hydroxylase (T10ßOH) and a newly putative ß-phenylalanyl-CoA ligase (PCL). Moreover, the source and formation of the taxane core and the details of the downstream synthetic pathway have been basically depicted, while the modification of the core taxane skeleton has not been fully reported, mainly concerning the developments from diol intermediates to 2-debenzoyltaxane. The acylation reaction mediated by specialized Taxus BAHD family acyltransferases (ACTs) is recognized as one of the most important steps in the modification of core taxane skeleton that contribute to the increase of taxol yield. Recently, the influence of acylation on the functional and structural diversity of taxanes has also been continuously revealed. This review summarizes the latest research advances of the taxol biosynthetic pathway and systematically discusses the acylation reactions supported by Taxus ACTs. The underlying mechanism could improve the understanding of taxol biosynthesis, and provide a theoretical basis for the mass production of taxol.

Phyto-miRNAs-based regulation of metabolites biosynthesis in medicinal plants.

Medicinal plants, are known to produce a wide range of plant secondary metabolites (PSMs) applied as insecticides, drugs, dyes and toxins in agriculture, medicine, industry and bio-warfare plus bio-terrorism, respectively. However, production of PSMs is usually in small quantities, so we need to find novel ways to increase both quantity and quality of them. Fortunately, biotechnology suggests several options through which secondary metabolism in plants can be engineered in innovative ways to: 1) over-produce the useful metabolites, 2) down-produce the toxic metabolites, 3) produce the new metabolites. Among the ways, RNA interference (RNAi) technology which involves gene-specific regulation by small non-coding RNAs (sncRNAs) have been recently emerged as a promising tool for plant biotechnologist, not only to decipher the function of plant genes, but also for development of the plants with improved and novel traits through manipulation of both desirable and undesirable genes. Among sncRNAs, miRNAs have been recorded various regulatory roles in plants such as development, signal transduction, response to environmental stresses, metabolism. Certainly, the use of miRNAs in metabolic engineering requires identification of miRNAs involved in metabolites biosynthesis, understanding of the biosynthetic pathways, as well as the identification of key points of the pathways in which the miRNAs have their own effect. Thus, we firstly consider these three issues on metabolic engineering of medicinal plants. Our review shows, application of miRNAs can open a novel perspective to metabolic engineering of medicinal plants.

Seed and pollen gene dispersal in Taxus baccata, a dioecious conifer in the face of strong population fragmentation.

Background and Aims: Dispersal is crucial due to its direct impact on dynamics of a species' distribution as well as having a role in shaping adaptive potential through gene flow. In plants forming scarce and small populations, knowledge about the dispersal process is required to assess the potential for colonizing new habitats and connectivity of present and future populations. This study aimed to assess dispersal potential in Taxus baccata, a dioecious gymnosperm tree with a wide but highly fragmented distribution. Methods: Seed and pollen dispersal kernels were estimated directly in the framework of the spatially explicit mating model, where genealogies of naturally established seedlings were reconstructed with the help of microsatellite markers. In this way, six differently shaped dispersal functions were compared. Key Results: Seed dispersal followed a leptokurtic distribution, with the Exponential-Power, the Power-law and Weibull being almost equally best-fitting models. The pollen dispersal kernel appeared to be more fat-tailed than the seed dispersal kernel, and the Lognormal and the Exponential-Power function showed the best fit. The rate of seed immigration from the background sources was not significantly different from the rate of pollen immigration (13.1 % vs. 19.7 %) and immigration rates were in agreement with or below maximum predictions based on the estimated dispersal kernels. Based on the multimodel approach, 95 % of seeds travel <109 m, while 95 % of pollen travels <704 m from the source. Conclusions: The results showed that, at a local spatial scale, yew seeds travel shorter distances than pollen, facilitating a rapid development of a kinship structure. At the landscape level, however, although yew exhibits some potential to colonize new habitats through seed dispersal, genetic connectivity between different yew remnants is strongly limited. Taking into account strong population fragmentation, the study suggests that gene dispersal may be a limiting factor of the adaptability of the species.

Relative strength of fine-scale spatial genetic structure in paternally vs biparentally inherited DNA in a dioecious plant depends on both sex proportions and pollen-to-seed dispersal ratio.

In plants, the spatial genetic structure (SGS) is shaped mainly by gene dispersal and effective population density. Among additional factors, the mode of DNA inheritance and dioecy influence SGS. However, their joint impact on SGS remains unclear, especially in the case of paternally inherited DNA. Using theoretical approximations and computer simulations, here we showed that the relative intensity of SGS measured in paternally and biparentally inherited DNA in a dioecious plant population depends on both the proportion of males and the pollen-to-seed dispersal ratio. As long as males do not prevail in a population, SGS is more intense in paternally than biparentally inherited DNA. When males prevail, the intensity of SGS in paternally vs biparentally inherited DNA depends on the compound effect of sex proportions and the pollen-to-seed dispersal ratio. To empirically validate our predictions, we used the case of Taxus baccata, a dioecious European tree. First, we showed that mitochondrial DNA (mtDNA) in T. baccata is predominantly (98%) paternally inherited. Subsequently, using nuclear DNA (nuDNA) and mitochondrial microsatellite data, we compared the fine-scale SGS intensity at both marker types in two natural populations. The population with equal sex proportions showed stronger SGS in mtDNA than in nuDNA. On the other hand, we found lower SGS intensity in mtDNA than in nuDNA in the population with 67% males. Thus, the empirical results provided good support for the theoretical predictions, suggesting that knowledge about SGS in paternally vs biparentally inherited DNA may provide insight into effective sex proportions within dioecious populations.

Response and Defense Mechanisms of Taxus chinensis Leaves Under UV-A Radiation are Revealed Using Comparative Proteomics and Metabolomics Analyses.

Taxus chinensis var. mairei is a species endemic to south-eastern China and one of the natural sources for the anticancer medicine paclitaxel. To investigate the molecular response and defense mechanisms of T. chinensis leaves to enhanced ultraviolet-A (UV-A) radiation, gel-free/label-free and gel-based proteomics and gas chromatography-mass spectrometry (GC-MS) analyses were performed. The transmission electron microscopy results indicated damage to the chloroplast under UV-A radiation. Proteomics analyses in leaves and chloroplasts showed that photosynthesis-, glycolysis-, secondary metabolism-, stress-, and protein synthesis-, degradation- and activation-related systems were mainly changed under UV-A radiation. Forty-seven PSII proteins and six PSI proteins were identified as being changed in leaves and chloroplasts under UV-A treatment. This indicated that PSII was more sensitive to UV-A than PSI as the target of UV-A light. Enhanced glycolysis, with four glycolysis-related key enzymes increased, provided precursors for secondary metabolism. The 1-deoxy-d-xylulose-5-phosphate reductoisomerase and 4-hydroxy-3-methylbut-2-enyl diphosphate reductase were identified as being significantly increased during UV-A radiation, which resulted in paclitaxel enhancement. Additionally, mRNA expression levels of genes involved in the paclitaxel biosynthetic pathway indicated a down-regulation under UV-A irradiation and up-regulation in dark incubation. These results reveal that a short-term high dose of UV-A radiation could stimulate the plant stress defense system and paclitaxel production.

Transcript profiling of jasmonate-elicited Taxus cells reveals a ß-phenylalanine-CoA ligase.

Plant cell cultures constitute eco-friendly biotechnological platforms for the production of plant secondary metabolites with pharmacological activities, as well as a suitable system for extending our knowledge of secondary metabolism. Despite the high added value of taxol and the importance of taxanes as anticancer compounds, several aspects of their biosynthesis remain unknown. In this work, a genomewide expression analysis of jasmonate-elicited Taxus baccata cell cultures by complementary DNA-amplified fragment length polymorphism (cDNA-AFLP) indicated a correlation between an extensive elicitor-induced genetic reprogramming and increased taxane production in the targeted cultures. Subsequent in silico analysis allowed us to identify 15 genes with a jasmonate-induced differential expression as putative candidates for genes encoding enzymes involved in five unknown steps of taxane biosynthesis. Among them, the TB768 gene showed a strong homology, including a very similar predicted 3D structure, with other genes previously reported to encode acyl-CoA ligases, thus suggesting a role in the formation of the taxol lateral chain. Functional analysis confirmed that the TB768 gene encodes an acyl-CoA ligase that localizes to the cytoplasm and is able to convert ß-phenylalanine, as well as coumaric acid, into their respective derivative CoA esters. ß-phenylalanyl-CoA is attached to baccatin III in one of the last steps of the taxol biosynthetic pathway. The identification of this gene will contribute to the establishment of sustainable taxol production systems through metabolic engineering or synthetic biology approaches.

[Cloning and functional characterization of a cDNA encoding isopentenyl diphosphate isomerase involved in taxol biosynthesis in Taxus media].

Taxol is one of the most potent anti-cancer agents, which is extracted from the plants of Taxus species. Isopentenyl diphosphate isomerase (IPI) catalyzes the reversible transformation between IPP and DMAPP, both of which are the general 5-carbon precursors for taxol biosynthesis. In the present study, a new gene encoding IPI was cloned from Taxus media (namely TmIPI with the GenBank Accession Number KP970677) for the first time. The full-length cDNA of TmIPI was 1 232 bps encoding a polypeptide with 233 amino acids, in which the conserved domain Nudix was found. Bioinformatic analysis indicated that the sequence of TmIPI was highly similar to those of other plant IPI proteins, and the phylogenetic analysis showed that there were two clades of plant IPI proteins, including IPIs of angiosperm plants and IPIs of gymnosperm plants. TmIPI belonged to the clade of gymnosperm plant IPIs, and this was consistent with the fact that Taxus media is a plant species of gymnosperm. Southern blotting analysis demonstrated that there was a gene family of IPI in Taxus media. Finally, functional verification was applied to identify the function of TmIPI. The results showed that biosynthesis of ß-carotenoid was enhanced by overexpressing TmIPI in the engineered E. coli strain, and this suggested that TmIPI might be a key gene involved in isoprenoid/terpenoid biosynthesis.

Taximin, a conserved plant-specific peptide is involved in the modulation of plant-specialized metabolism.

Small peptides play important roles in the signalling cascades that steer plant growth, development and defence, and often crosstalk with hormonal signalling. Thereby, they also modulate metabolism, including the production of bioactive molecules that are of high interest for human applications. Yew species (Taxus spp.) produce diterpenes such as the powerful anticancer agent paclitaxel, the biosynthesis of which can be stimulated by the hormone jasmonate, both in whole plants and cell suspension cultures. Here, we identified Taximin, as a gene encoding a hitherto unreported, plant-specific, small, cysteine-rich signalling peptide, through a transcriptome survey of jasmonate-elicited T. baccata suspension cells grown in two-media cultures. Taximin expression increased in a coordinated manner with that of paclitaxel biosynthesis genes. Tagged Taximin peptides were shown to enter the secretory system and localize to the plasma membrane. In agreement with this, the exogenous application of synthetic Taximin peptide variants could transiently modulate the biosynthesis of taxanes in T. baccata cell suspension cultures. Importantly, the Taximin peptide is widely conserved in the higher plant kingdom with a high degree of sequence conservation. Accordingly, Taximin overexpression could stimulate the production of nicotinic alkaloids in Nicotiana tabacum hairy root cultures in a synergistic manner with jasmonates. In contrast, no pronounced effects of Taximin overexpression on the specialized metabolism in Medicago truncatula roots were observed. This study increases our understanding of the regulation of Taxus diterpene biosynthesis in particular and plant metabolism in general. Ultimately, Taximin might increase the practical potential of metabolic engineering of medicinal plants.

Metabolic engineering approaches for production of biochemicals in food and medicinal plants.

Historically, plants are a vital source of nutrients and pharmaceuticals. Recent advances in metabolic engineering have made it possible to not only increase the concentration of desired compounds, but also introduce novel biosynthetic pathways to a variety of species, allowing for enhanced nutritional or commercial value. To improve metabolic engineering capabilities, new transformation techniques have been developed to allow for gene specific silencing strategies or stacking of multiple genes within the same region of the chromosome. The 'omics' era has provided a new resource for elucidation of uncharacterized biosynthetic pathways, enabling novel metabolic engineering approaches. These resources are now allowing for advanced metabolic engineering of plant production systems, as well as the synthesis of increasingly complex products in engineered microbial hosts. The status of current metabolic engineering efforts is highlighted for the in vitro production of paclitaxel and the in vivo production of ß-carotene in Golden Rice and other food crops.

[Research progress and trend analysis of biology and chemistry of Taxus medicinal resources].

Taxus is the source plant of anti-cancer drug paclitaxel and its biosynthetic precursor, analogs and derivatives, which has been studying for decades. There are many endemic Taxus species in China, which have been studied in the field of multiple disciplines. Based on the recent studies of the researchers, this review comments on the study of Taxus biology and chemistry. The bibliometric method is used to quantify the global scientific production of Taxus-related research, and identify patterns and tendencies of Taxus-related articles. Gaps are present in knowledge about the genomics, epigenomics, transcriptomics, proteomics, metabolomics and bioinformatics of Taxus and their endophytic fungi. Systems biology and various omics technologies will play an increasingly important role in the coming decades.

[Identification of EST-SSRs in Taxus cuspidata based on high-throughput sequencing].

OBJECTIVE: Taxus species are highly valued for the production of taxol. Based on high-throughput sequenceing, EST-SSRs were explored and studied in the transcriptome of Taxus cuspidata. METHOD: T cuspidata leaf cDNA was extracted and sequenced by 454 GS FLX Titanium. High-quality sequences were assembled using Newbler Assembler Software, which produced unique sequences. SSRs motif was explored using simple sequence repeat identification tool (Perl Script). Primers were designed using PRIMER3. RESULT: A total of 81 148 high-quality reads from the needles of T. cuspidata were produced using the Roche GS FLX Titanium system. A total of 20 557 unique sequences were obtained. There were 753 simple sequence repeat motifs identified. Primers of PCR were obtained for 519 EST-SSRs, randomly selected cloning sequencing revealed that 87.5% of ESTs were the same as the results of Sanger sequencing. CONCLUSION: The results provide the first EST-SSRs collection in Taxus and are essential for future efforts of gene discovery, functional genomics, and genome annotation in related species.

The first insight into the Taxus genome via fosmid library construction and end sequencing.

Taxus mairei is a critically endangered and commercially important cultured medicinal gymnosperm in China and forms an important medicinal resource, but the research of its genome is absent. In this study, we constructed a T. mairei fosmid library and analyzed the fosmid end sequences to provide a preliminary assessment of the genome. The library consists of one million clones with an average insert size of about 39 kb, amounting to 3.9 genome equivalents. Fosmid stability assays indicate that T. mairei DNA was stable during propagation in the fosmid system. End sequencing of both 5' and 3' ends of 968 individual clones generated 1,923 sequences after trimming, with an average sequence length of 839 bp. BLASTN searches of the nr and EST databases of GenBank and BLASTX searches of the nr database resulted in 560 (29.1%) significant hits (E < e(-5)). Repetitive sequences analysis revealed that 20.8% of end sequences are repetitive elements, which were composed of retroelements, DNA transposons, satellites, simple repeats, and low complexity sequences. The distribution pattern of various repeat types was found to be more similar to the gymnosperm Pinus and Picea than to the monocot and dicot. The satellites of T. mairei were significantly longer than those of P. taeda and P. glauca. The tetra-nucleotide repeats of T. mairei were much longer than those of P. glauca and P. taeda. The fosmid library and the fosmid end sequences, for the first time, will serve as a useful resource for large-scale genome sequencing, physical mapping, SSR marker development and positional cloning, and provide a better understanding of the Taxus genome.

Transcriptome analysis of Taxus cuspidata needles based on 454 pyrosequencing.

Taxus species are highly valued as renewable resources for the production of Taxol. Despite the commercial and medicinal importance of Taxus, little genomic information is available for yew species, and Taxol biosynthesis still needs to be fully elucidated. In this study, 454 pyrosequencing technology was employed to produce an expressed sequence tag (EST) from the needles of Taxus cuspidata. In all, 81 148 high-quality reads from the needles of T. cuspidata were produced using Roche GS FLX Titanium. A total of 20,557 unique sequences were obtained, including 12 975 singletons and 7582 contigs. Approximately 14,095 unique sequences were annotated by a similarity search against five public databases. Gene ontology revealed 11,220 unique sequences that could be assigned to 45 vocabularies. In the Kyoto Encyclopedia of Genes and Genomes mapping, 2403 transcripts were established as associated with 3821 biochemical pathways. Enzymes in the plastidial 2-C-methyl-D-erythritol 4-phosphate pathway were well represented. Candidates of the putative genes of Taxol biosynthesis were revealed, including those in the remaining steps. In total, 291 transcripts were identified, representing putative homologues of transcription factors. Furthermore, 753 simple sequence repeat motifs, which are potential molecular markers for genetic application, were identified. These results provide the largest EST collections in TAXUS and will contribute to biosynthetic and biochemical studies that lead to drug improvement.

Phylogenetic relationships of the genus Taxus inferred from chloroplast intergenic spacer and nuclear coding DNA.

A cladistic analysis of the medicinal plant Taxus, using the sequences of one chloroplast (trnS-trnQ spacer) and three nuclear taxadiene synthase (TS), 10-deacetylbaccatin III-10beta-O-acetyltransferase (DBAT), and 18S rDNA) molecular markers, was carried out by distance, parsimony, likelihood, and Bayesian methods. Three of the four New World species (T. brevifolia, T. floridana and T. globosa) form a well-supported clade, whereas T. canadensis initially branches-appearing distantly related to both Old World taxa and New World species. In Asia, Taxus chinensis, T. mairei, T. sumatrana and T. wallichiana cluster together and are sister to a clade containing T. baccata and T. contorta. Taxus yunnanensis is more closely related to T. wallichiana than to four other Taxus species in our study from China; T. contorta is closer to the Euro-Mediterranean T. baccata than to the Asian species. This study provides a genetic method for authentication of economically important Taxus species and proposes a robust phylogenetic hypothesis for the genus. Using trnS-trnQ spacer sequences, we were able to distinguish T. mairei from all other species of Taxus.

Molecular cloning and characterization of the yew gene encoding squalene synthase from Taxus cuspidata.

The enzyme squalene synthase (EC 2.5.1.21) catalyzes a reductive dimerization of two farnesyl diphosphate (FPP) molecules into squalene, a key precursor for the sterol and triterpene biosynthesis. A full-length cDNA encoding squalene synthase (designated as TcSqS) was isolated from Taxus cuspidata, a kind of important medicinal plants producing potent anti-cancer drug, taxol. The full-length cDNA of TcSqS was 1765 bp and contained a 1230 bp open reading frame (ORF) encoding a polypeptide of 409 amino acids. Bioinformatic analysis revealed that the deduced TcSqS protein had high similarity with other plant squalene synthases and a predicted crystal structure similar to other class I isoprenoid biosynthetic enzymes. Southern blot analysis revealed that there was one copy of TcSqS gene in the genome of T. cuspidata. Semiquantitative RT-PCR analysis and northern blotting analysis showed that TcSqS expressed constitutively in all tested tissues, with the highest expression in roots. The promoter region of TcSqS was also isolated by genomic walking and analysis showed that several cis-acting elements were present in the promoter region. The results of treatment experiments by different signaling components including methyl-jasmonate, salicylic acid and gibberellin revealed that the TcSqS expression level of treated cells had a prominent diversity to that of control, which was consistent with the prediction results of TcSqS promoter region in the PlantCARE database.

Identification of a cDNA clone specific for the taxol synthesis phase of Taxus chinensis cells by mRNA differential display.

In order to identify genes related to Taxol biosynthesis, the mRNA differential display method was employed to compare mRNA populations from suspension cultured Taxus chinensis cells before and after beginning to produce Taxol. From a total display of about 4000 PCR products, 104 were derived from cells during the Taxol synthesis phase but not the non-Taxol synthesis phase. These products were cloned, and one such cDNA clone, named TS1, was confirmed to be specifically expressed in the Taxol synthesis phase by northern blot analysis. The length of the transcript corresponding TS1 was approximately 2.1 kb. DNA sequencing and homology search showed the sequence of TS1 contains a partial open reading frame and has no homologies with other known genes. Hence, this report demonstrated the potential of mRNA differential display for the isolation of genes specific for the period of secondary metabolite production as well as the feasibility of the approach for the identification of genes potentially related to the synthesis of secondary metabolites in higher plants.

Phylogeny of taxaceae and cephalotaxaceae genera inferred from chloroplast matK gene and nuclear rDNA ITS region.

Phylogeny of the Taxaceae genera and the monotypic family Cephalotaxaceae has been extraordinarily controversial. In this paper chloroplast matK genes and nuclear ITS sequences were determined for all six genera of the two families and representatives of other conifer families. Analysis using either the nonsynonymous sites or the deduced amino acid sequences of matK genes strongly indicates that taxad genera and Cephalotaxaceae are monophyletic, with the Taxodiaceae/Cupressaceae clade as their sister group. Cephalotaxus is basal to the taxad genera, among which two clades, Torreya/Amentotaxus and Taxus/Pseudotaxus/Austrotaxus, are resolved. They correspond to Janchen's two tribes, Torreyeae and Taxeae. In Taxeae, Austrotaxus is the first to branch off. Analyses of the nuclear ITS sequence data corroborated the topology of the matK gene tree. These results refute the views that Cephalotaxaceae has no alliance with Taxaceae and that Austrotaxus and Amentotaxus should be excluded from the Taxaceae. We estimated the divergence time between the Taxodiaceae/Cupressaceae and the Cephalotaxaceae/Taxaceae clades to be 192-230 Myr ago and the divergence time between taxads and Cephalotaxus to be 149-179 Myr ago. Soon after the latter divergence event, within 6-8 Myr, the two taxad tribes originated. In conclusion, our data do not support Florin's claim that taxads could be traced to Devonian psilophytes (359-395 Myr ago).

Molecular cloning of a taxa-4(20),11(12)-dien-5alpha-ol-O-acetyl transferase cDNA from Taxus and functional expression in Escherichia coli.

The taxa-4(20),11(12)-dien-5alpha-ol-O-acetyl transferase which catalyzes the third step of Taxol biosynthesis has been isolated from methyl jasmonate-induced Taxus cells, and partially purified and characterized (K. Walker, R. E. B. Ketchum, M. Hezari, D. Gatfield, M. Golenowski, A. Barthol, and R. Croteau, Arch. Biochem. Biophys. 364, 273-279 1999). A revised purification method allowed internal amino acid microsequencing of the enzyme, from which primers were designed and employed to amplify a transacetylase gene-specific fragment. This radiolabeled, 900-bp amplicon was used as a hybridization probe to screen a cDNA library constructed from poly(A)(+) RNA isolated from induced Taxus cells, from which a full-length transacetylase sequence was obtained. Expression of this clone from pCWori(+) in Escherichia coli JM109 cells yielded the functional enzyme, as determined by radiochemical assay and combined capillary gas chromatographic-mass spectrometric verification of the acetylated product. The full-length DNA has an open-reading frame of 1317 nucleotides corresponding to a deduced amino acid sequence of 439 residues that exhibits high sequence identity to the proteolytic fragments of the native enzyme, which the recombinant transacetylase resembles in properties. Consistent with the size of the operationally soluble native enzyme, the DNA appears to encode a monomeric protein of molecular weight 49,079 that bears no N-terminal organellar targeting information. Sequence comparison of the taxadien-5alpha-ol-O-acetyl transferase with the few other known acyl transferases of plant origin indicates a significant degree of similarity between these enzymes (64-67%). The efficient conversion of taxadien-5alpha-yl acetate to further hydroxylated intermediates of the Taxol pathway confirms the significance of this acylation step and suggests this taxadienol transacetylase to be an important target for genetic manipulation to improve Taxol production.