| dc.description.abstract | Taxus L. is a genus of trees and shrubs with high value in horticulture,and in medicine as a source of the anticancer drug Paclitaxel. The taxonomy of the group is highly complex due to the lack of diagnostic morphological characters and the high degree of phenotypic plasticity among species. This often leads to misidentification and problems with classification using traditional taxonomic methods that rely solely on morphological characters. The complexity has become an issue for the pharmaceutical industry because of the anti-cancer properties of Paclitaxel. Paclitaxel is a diterpenoid, which is produced by several species of Taxus and is used to treat various forms of cancer. Taxushas a wide global geographic distribution and some taxonomists recognise only a single species with geographically defined subgroups. However, others have preferred to split the genus into several species. To address these differences, we conducted a thorough phylogenetic analysis (Maximum Likelihood, Bayesian Inference and TCS haplotype network analyses) of all its species and many subtaxa using nuclear and plastid gene regions (nrITS and plastid trnL intron and trnL-Fintergenic spacer). Results support the recognition of nine distinct species (T. baccata,T. brevifolia, T. canadensis, T. cuspidata, T. floridana, T. fauna, T. globosa,T. sumatranaand T. wallichiana) but evidence is found for less species distinction and considerable reticulation within the T.baccata, T. canadensis and T. cuspidata group. There is some biogeographic structure in the nrITS data showing that Taxus brevifolia is sister to T. globosa and T. floridana. Taxus fauna groups with T. contorta. Taxus wallichiana is resolved as monophyletic but its varieties T. wallichiana var. mairei, var.chinensis and var.wallichianaare not monophyletic although individuals within varieties generally group well together. Evidence is also presented for the sister group status of Pseudotaxusto Taxusand the inclusion of Amentotaxus, Austrotaxus, Cephalotaxusand Torreyawithin Taxaceae. We compare the results to known taxonomy, and present preliminary new leaf anatomical data using leaf impressions to visualise epidermal and stomatal characters.We also investigate the hybrids T.xmediaand T. xhunnewelliana and present nrITSdata identifying the origin of these taxa and their putative parental species. The biosynthesis of Paclitaxel involves 19 steps. We characterised two genes, Taxadiene synthase (TS) and 10-deacetyl baccatin III-10-0-acetyltranferase (DBAT), involved in the
pathway and compared variation in the genes among species of Taxus. TS is involved in the first committed step of the pathway. We developed new molecular PCR primers to amplify and study the TS1 to 5 exon regions. The primary aim was to assess molecular variation in the TSgene at DNA and protein levels and to test for evidence of selection on the gene. However, we also sequenced a section of the DBATgene,for a sample of taxa, to look for evidence of molecular variation in that gene and its presence in relatives of Taxus. The substitution rate is high in the TS gene with on average 43 changes per kbp and there is considerable variation in the amino acid translation, which varies with exon. TS gene trees based on all codon positions are not consistent with the ITS and trnL-Ftreeshinting at selection acting on the gene. Our results show that there is much variation in the sequences of TS and DBATacross species, that many are non-synonymous and that these changes are possibly due to parallel changes caused by selection on the genes. These results bring more clarification to the taxonomy of Taxus and highlight the high levels of variation in Paclitaxel genes. It may provide the pharmaceutical industry with promising targets for genetically engineering more efficient biosynthetic production of Paclitaxel and its precursors. | en |
