Arabidopsis thaliana and other members of the Brassicaceae accumulate the hydroxycinnamic acid esters sinapoylmalate in leaves and sinapoylcholine in seeds. Our recent understanding of the phenylpropanoid pathway although complex has enabled us to perturb the sinapine biosynthesis pathway in plants. Sinapine (sinapoylcholine) is the most abundant antinutritional phenolic compound in seeds of cruciferous species and therefore is a target for elimination in canola (Brassica napus) meal. We analysed A. thaliana mutants with specific blocks in the phenylpropanoid pathway and identified mutant lines with significantly altered sinapine content. Knowledge gained from A. thaliana was extended to B. napus and the corresponding phenylpropanoid pathway genes were manipulated to disrupt sinapine biosynthesis in B. napus. Based on our understanding of the A. thaliana genetics, we have successfully developed transgenic B. napus lines with ferulic acid 5-hydroxylase (FAH) and sinapoylglucose:choline sinapoyltransferase (SCT)-antisense. These lines with concomitant downregulation of FAH and SCT showed up to 90% reduction in sinapine. In addition to reduced sinapine content, we detected higher levels of free choline accumulation in the seeds. These results indicate that it is possible to develop plants with low sinapine and higher choline by manipulating specific steps in the biosynthetic pathway. These improvements are important to add value to canola meal for livestock feed.

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Hydrocinnamic acid esters, lignin, flavonoids, glucosinolates, and salicylic acid protect plants against UV exposure, oxidative stress, diseases, and herbivores. Through the phenylpropanoid pathway, certain Brassicaceae family members, including Arabidopsis thaliana and Brassica napus, accumulate large amounts of the anti-nutritive sinapoylcholine (sinapine) in the seed. We successfully down-regulated activities of key enzymes in the pathway including F5H and SCT and achieved reduction of sinapine and lignin in B. napus seeds. Despite this success, it was unclear how multiple agronomic traits were affected in the transgenic plants. Here, we report altered large-scale gene expression of new alleles of f5h and sct mutants of A. thaliana and resultant accumulation of sinapoylglucose, disinapoylglucose, quercetin-3-O-rhamnoside, salicylic acid glucoside, and total indolyl glucosinolates in the two mutants. Expression of several flowering genes was altered in these mutants when grown under drought and NaCl treatments. Furthermore, both mutants were more susceptible to fungal infection than the wild type. Microarray experiments identified distinctive spatial and temporal expression patterns of gene clusters involved in silique/seed developmental processes and metabolite biosynthesis in these mutants. Taken together, these findings suggest that both f5h and sct mutants exhibit major differences in accumulation of diverse metabolites in the seed and profound changes in global large-scale gene expression, resulting in differential pleiotropic responses to environmental cues. Electronic supplementary material The online version of this article (doi:10.1007/s00425-009-1007-2) contains supplementary material, which is available to authorized users.

2009. Perturbation of lignin biosynthesis pathway in Brassica napus (canola) plants using RNAi. Can. J. Plant Sci. 89: 441Á453. Brassica napus meal contains high levels of lignin, which is one of the most important compositional factors affecting feed utilization by ruminants. We attempted to modify the concentration and composition of lignin in B. napus plants using the RNAi approach. Four genes were targeted for silencing by this approach either independently or in combination; caffeic acid O-methyltransferase (COMT), cinnamate 4-hydroxylase (C4H); coumarate 3-hydroxylase (C3H); ferulic acid 5-hydroxylase (F5H). We successfully developed transgenic B. napus lines expressing CaMV35S:C3H-C4H RNAi, CaMV35S:F5H-COMT RNAi, and Cruciferin:COMT RNAi that contained up to 40% less seed lignin in the transgenic seeds compared to the control. Despite successfully achieving suppression of these lignin biosynthesis genes and reduction in lignin content in B. napus seeds, we observed minor phenotypic effects on the transgenic plants. In lines carrying the cruciferin: COMT RNAi construct we observed a decrease in lignin content (40%) in the seed and anatomical variations when stem sections were examined. While our silencing had no major negative effect on plant growth it resulted in deformation of vessel elements, and minor changes in S-units. Taken together, these results clearly show that by employing RNAi strategy, it is possible to alter seed lignin content and composition in a manner non-detrimental to B. napus plants.. et Hannoufa, A. 2009. Alte´rationAlte´ration d'un circuit de labiosynthè se de la lignine chez le canola (Brassica napus) graˆcegraˆce a` l'ARNi. Can. J. Plant Sci. 89: 441Á453. Le tourteau de Brassica napus renferme beaucoup de lignine, un facteur qui, dans la composition des aliments, affecte conside´rablementconside´rablement son utilisation par les ruminants. Les auteurs ont tente´detente´de modifier la concentration et la composition de la lignine chez les plants de B. napus en recourant a` l'ARNi. Ils ont cible´quatrecible´quatregè nes pour court-circuiter, par leur action soit inde´pendanteinde´pendante, soit combineé, la production de lignine, en l'occurrence ceux de la O-me´thyltransfe´raseme´thyltransfe´me´thyltransfe´rase de l'acide cafeíque (COMT), de la cinnamate 4-hydroxylase (C4H), de la coumarate 3-hydroxylase (C3H) et de la 5-hydroxylase de l'acide fe´ruliquefe´rulique (F5H). Ils ont reússi a` creér des ligneés transge´niquestransge´niques de B. napus exprimant l'ARNi CaMV35S:C3H-C4H, l'ARNi CaMV35S:F5H-COMT et l'ARNi crucife´rinecrucife´rine:COMT dont les semences contenaient jusqu'a` 40 % moins de lignine que celles du cultivar te´mointe´moin. Bien qu'ils soient parvenus a` faire taire l'expression desgè nes codant labiosynthè se de la lignine et a` re´duirere´duire la concentration de cettederniè re dans les graines du canola, les auteurs ont releve´dereleve´de le´gersle´gers effets sur le phe´notypephe´notype des plantes transge´niquestransge´niques. Ainsi, chez les ligneés portant l...

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