The biochemistry of headgroup exchange during triacylglycerol synthesis in canola.

Many pathways of primary metabolism are substantially conserved within and across plant families. However, significant differences in organization and fluxes through a reaction network may occur, even between plants in closely related genera. Assessing and understanding these differences is key to appreciating metabolic diversity, and to attempts to engineer plant metabolism for higher crop yields and desired product profiles. To better understand lipid metabolism and seed oil synthesis in canola (Brassica napus), we have characterized four canola homologues of the Arabidopsis (Arabidopsis thaliana) ROD1 gene. AtROD1 encodes phosphatidylcholine:diacylglycerol cholinephosphotransferase (PDCT), the enzyme that catalyzes a major flux of polyunsaturated fatty acids (PUFAs) in oil synthesis. Assays in yeast indicated that only two of the canola genes, BnROD1.A3 and BnROD1.C3, encode active isozymes of PDCT, and these genes are strongly expressed during the period of seed oil synthesis. Loss of expression of BnROD1.A3 and BnROD1.C3 in a double mutant, or by RNA interference, reduced the PUFA content of the oil to 26.6% compared with 32.5% in the wild type. These results indicate that ROD1 isozymes in canola are responsible for less than 20% of the PUFAs that accumulate in the seed oil compared with 40% in Arabidopsis. Our results demonstrate the care needed when translating results from a model species to crop plants.

Identification, characterization and field testing of Brassica napus mutants producing high-oleic oils.

Producing healthy, high-oleic oils and eliminating trans-fatty acids from foods are two goals that can be addressed by reducing activity of the oleate desaturase, FAD2, in oilseeds. However, it is essential to understand the consequences of reducing FAD2 activity on the metabolism, cell biology and physiology of oilseed crop plants. Here, we translate knowledge from studies of fad2 mutants in Arabidopsis (Arabidopsis thaliana) to investigate the limits of non-GMO approaches to maximize oleic acid in the seed oil of canola (Brassica napus), a species that expresses three active FAD2 isozymes. A series of hypomorphic and null mutations in the FAD2.A5 isoform were characterized in yeast (Saccharomyes cerevisiae). Then, four of these were combined with null mutations in the other two isozymes, FAD2.C5 and FAD2.C1. The resulting mutant lines contained 71-87% oleic acid in their seed oil, compared with 62% in wild-type controls. All the mutant lines grew well in a greenhouse, but in field experiments we observed a clear demarcation in plant performance. Mutant lines containing less than 80% oleate in the seed oil were indistinguishable from wild-type controls in growth parameters and seed oil content. By contrast, lines with more than 80% oleate in the seed oil had significantly lower seedling establishment and vigor, delayed flowering and reduced plant height at maturity. These lines also had 7-11% reductions in seed oil content. Our results extend understanding of the B. napusFAD2 isozymes and define the practical limit to increasing oil oleate content in this crop species.

A novel isoform of sucrose synthase is targeted to the cell wall during secondary cell wall synthesis in cotton fiber.

Sucrose (Suc) synthase (Sus) is the major enzyme of Suc breakdown for cellulose biosynthesis in cotton (Gossypium hirsutum) fiber, an important source of fiber for the textile industry. This study examines the tissue-specific expression, relative abundance, and temporal expression of various Sus transcripts and proteins present in cotton. A novel isoform of Sus (SusC) is identified that is expressed at high levels during secondary cell wall synthesis in fiber and is present in the cell wall fraction. The phylogenetic relationships of the deduced amino acid sequences indicate two ancestral groups of Sus proteins predating the divergence of monocots and dicots and that SusC sequences form a distinct branch in the phylogeny within the dicot-specific clade. The subcellular location of the Sus isoforms is determined, and it is proposed that cell wall-localized SusC may provide UDP-glucose for cellulose and callose synthesis from extracellular sugars.