Modification of Brassica seed oil by antisense expression of a stearoyl-acyl carrier protein desaturase gene.

Abstract

Molecular gene transfer techniques have been used to engineer the fatty acid composition of Brassica rpa and Brassica napus (canola) oil. Stearoyl-acyl carrier protein (stearoyl-ACP) desaturase (EC 1.14.99.6) catalyzes the first desaturation step in seed oil biosynthesis, converting stearoyl-ACP to oleoyl-ACP. Seed-specific antisense gene constructs ofB. rapa stearoyl-ACP desaturase were used to reduce the protein concentration and enzyme activity of stearoyl-ACP desaturase in developing rapeseed embryos during storage lipid biosynthesis. The resulting transgenic plants showed dramatically increased stearate levels in the seeds. A continuous distribution of stearate levels from 2% to 40% was observed in seeds of a transgenic B. napus plant, illustrating the potential to engineer specialized seed oil compositions.Canola and other temperate vegetable oils are composed predominantly of unsaturated 18-carbon fatty acids: the monounsaturated oleic (18:1) and polyunsaturated linoleic (18:2) and linolenic (18:3) acids. In addition to these fatty acids, most oils also contain small but significant amounts of the saturated palmitic (16:0) and stearic (18:0) acids (1). The plastid-localized enzyme stearoyl-acyl carrier protein (stearoyl-ACP) desaturase (EC 1.14.99.6) catalyzes the initial desaturation reaction in fatty acid biosynthesis (Fig. 1A) and thus plays a key role in determining the ratio oftotal saturated to unsaturated fatty acids in plants (2,4,5).Specialized fatty acid compositions desired for edible and industrial purposes have been produced in oilseed crops through traditional breeding and selection alone or in combination with mutagenesis programs (6-9). Although the molecular basis for the changes is largely unknown, examples such as the removal of erucic acid from rapeseed oil to create canola (10), reduction of linolenic acid content in flax seed (11), and increases in stearate content of up to six times the wild-type level in safflower (up to 12% stearate) (12) and soybean (up to 30%o stearate) oil (13,14) demonstrate the plasticity of fatty acid composition in seed oil. It should also be possible to modify seed oil composition by the use of genetic engineering techniques (15-17). Antisense RNA technology has proven to be an effective means of reducing the level of specific enzymes in plants (18-21). Because fatty acid biosynthesis is an essential metabolic pathway in all tissues ofthe plant, modification of seed oil biosynthesis may require tissue-specific control of antisense RNA expression. Reduction of stearoyl-ACP desaturase in seeds should alter the ratio of saturated to unsaturated fatty acids and lead to the production of a novel storage oil without compromising the integrity of membrane lipids in leaf and other plant tissues.We report the isolation of a Brassica rapa (syn. Brassica campestris, turnip rape) stearoyl-ACP desaturase cDNAt and expression of antisense stearoyl-ACP desaturase constructs in seeds of B. rapa and Brassica napus. The activity and amount of stearoyl-ACP desaturase...

Transplastomic tobacco plants expressing a fatty acid desaturase gene exhibit altered fatty acid profiles and improved cold tolerance

“…The down-regulation of the expression of genes encoding stearoyl-ACP desaturases has been repeatedly reported and is associated with changes in fatty acid composition and alterations in various physiological traits and growth of mutants or transgenic plants (Knutzon et al 1992;Lightner et al 1994a, b;Merlo et al 1998;Kachroo et al 2001;Nandi et al 2003). However, there are relatively few data concerning up-regulation of the same genes.…”

Acyl-lipid desaturases and their importance in the tolerance and acclimatization to cold of cyanobacteria

“…It was purified to homogeneity from avocado mesocarp [59] and developing embryos of safflower seed [60]. cDNAs encoding stearoyl-ACP desaturases were isolated from castor bean [59,61], safflower seeds [60], cucumber [62], spinach [63], B. rapa [64] and B. napus [65]. cDNA encoding the 16:0-ACP desaturase that is involved in the biosynthesis ofpetroselinic acid was isolated from coriander [66,67].…”

Strategies for Modifying Fatty Acid Composition in Transgenic Plants

“…In some cases the total amount of seed oil was reduced and germination was impaired. However, some B. napus lines that produced seed TAGs with up to 40% stearic acid germinated normally and contained normal quantities of seed oil (Knutzon et al, 1992).…”