Glucosinolate Uptake by Developing Rapeseed Embryos

Abstract

Embryos excised from the seed of rapeseed (Brassica napus L.) accumulated glucosinolate from the culture medium. Uptake was saturable, subject to inhibition, varied with the developmental stage of the embryo but correlated with the time of accumulation of glucosinolates In situ. It is suggested that a carrier-mediated transport system is operating in the developing embryo.During embryogenesis, glucosinolates accumulate in the seed of rapeseed (Brassica napus L). At maturity, the embryo comprises from 80 to 90% of the seed dry weight and is the site where most of the glucosinolates contained in the seed are located (7,14). The subcellular location of the glucosinolates within the embryo is not known for certain, although they are thought to be localized in globoid bodies associated with aleurone grains (17), structures which form within and eventually fill the vacuoles. Inheritance studies (7,14) and grafting experiments (9) have indicated that seed glucosinolate content is maternally controlled. Whether embryos have any synthetic capability, or simply act as a sink for glucosinolates synthesized in the pod or other plant parts, is not known.Immature rapeseed embryos germinate when removed from the seed if placed in nutrient medium or water (3). Precocious germination may be prevented by increasing the osmotic potential of the medium or by adding ABA (4, 5). Immature embryos cultured on high osmoticum develop normally in that they continue to synthesize and accumulate embryo-specific storage protein (4). This investigation presents evidence that immature rapeseed embryos cultured on high osmoticum do not synthesize and/or accumulate glucosinolates ifglucosinolates are absent from the culture medium, but can take up exogenously supplied glucosinolate in a carrier-mediated fashion. This ability to take up glucosinolate varies with development of the embryo, and may account for glucosinolate accumulation in situ. MATERIALS AND METHODS Plant MaterialsRapeseed (Brassica napus L. cv Golden) plants were grown in soil-free medium (15) Accumulation ExperimentsAccumulation of glucosinolate in situ was examined from 16 to 40 DAP2 by excising embryos from seeds at 2 to 4 d intervals. Accumulation of glucosinolates in vitro was examined by aseptically excising embryos from the seeds and culturing 10 per Petri plate (6 x 1.5 cm) in 5 mL of solid Monnier's culture medium (12) (filter sterilized, 0.4% agarose) according to the method of Crouch and Sussex (3) with and without the addition of 1.0 mm 2-propenyl-, 3-butenyl-, or benzylglucosinolate. Potassium salt of 2-propenylglucosinolate (sinigrin) was purchased from Sigma. The tetramethylammonium salts of 3-butenylglucosinolate and benzylglucosinolate were purified from the seeds of rapeseed (Brassica campestris L. cv R500) and nasturtium (Tropaeolum majus L.), respectively, by the method described by Hanley et al. (6). Each plate constituted a sample.Glucosinolate uptake by embryos over a 26 h period was examined by excising embryos from seeds, 10 to 20 per sample, an...

Biochemical Genetics of Plant Secondary Metabolites in Arabidopsis thaliana

“…The biosynthesis of seed glucosinolates could involve the transport of glucosinolate biosynthetic enzymes, the RNA encoding those enzymes, or glucosinolates themselves, from the maternal tissues to the embryo. Consistent with the latter hypothesis are recent physiological studies that have suggested that embryos of B. napus developing in vitro acquire their glucosinolates from the exogenous medium rather than synthesizing them de novo (5,7).…”

“…The effect of the gsml-l allele on shoot growth was tested as follows. Flats were planted with alternating rows of wild-type and TU1 plants and kept at 4°C for 7 . Glucosinolates in TUl shoots after feeding various compounds.…”

“…The w-methylthio glucosinolates (Table I; glucosinolates 1-6) may be derived from methionine by way of a chain-extension pathway (26) to produce a homologous series of amino acids (R = CH3S(CH2)n; n = 3-8) which, in turn, serve as precursors for glucosinolate biosynthesis. Additional modification of aliphatic side chains could generate the corresponding series of w-methylsulfinyl glucosinolates (Table I, [7][8][9][10][11][12], the benzoyloxy glucosinolates (Table I, 'Abbreviations: LC-MS, liquid chromatography-mass spectrometry; 2-A-5-MTP, 2-amino-5-methylthiopentanoic acid; 2-A-6-MTH, 2-amino-6-methylthiohexanoic acid; 2-A-7-MTH, 2-amino-7-methylthioheptanoic acid; 2-A-8-MTO, 2-amino-6-methylthiooctanoic acid; M2, progeny of the mutagenized generation; M3, two generations following mutagenesis. (1 ,umol) in the tip of a 1.5 mL microfuge (feeding) tube.…”

Metabolite profiling of Arabidopsis seedlings in response to exogenous sinalbin and sulfur deficiency

“…Up to now, however little was known about the influence of exogenous glucosinolates on plant morphology. A report by Gijzen et al (1989) found that the addition of 2-propenylglucosinolate to a growth medium containing rapeseed embryos resulted in a decline in embryo fr. wt.…”

Regulation of Isoflavonoid Biosynthesis in Soybean Seeds

“…Similarly glucosinolates are synthesized in leaves and translocated to seeds during development in Tropaeolus majus (Lykkesfeldt & Moller 1993). A carrier mediated system has been proposed for accumulation of glucosinolates in developing rapeseed embryos (Chen & Halkier 2000, Gijzen et al 1989. To investigate if similar pattern of transport and accumulation occurs in soybean seed isoflavonoids, the ability of soybean embryos to synthesize isoflavonoids were assessed and maternal effect on seed isoflavonoids was determined (Dhaubhadel et al 2003).…”