Abscisic acid induction of elongase activity, biosynthesis and accumulation of very long chain monounsaturated fatty acids and oil body proteins in microspore-derived embryos of Brassica napus L. cv Reston

Abstract

Microspore-derived (MD) embryos of Brassica napus cv Reston, were used as a model to test the effect of abscisic acid (ABA) on the accumulation of very long chain monounsaturated fatty acids (VLCMFAs) and oil body proteins during embryogenesis. Developing early cotyledonary MD embryos (13–16 days in culture) were treated with 10 μM ABA + 0.05%DMSO (+ ABA) or 0.05%DMSO only (control), for 3 days, transferred to fresh culture medium (without ABA) 5–8 days and then examined at 24 days in culture for effects on fatty acid and oil body protein biosynthesis. ABA-treated embryos had a total fatty acid content 40% higher on a mg dry weight basis and 50% higher on a mg protein basis, than the control treatment. In particular, the levels of eicosenic (20:1) and erucic (22:1) acids in the total lipid extract and especially in triacylglycerols (TAGs), increased 3–4-fold after ABA treatment compared to controls. The ABA effect was shown to be related to an increased specific activity of elongase(s) assayed in vitro and resulted in the accumulation of TAGs containing a greater proportion of VLCMFAs as demonstrated by direct probe mass spectrometry. Time-course experiments revealed that the first 72 h of ABA treatment were critical in the induction phenomenon. The net effect of the ABA treatment was the accumulation of VLCMFAs at a point earlier in embryo development than normally observed (control). De novo biosynthesis of oil body proteins (oleosins) was similarly affected by treatment with ABA and suggested that the initial 3-day ABA treatment induced more mRNA and/or resulted in a more stable oleosin mRNA. Western analysis showed an increased mass of oleosin as a proportion of total protein in the ABA-treated samples, first clearly discernible at 24 h and strongly evident by 72 and 168 h. The temporal relatedness of VLCMFA accumulation and oleosin biosynthesis as affected by ABA in microspore-derived embryos of B. napus, suggest that the latter may be useful system to gain insights into the ontogeny of oil bodies and the biosynthesis of developmentally-associated lipids under phytohormonal regulation.