Immunocytochemical and biochemical studies of the mobilisation of storage oil-bodies and proteins in germinating cotyledons of oilseed rape,Brassica napus

Seeds of Brassica napus L cv Mikado contain about 25% w/w protein in addition to 40–50% w/w storage oil. About 50% of the seed protein is the legumin-like neutral protein, cruciferin, and a further 20% is the small basic protein, napin. The only other major seed protein (20% of total) is a polypeptide of apparent molecular mass (M_r) 19 000±200, which is associated with the membranes of the storage oil-bodies. The purification of this protein and preparation of monospecific antibodies have recently been reported. The kinetics of protein and oil mobilisation and the subcellular distribution of the M_r 19 000 oil-body protein have been studied by techniques including sodium dodecyl sulphate-polyacrylamide gel electrophoresis (SDS-PAGE), gas-liquid chromatography (GLC), sucrose density gradient fractionation, and electron microscop-immunocytochemistry. The results show that the mobilisation of the storage products of rapeseed occurs in at least three distinct phases: (1) a lag phase of 10–15 h, (2) breakdown of cruciferin and napin from 12 h until day 3, (3) breakdown of storage oil and oil-body membranes from day 2 until day 7. The M_r 19000 protein was localised on oil-body membranes in early stages of germination but was later associated with a light membrane fraction, which probably contained oil-body ghosts. Relatively little difference in the kinetics of the mobilisation of storage oils and proteins was found whether seedlings were grown in the light or in the dark. The implications of these results for the mechanism of storage oil mobilisation in oilseeds are discussed.