Isolation and Characterization of the Lipid Reserve Bodies, Spherosomes, from Layers of Wheat

Abstract

Lipid reserve bodies (spherosomes) isolated from aleurone layers of wheat (Triticum aestivum L.) by combined differential and density gradient centrifugation were heterogeneous in size and density and distributed throughout continuous sucrose density gradients. The lightest spherosomes sedimented as floating lipid (d [density] < 1.0), and the heaviest spherosomes sedimented with a density near that for mitochondria (d > 1.18). While all classes of spherosomes were lipid-rich, the light spherosomes contained more triglycerides and the heavy spherosomes contained more phospholipids and protein. Acid phosphatase activity of aqueous homogenates occurred mainly in the microsome-free supernatant with only trace levels of the enzyme associated with spherosomes. Among the particulate fractions, only aleurone grains obtained by nonaqueous procedures or aleurone fragments had acid phosphatase activity in biochemical and cytochemical assays. Cytochemical analysis showed that the acid phosphatase activity in the spherosome fractions was not due to spherosomes per se but to adhering membrane fragments. In biochemical studies, acid lipase and phospholipase D activities paralleled the distribution of acid phosphatase among the cell fractions, demonstrating an initial association with aleurone grains rather than with spherosomes. Upon incubation with GA3 these enzymes disappeared from the aleurone grain fraction. Apparently spherosomes of wheat aleurone are not lysosomal in nature but cellular repositories of lipids. Aleurone grains appear to function as the main repository of hydrolytic enzymes in ungerminated caryopsis.