Characterization of Seed Oil Bodies and Their Surface Oleosin Isoforms from Rice Embryos

Abstract

Plant seeds store triacylglycerols in discrete organelles called oil bodies. An oil body stores a matrix of triacylglycerols surrounded by phospholipids and alkaline proteins termed oleosins. Oil bodies in rice seeds are present in embryos and aleurone layers. They do not coalesce in crowded environments, as observed on electron microscopy. The detected isoelectric point of purified rice oil bodies is pH 6.2. This implies that rice oil bodies possess a negatively charged surface at neutral pH. The suspension of rice oil bodies in pH 6.5 buffer induces aggregation. Presumably, the negatively charged surface causes electrostatic repulsion that maintains rice oil bodies as discrete organelles. Rice oil bodies lose their integrity on trypsin treatment. Undoubtedly, oleosins play an important role in the stability of oil bodies. There are two oleosin isoforms in rice oil bodies. Antibodies raised against these two homologous isoforms do not cross-recognize each other. Both isoforms are restricted to oil bodies, as detected on immuno-assaying. Partial amino acid sequences of these two isoforms were obtained, and compared with the deduced sequences of two maize and two rice oleosin genes. The comparison confirmed that the two major proteins in rice oil bodies are the two oleosin isoforms.

Gene Family of Oleosin Isoforms and Their Structural Stabilization in Sesame Seed Oil Bodies

“…In several species, the diŠerence in molecular mass between H-oleosin and L-oleosin is 2 kDa: maize has 18-and 16-kDa oleosins; 8) Arabidopsis thaliana has 21-and 19-kDa oleosins; 24,25) barley has 18-and 16-kDa oleosins; 26) rice has 18-and 16-kDa oleosins; 27) and sesame has 17-and 15-kDa oleosins (this report). Presumably, the diŠerence is constant because of the insertion of 18 residues (¿2 kDa) in the C-terminal domain of the H-oleosins.…”

Integral Proteins in Plant Oil Bodies

“…Right after the kick-off studies on maize oleosins, homologous oleosin isoforms were subsequently identified in oil bodies of rapeseed (Brassica napus), soybean (Glycine max L.), carrot (Daucus carota), sunflower (Helianthus annuus), Arabidopsis thaliana, and cotton (Gossypium hirsutum) with their corresponding cDNA fragments cloned [27][28][29][30][31][32][33][34]. It seems that oleosin isoforms are universally present in oil bodies of angiosperm seeds including both monocotyledonous and dicotyledonous species [35][36][37][38][39]. Furthermore, oleosin isoforms found as the major proteins (approximately 80-90%) in seed oil bodies were demonstrated to shield the whole surface of the organelles in the company of phospholipids [40].…”

Oleosin Is Bifunctional Enzyme That Has Both Monoacylglycerol Acyltransferase and Phospholipase Activities

“…The central long hydrophobic core has a unique proline knot, PX 5 SPX 3 P, that is conserved across various species and is used for targeting the protein to oleosomes (9, 10) but is not required for integration into the membrane of the endoplasmic reticulum. A mutation in the conserved residues of the proline knot resulted in inefficient targeting to the oil body; however, oleosin maintained its normal level in seeds (11,12). Oleosins are not only restricted to oil bodies but are also expressed specifically in the floral tapetum cells of the pollen tube in Arabidopsis (13).…”

“…The extract was filtered through two layers of cheesecloth, and the filtrate was differentially centrifuged to fractionate the intracellular components. The oleosomes were prepared as described previously (12).…”