The properties of a discrete membranous fraction isolated on sucrose gradients from castor bean endosperm have been examined . This fraction was previously shown to be the exclusive site of phosphorylcholine-glyceride transferase . The distribution of NADPH-cytochrome c reductase and antimycin insensitive NADH-cytochrome c reductase across the gradient followed closely that of the phosphorylcholine-glyceride transferase . This fraction also had NADH diaphorase activity and contained cytochromes b5 and P 450. On sucrose gradients containing 1 mM EDTA this fraction had a mean isopycnic density of 1 .12 g/cma and sedimented separately from the ribosomes ; electron micrographs showed that it was comprised of smooth membranes . When magnesium was included in the gradients to prevent the dissociation of membrane-bound ribosomes, the isopycnic density of the membrane fraction with its associated enzymes was increased to 1 .16 g/cma and under these conditions the electron micrographs showed that the membranes had the typical appearance of rough endoplasmic reticulum . Together these data show that the endoplasmic reticulum is the exclusive site of lecithin formation in the castor bean endosperm and establish a central role for this cytoplasmic component in the biogenesis of cell membranes .
The ability of rice, wheat, and oat seedlngs to germinate and grow as the 02 concentration was lowered to zero was compared. The germination of rice was completely unaffected by 02 supply, whereas that of oats and wheat was strongly retarded at levels below 5% 02. In contrast to the coleopties of oats and wheat and to roots of all three species where growth was progressively diminished as the 02 concentration was lowered, that of the rice coleoptile was progressively increased. However, the dry weight and content of protein, sugars, and cellulose were all depressed In the rice coleoptile in anoxia, and the levels of several respiratory enzymes, particularly those of mitochondria, were also much lower than those of the coleoptiles grown in air. In 1% 02, the growth of the rice coleoptile was similar to that in air. The effect of ethanol concentration on germination and growth of rice was measured. Coleoptile growth was reduced when the ethanol concentration exceeded 40 mfllimolarity, and root growth was somewhat more sensitive. Coleoptiles of all three species grown in air were transferred to N2, and ethanol accumulation was measured over 24 hours. The rate of ethanol accumulation in oats was close to that in rice, and in all three species the amounts of ethanol lost to the surrounding medium were those expected from simple diffusion from the tissue. The ability of the rice coleoptile to grow in anoxia is apparently not due to a particularly low rate of ethanol formation or to unusual ethanol tolerance. Any explanation of the success of rice in anoxia must encompass the much lower rate of ATP synthesis than that in air and account for the biochemical deficiencies of the coleoptile.When deprived of 02, the seeds of most higher plant species fail to germinate, and for plants growing in air the imposition of anaerobic conditions prevents their further growth. Rice is one of the few species whose seeds can germinate in anaerobic conditions. Although there is now an extensive literature on some aspects of growth and anaerobic metabolism in rice, the biochemical basis for its unusual adaptation to anoxia is not known (8). It is frequently inferred that rice germinates better in the absence of 02, although several authors have emphasized that only the coleoptile elongates and no roots are produced under these conditions (4,11,17,22,31,33,34). In this paper, we present data on the growth and some biochemical features of rice seeds exposed to different levels of 02 Seeds sterilized in 1% NaOCl were germinated in darkness in 250-ml flasks containing five Whatman No. 1 filter papers and 7 ml H20. A moistened gas stream of air, 5% 02, 1% 02 or N2 (99.998% purity) was passed through the flasks at 3 ml/min throughout the experiment period.Growth Measurements. Increases in fresh and dry weights of roots, coleoptiles, and primary leaves were recorded over a 7-d period.Effect of Ethanol on Germination. Seeds were germinated in air in darkness in the presence of a graded series of ethanol concentrations from 0.08% (14 mM...
Ripe seeds of the castor bean (Ricinus communis) are very rich in fat ( > 60% of the dry weight) that is contained in endosperm tissue surrounding the embryo. During germination there is a precipitous fall in the total fat content, and sugars, principally sucrose, are formed. These sugars are absorbed and utilized by the growing embryo, but during the fifth day of germination, at 30' C, when fat breakdown is at its height, a gram of sugar accumulates for each gram of fat consumed.l The Pathway of Gluconeogenesis from Fat: The role o f the glyoxylate CycleThe elucidation of the pathway of this remarkably efficient conversion had to await the discovery of the glyoxylate cycle by Kornberg and Krebs2 in 1957. It was already known at that time that acetate-14C was converted to sucrose by endosperm slices3 and the process of P-oxidation in plants had been elucidated by Stumpf and his colleagues.4 The fact that glycolytic enzymes, phosphoenolpyruvate carboxykinase and sucrose synthetase5.6 were also present in the tissue allowed the formulation of a pathway in which oxalacetate could be converted to sucrose using ATP and NADH generated during the conversion of fatty acids to acetate. The glyoxylate cycle offered a possible solution to the problem of the net conversion of acetate to oxalacetate, and in 1957 Kornberg and I showed that malate synthetase and isocitrate lyase, the key enzymes of the cycle, were indeed present in castor bean endosperm and the following pathway was proposed:Glyoxylate cycleFat -+ Fatty acid + Acetyl CoA -+ + + + Oxalacetate + PEP + Hexose Studies on the distribution of malate synthetase and isocitrate lyase and on the changes in their activity during germination829 reinforced the proposal that the glyoxylate cycle was an important stage in gluconeogenesis from fat. In surveys of some 20 plant tissues, the two enzymes were found together only in those converting fat to sugar, and the activities rose from virtually zero in the ungerminated seed to a peak at the time of most rapid fat breakdown, and disappeared when this process was complete (FIGURE 7).Subsequently, experiments in which specifically labeled acetates and other intermediates were provided to endosperm slices, gave definitive evidence that established that the pathway of conversion was the one proposed.1° As predicted, C-1 of acetate appeared equally in COz and sucrose and C-2 contributed only to sucrose. The kinetics of labeling of intermediates and the labeling patterns in malate and hexose showed that acetate was metabolized almost exclusively by this sequence, which predicts a maximum conversion of 75% of fatty-acid carbon to sucrose (FIGURE 1 ) .
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