Biotransformation of digitoxigenin by ginseng hairy root cultures.

Five new compounds (three esters and two glycosides) and seven previously reported compounds were isolated as biotransformation products of digitoxigenin by ginseng hairy root cultures. The new esters and glycosides were elucidated as digitoxigenin stearate, digitoxigenin palmitate, digitoxigenin myristate, 3-epidigitoxigenin beta-D-gentiobioside and digitoxigenin beta-D-sophoroside using 1H and 13CNMR and FAB mass spectral data. Biotransformations involving esterification (of stearic acid, palmitic acid, myristic acid and lauric acid) and glycosylation (of gentiobiose and sophorose) of digitoxigenin have been demonstrated for the first time in the plant cell and tissue cultures. The hairy roots showed high glycosylation ability to the digitoxigenin molecule.     

Cooperative regulation of cytoplasmic streaming and ca fluxes by pfr and photosynthesis in vallisneria mesophyll cells

In mesophyll cells of Vallisneria gigantea Graebner, Ca²⁺ regulates the induction and cessation of cytoplasmic streaming. Streaming is induced when the level of calcium in the cytoplasm is lowered through light-accelerated release of Ca²⁺ from the cells (S Takagi, R Nagai [1988] Plant Physiol 88: 228-232). We have now initiated an investigation on the nature of the photoreceptor(s) that are involved in the regulation of Ca²⁺ movements across the cell membrane and of streaming. Streaming is induced only when phytochrome exists in the phytochrome—far redabsorbing form (Pfr)—and photosynthesis is allowed to take place for at least 4 minutes. The former effect is typically photoreversible by red and far-red light, and phytochrome is spectro-photometrically detectable in the crude extract from the leaves. The latter effect is assessed in terms of the wavelength dependency and the effects of diuron and atrazine, two inhibitors of photosynthesis. A similar requirement for Pfr and photosynthesis is found to be associated with the acceleration of Ca²⁺ efflux in the protoplasts. The results suggest that phytochrome and photosynthetic pigment(s) cooperatively regulate cytoplasmic streaming via modulation of the Ca²⁺ transport in the cell membrane.     

Phytochrome-induced intercellular signalling activates cab::luciferase gene expression

The phytochrome-induced expression pattern of the chlorophyll a/b binding protein (cab) gene was studied using a cab2::luciferase reporter transgene in the tobacco cotyledon. The role of developmentally regulated competence, cooperativity among cells and signal propagation was investigated by red-light microbeam irradiation of distinct areas of the cotyledon. Even with a minimal fluence, the response was not restricted to the irradiated cells. Following irradiation of the cotyledon base, the luciferase activity revealed a robust propagation of activating signals to areas that had not received a light stimulus. The acropetal outspread formed distinct expression patterns depending on the site of the irradiation. The combination of imaging luciferase activity in living seedlings and microbeam microscopy provides significant experimental evidence of how cellular light perception and intercellular signalling contribute to the cab gene expression pattern.     

Effects of the Timing of Flashes of Light during the Course of Cellular Rotation on Phototactic Orientation of Individual Cells of Cryptomonas

The swimming movement of Cryptomonas sp. cells generates a helical path, as a result of rotations with an average period of 500 milliseconds. When a flash of light at 570 nanometers for 20 microseconds was applied unidirectionally at intervals of 500 milliseconds, only a fixed side of each rotating cell was repeatedly exposed to the flashes of light. The relationship between the irradiated side of a cell and the phototactic orientation of the cell, rotating with a period of 475 to 525 milliseconds, was determined by infrared videomicrography. Only when the ventral sides of the cells were exposed to the flashes of light did their courses shift predominantly toward the light source. This result suggests that light is efficiently detected by the ventral side of these organisms.     

Synthetic partial extension peptides of P-450(SCC) and adrenodoxin precursors: effects on the import of mitochondrial enzyme precursors

Various portions of the extension peptides of P-450(SCC) precursor were chemically synthesized. The effects of these peptides on the import of enzyme precursors into mitochondria were examined. Peptides SEP1-15 and SEP1-20, corresponding to the amino terminal portion of the extension peptides, strongly inhibited the import of P-450(SCC) precursor into mitochondria. These peptides were effective at concentrations above 30 microM, and complete inhibition was observed at 100 microM. SEP1-11, which is shorter than SEP1-15 and SEP1-20, showed very weak inhibition. SEP25-39, which corresponds to the carboxy terminal portion of the extension peptide, did not affect the import of the precursor. The import of P-450(11 beta) and adrenodoxin precursors were also inhibited by SEP1-15. Another peptide, AEP1-14, which corresponds to the amino terminal portion of the extension peptide of adrenodoxin precursor, was also synthesized. The peptide inhibited the import of both adrenodoxin and P-450(SCC) precursors into mitochondria. The import of malate dehydrogenase was also inhibited by SEP1-15 and AEP1-14. The rate of the internalization of the precursor into mitochondria was decreased by the peptides. The amount of the precursor bound to the surface of mitochondria and the processing of adrenodoxin precursor were not affected. The respiratory activities of isolated mitochondria were not influenced by SEP1-15 even at 100 microM. We conclude that the inhibitory activities of the synthetic partial extension peptides on the import of enzyme precursors into mitochondria require the presence of about fifteen amino acid residues in the amino terminal portion of the extension peptides, and the inhibition of the import by the peptides was dependent on the blockage of the internalization of the precursors into mitochondria.     

Volume changes in activated human neutrophils: The role of Na+/H+ exchange

The apparent volume of neutrophils, as measured electronically with the Coulter counter, has been reported to increase upon treatment with chemotactic factors. The occurrence of a volume change was confirmed by forward angle light scattering and by isotopic measurements of intracellular water space in cells treated with 12-O-tetradecanoylphorbol 13, acetate (TPA) or formyl-methionyl-leucyl-phenylalanine (FMLP). Cell swelling was associated with an increase in the osmotic content of the cells, determined from Boyle-van't Hoff plots, and with an increase in Na⁺ content, measured by flame photometry. The volume change was inhibited by replacement of extracellular Na⁺ with K⁺ or N-methyl-D-glucamine⁺, or by addition of amiloride. Swelling was also inhibited by the 5-N-substituted analogs of amiloride, which are potent specific inhibitors of the Na⁺/H⁺ antiport. This pathway is activated in neutrophils by both TPA and FMLP. Activation of Na⁺/H⁺ exchange, determined as a Na⁺-dependent and amiloride-sensitive cytoplasmic alkalinization, was also found when neutrophils were treated with hypertonic solutions. The hypertonic activation of the antiport was similarly followed by cell swelling, detectable by electronic sizing. The results indicate that activation of Na⁺/H⁺ exchange can lead to significant cell swelling in neutrophils.