Isolation, partial chemical and biological characterization of thymone B

A new approach to the study of the molecular arrangements of proteins in membranes is described. Irradiation with visible light of native erythrocytes or washed erythrocyte membranes suspended in buffers containing a) riboflavin, fluorescein or fluorescein coupled to dextran and b) ³H-labelled tryptophan resulted in incorporation of radioactivity into the membrane proteins. Polyacrylamide gel electrophoresis of solubilized membranes followed by radioactivity measurements of the separated membrane proteins revealed that in native erythrocytes the protein components known to be located at the exterior cell surface, Band 3 and the major sialoglycoproteins became specifically labelled, whereas in washed lysed cells all of the major membrane proteins were labelled.     

Activation of fatty acids by non-glyoxysomal peroxisomes

Ethylene treatment (approx. 20 l ·1^-1 in air for 2 d) of tobacco (Nicotiana tabacum L. cv. Havana 425) plants markedly increases the endo--1,3-glucanase (EC 3.2.1.39) content of leaves. The antigenic form of the enzyme induced is the same one whose production is blocked by treating cultured cells with combinations of auxin (1.1 · 10^-5 M -naphthaleneacetic acid) and cytokinin (1.4 · 10^-6 M kinetin). Evidence is presented that cultured tobacco cells require ethylene for -1,3-glucanase accumulation: i) ethylene treatment increased the accumulation of \-1,3-glucanase in callus tissues >10 d after subculturing and in cell-suspension cultures; ii) callus tissues can produce ethylene; iii) conditions known to inhibit ethylene production (1 mM CoCl₂; 33° C treatment) or ethylene action (approx. 1.6 mmol · 1^-1 norbornadiene in air) inhibited -1,3-glucanase accumulation by callus tissues treated for 4 d following subculturing; and, these inhibitory effects were prevented by exogenous ethylene. Combinations of auxin and cytokinin blocked ethylene-induced accumulation of -1,3-glucanase by cell-suspension cultures. The results favor a model in which ethylene induces results favor a model in which ethylene induces 1,3-glucanase accumulation, and auxin and cytokinin inhibit this induction process.Abbreviations NAA -naphthaleneacetic acid - NDE norbornadiene     

Effects of growth regulators on light-dependent anthocyanin production in Zea mays seedlings

Rengel, Z. and Kordan, H. A. 1987. Effects of growth regulators on light-dependent anthocyanin production in Zea mays seedlings.
The effects of ethylene, indolyl- and naphthylacetic acids, zeatin, benzyladenine, gib-berellic acid and triiodobenzoic acid on anthocyanin production in seedlings of Zea mays L. cv. Golden Bantam were investigated. Endogenously produced and exogen-ously supplied ethylene, as well as the other growth regulators tested markedly suppressed anthocyanin formation. Except for triiodobenzoic acid, the other growth regulators stimulated ethylene production, the amounts produced in the light being larger than those in the dark. Absorption of ethylene by permanganate as well as inhibition of ethylene production or action by Co²⁺ or Ag⁺ increased anthocyanin formation in maize seedlings above the level found in the control plants. The inhibiting effect of auxins and cytokinins on anthocyanin production was reversed by Co²⁺ or Ag⁺. In contrast, decreased anthocyanin formation caused by gibberellic acid or triiodobenzoic acid seemed unrelated to ethylene and could not be alleviated by Co²⁺ or Ag+.     

Induction of secondary dormancy in sunflower seeds by high temperature. Possible involvement of ethylene biosynthesis

High temperature (45°C) inhibits seed germinition and seedling sunflower ( Helianthus annuus L. cv. Mirasol). Treatment of imbibed seeds at 45°C for more than 48 h induces a secondary dormancy, which is associated with progressive decrease of germination ability at optimal temperature (25°C) as well as with abnormal seedling growth. Ethylene (55μl l⁻¹) and 2-chloroethylphosphonic acid (ethephon) (2.5 m M ) improve germination of thermodormant seeds at 25°C. but the abnormal growth of the seedlings remains. O₂-enriched atmosphere and dry storage improve germination and normal seedling growth. The induction of thermodormancy in sunflower seeds seems associated with loss of their ability to convert 1-aminocyclopropane-1-carboxylic acid (ACC) to ethylene. Possible effects of high temperature on membranes and ethylene forming enzyme (EFE) are discussed.     

Ethephon and auxin induce mycorrhiza-like changes in the morphology of root organ cultures of Mugo pine

The effects of ethylene and auxin on the morphology and anatomy of root organ cultures of Pinus mugo Turra var. mugo were investigated to test the hypothesis that changes in root morphology associated with formation of ectomycorrhizae may be related to ethylene produced by ectomycorrhizal fungi or by host plant roots in response to fungus-produced auxin. Morphological changes characteristic of mycorrhizal infection include dichotomous branching of lateral roots, inhibition of root hair formation and enlargement of cortical cells. Lateral roots on non-mycorrhizal root organ cultures, grown in a defined medium, underwent dichtotomous branching while root hair formation was inhibited in response to the ethylene released by 50 and 100 μ M ethephon (2-chloroethylphosphonic acid), but no effect on cortical cell dimensions was observed. The auxin, naphthaleneacetic acid (1 and 10 μ M ) also stimulated dichotomous branching and inhibited root hair formation, but to a lesser extent and with a greater lag time than ethephon. Auxin-stimulated ethylene production by root organ cultures was demonstrated. This appeared to be responsible, at least in part, for the auxin-induced dichotomous branching since the ethylene action inhibitor, silver thiosulfate (0.1 m M ) inhibited the response to auxin by 35%.     

Vasoactive intestinal peptide effects on GH3 pituitary tumor cells: high affinity binding, affinity labeling, and adenylate cyclase stimulation. Comparison with peptide histidine isoleucine and growth hormone-releasing factor

The vasoactive intestinal polypeptide (VIP) receptor was characterized on the GH3 rat pituitary tumor cell line using competitive binding studies with peptides having sequence homology with VIP. Further studies investigated receptor coupling to the adenylate cyclase complex by measurement of cAMP levels. Finally, the molecular weight of the receptor was estimated by affinity labeling techniques. Studies using 125I-VIP and unlabeled competing peptides revealed a single class of high affinity binding sites with a dissociation constant (KD) of 17 +/- 2 nM (mean +/- S.E.M.) for VIP, 275 +/- 46 nM for peptide histidine isoleucine (PHI), and 1380 +/- 800 nM for human pancreatic growth hormone releasing factor (GHRF). VIP and PHI each stimulated intracellular cAMP accumulation in a dose-dependent manner; both peptides demonstrated synergism with forskolin. In contrast, GHRF neither stimulated accumulation of cAMP nor demonstrated synergism with forskolin. VIP plus PHI (1 microM each) caused no significant increase in cAMP over either VIP or PHI alone, implying that the two peptides act through the same receptor. Covalent crosslinking of 125I-VIP to its binding site using either disuccinimidyl suberate (DSS) or ethylene glycol bis(succinimidyl succinate) (EGS) was followed by SDS-PAGE and autoradiography. The result is consistent with an Mr 47 000 VIP-binding subunit comprising or being associated with the VIP receptor of GH3 pituitary tumor cells.     

Seed germination in Chenopodium album L.: further evidence for the dependence of the effects of growth regulators on nitrate availability

Abstract Chenopodium album L. plants, grown under controlled environmental conditions on different levels of soil nitrate, produced seeds with proportionately different NO^?₃ contents. Regardless of the endogenous NO^?₃ content, few seeds germinated in water or upon treatment with KNO₃. Ethylene promoted germination, and the extent of germination was positively correlated with the endogenous seed NO^?₃ content. Combined application of ethylene and KNO₃ in the dark had a synergistic effect on NO^?₃ -deficient seed. The synergism between ethylene and KNO₃ was attributable to the NO^?₃ moiety of the nitrate salt. Ethylene and light showed moderate synergism in seeds with low or high endogenous nitrate. Addition of nitrate, however, masked the interaction between ethylene and light. Gibberellic acid₄₊₇ (GA₄₊₇) or red light, each alone or combined with KNO₃, had little effect on germination. When applied together in the dark, ethylene and GA₄₊₇ synergistically enhanced the germination of NO^?₃-deficient seed. The combined effects of the two hormones on this seed were further enhanced by the addition of KNO₃. There was no synergism between ethylene and GA₄₊₇ in NO^?₃-rich seed. These interactions among GA₄₊₇, ethylene and KNO₃ were not affected by light. The results confirm and further elaborate our earlier finding that the sensitivity of C. album seeds to ethylene may depend on nitrate availability.     

Receptor-mediated regulation of calcium mobilization and cyclic GMP synthesis in neuroblastoma cells

In neuroblastoma N1E 115 cells, carbachol, histamine and PGE1 elevated cyclic GMP content and, induced the efflux of preloaded 45Ca2+, the release of membrane-bound Ca2+ measured by fluorescent CTC, and the increase in [Ca2+]i as measured by Quin 2 fluorescence. The time course of the responses, the absolute requirement of extracellular Ca2+, the inhibition by receptor blockers, and the concentration dependency on histamine were all similar between these responses. The observation indicates that the mobilization of Ca2+, especially the increase of [Ca2+]i, may be intimately linked to the synthesis of cyclic GMP in the cells.     

The effect of drought on levels of abscisic acid,cytokinins, gibberellins and ethylene in aeroponically-grown sunflower plants

Abscisic acid (ABA), cytokinins and gibberellin-like substances (GAs) were extracted from the roots and shoots of 17-day-old sunflower seedlings which had been droughted or were unstressed. Plants were grown in an aeroponic chamber which allowed for good control over degree of water stress and easy access to roots. Following methanolic extraction of lyophilized material, cytokinins were separated from the acidic growth-regulators on a cellulose PO₄ cationic exchange column. The cytokinins were analysed by paper chromatography and HPLC and the soybean hypocotyl section assay. Semipurified acidic regulators were chromatographed on SiO₂ columns and HPLC and aliquots assayed with the dwarf rice cv. Tan-ginbozu bioassay for GAs. Fractions known to contain ABA were purified by sequential reverse-phase HPLC of the acid and then of the methyl ester forms followed by quantitation as Me-ABA on GLC-EC. ABA losses were measured by using an internal standard [³H]-ABA). Ethylene production was also monitored in stressed and unstressed seedlings.The effect of drought on GAs and ethylene was minimal. The ABA levels were markedly higher in droughted plants. Stressed roots had 32 times more ABA than controls. The levels of cytokinins in the shoots of droughted plants were about half those in unstressed shoots, and qualitative differences occurred in the roots. Under stress a large peak of activity was present similar to zeatin glucoside which was not present in the unstressed condition. The results are discussed in relation to drought-effects on metabolism.     

Disassembly of microtubules by the action of calmodulin-dependent protein kinase (Kinase II) which occurs only in the brain tissues

Microtubules assembled by the incubation of GTP at 37 °C were disassembled by the action of calmodulin-dependent protein kinase (Kinase II) which occrs only in the brain tissues. This disassembly required the presence of ATP and physiological concentrations of Ca²⁺ and calmodulin.