Protein and Ribonucleic Acid Synthesis by Plant Mitochondrial Systems

The incorporation of ¹⁴C-labelled phenylalanine into proteins of the mitochondrial systems obtained from 48-h germinating seeds of Vigna sinensis (L.) Savi can be stimulated by polyuridylic acid [poly (U)] and depressed by rifampicin, which is, however, ineffective if poly (U) is allowed to interact with the incorporating system before the antibiotic has access to it. A system consisting of a mitochondrial S-100 fraction and ribosomes from the same source with other cofactors can bring about polymerization of phenylalanine. The incorporation of ¹⁴C-labelled uracil into RNA by the plant mitochondria is greatly dependent on the exogenous addition of adenine, guanine, cytosine and also on 5-phosphoribosyl-l-pyrophosphate (5-PRPP). It is greatly suppressed by rifampicin and ethidium bromide.     

Interrelationship between ethylene and growth regulators in the senescence of lettuce leaf discs

The interrelationship between ethylene and growth regulators in the senescence of romaine lettuce (Lactuca sativa L.) leaves was studied. Gibberellic acid (GA₃), kinetin, and 3-indoleacetic acid (IAA) retarded chlorophyll loss from leaf discs which were floated on hormone solutions. Abscisic acid (ABA) and ethephon enhanced chlorophyll loss and antagonized the senescence-retarding effect of GA₃ and kinetin. A high concentration of IAA (10^–4 M) caused accelerated chlorophyll loss, whereas a similar concentration of kinetin neither retarded nor promoted chlorophyll loss. The ineffectiveness of IAA and kinetin at their supraoptimal concentrations in retarding leaf senescence was related to increased production of ethylene induced in the treated leaf discs. GA₃ was the most effective in retarding chlorophyll loss and did not stimulate ethylene production at all. The senescence-enhancing effect of ABA was not mediated by ethylene. However, the moderately increased production of ethylene, induced by relatively high concentrations of ABA, could act synergistically with the latter to accelerate chlorophyll loss. It is proposed that the effectiveness of exogenously applied hormones, both in enhancing and retarding senescence, is greatly affected by the endogenous ethylene concentration of the treated plant tissue.Contribution from the Agricultural Research Organization, The Volcani Center, Bet Dagan, Israel, No. 2571-E, 1988 series.     

Mode of action of antitumour antibiotics-III: Modulation of permeability of nuclear membrane in the presence of the antibiotics

The binding characteristics of the antibiotics to nuclei and their effect on the permeability of nuclear membrane with respect to histones and ribonucleic acids have been investigated. The binding constant for chromomycin A₃ was found to be 1.4 × 10⁴M^?1 and number of binding sites was equal to 3.48 ± 1.08 × 10¹² molecules/nuclei. The antibiotic chromomycin A₃ enhanced the uptake of lysine-rich histone, actinomycin D decreased the uptake and ethidium bromide had no effect. Chromomycin A₃ also enhanced the release of acid insoluble fraction containing RNA from the nuclei, actinomycin D and ethidium bromide inhibited the release of acid insoluble fraction containing RNA. The relevance of this finding to the role of nuclear envelope in understanding the mechanism of action of the antibiotic has been discussed.     

Plant enzyme synthesis: Hormonal regulation of invertase and peroxidase synthesis in sugar cane

Summary Using sugar-cane internodal tissue in which RNA synthesis was ratelimiting for invertase of peroxidase synthesis, measurements were made of enzymeforming-capacity after blocking further RNA synthesis with actinomycin D or 6-methylpurine. In this way it was possible to determine whether added auxin (naphthaleneacetic acid) or gibberellic acid (GA₃) affected steps prior or subsequent to synthesis of the RNA fractions specifically required for synthesis of either enzyme. Both auxin and GA₃ increased the enzyme-forming-capacity for invertase but not for peroxidase. The effects of the two hormones are interpreted as causing stabilization of mRNA for invertase.Abscisic acid (ABA) increased the rate of synthesis of invertase but not peroxidase. ABA did not change the rate of loss of invertase when peptide-bond formation was blocked with cycloheximide, but stimulated its synthesis when RNA synthesis was blocked with 6-methyl purine. Hence, the site of action of ABA is subsequent to invertase-mRNA formation and prior to invertase destruction.Kinetin had no short-term effects when RNA synthesis was limiting for invertase production, and does not appear to directly modulate mRNA synthesis or stabilization, or amino-acid-polymerization steps. In treatments longer than 5 hours, kinetin inhibited synthesis of all three enzymes studied, so that its effect on enzyme synthesis in this tissue appears to be unspecific.Abbreviations used throughout text ABA (±)-abscisic acid (abscisin II, dormin) - GA₃ gibberellic acid - NAA -naphthaleneacetic acid     

Effets du GA3 et de la BAP sur la respiration,l’absorption et le métabolisme du phosphate chez des disques foliaires dePelargonium zonale après survie

Respiration rate of foliarPelargonium discs was insensitive to ageing. The addition of BAP or GA₃ to the ageing medium did not produce any effect. The presence of GA₃ or BAP in the ageing medium induced an increase (27 %) or a decrease (45 %) of the phosphate uptake. The analysis of phosphorylated compound labelling showed that these two hormones decreased³²P incorporation in the non-acid soluble fraction and increased³²P incorporation in the acid soluble organic fraction. GA₃ and BAP had little effect on the distribution of radioactivity between the different acid soluble compounds, but they increased the ATP level. These results suggest that both GA₃ and BAP increase the basal metabolism, but they seem to act differently on the development of the uptake mechanism during ageing.     

Phytohormone effects on hydrolytic activity of phosphohydrolases in red beet (Beta vulgaris L.) tonoplasts

The effects of indole-3-acetic acid (IAA), abscisic acid (ABA), gibberellic acid (GA₃) and kinetin on the hydrolytic activity of proton pumps (adenosine triphosphatase, H⁺-ATPase, pyrophosphatase, H⁺-PPase) of tonoplasts isolated from stored red beet (Beta vulgaris L. cv. Bordo) roots were studied. Results suggest that the phytohormones can regulate the hydrolytic activities of H⁺-ATPase and H⁺-PPase of the vacuolar membrane. Each of the proton pumps of the tonoplast has its own regulators in spite of similar localization and functions. IAA and kinetin seem to be regulators of the hydrolytic activity for H⁺-PPase whereas for H⁺-ATPase it may be GA₃. Stimulation of enzyme activity by all hormones occurred at concentrations of 10^–6 to 10^–7 M.Abbreviations IAA indole-3-acetic acid - ABA abscisic acid - GA₃ gibberellic acid - H⁺-ATPase adenosine triphosphatase - H⁺-PPase pyrophosphatase - ATP adenosine triphosphate - Tris Tris (hydroxymethyl)-aminomethane - MES (2[N-Morpholino]) ethane sulfonic acid - EDTA ethylene diamine tetraacetic acid - P_i inorganic phosphate     

Ribonucleic Acid Synthesis in Plant Mitochondria: Effects of Some Inhibitors and Cyclic Nucleotides

Mitochondria isolated from 48-h germinating Vigna sinensis (L.) Savi can incorporate [³H]uridine into acid-insoluble material. The incorporation is highly sensitive to rifampicin and partially so to ethidium bromide, two specific inhibitors of template function. The inhibitory effect of rifampicin can be partly counteracted by cyclic 3′:5′-AMP but not by cyclic 3′:5′-GMP, if they are allowed to interact with the synthetic system before the treatment with rifampicin. This indicates that cyclic AMP and rifampicin compete for a common site on the RNA polymerase responsible for DNA-dependent RNA synthesis. Inhibition by ethidium bromide is unaffected by prior nucleotide interaction with the system.     

In Vitro DNA Synthesis by Chloroplasts Isolated from Marchantia polymorpha L. Cell Suspension Cultures

Lysate of chloroplasts prepared from liverwort Marchantia polymorpha L. cell suspension cultures incorporated [³H]-dTTP into acid insoluble materials when DNA was added exogenously as a template. The incorporation was highly dependent on the addition of template DNA, four deoxynucleoside triphosphates and magnesium ions (maximum incorporation at 5mM). Magnesium ions could be replaced by manganese ions. DNA synthesis inhibitors, N-ethylmaleimide (NEM) and ethidium bromide (EtBr), strongly inhibited the incorporation. Dideoxythymidine triphosphate (ddTTP), an inhibitor of DNA polymerases β and γ, inhibited the incorporation at the concentration of 50 μM (molar ratio of ddTTP/dTTP = 17). On the other hand, the incorporation by the chloroplast lysate was resistant to arabinofuranosyl cytosine triphosphate (araCTP) and aphidicolin as well as the RNA polymerase inhibitors, rifampicin and α-amanitin. The chloroplast lysate highly utilized denatured calf thymus DNA and bacteriophage ?X174 single-stranded DNA as templates when added exogenously, while a synthetic homopolymer, poly(rA)-oligo(dT)₁₂ ~ ₁₈, did not stimulate the incorporation at all. Autoradiographic analysis of DNA synthesized in isolated chloroplasts showed that the chloroplast DNA synthesis took place at several specific sites on the chloroplast DNA from cells of the liverwort, Marchantia polymorpha.     

Effects of kinetin and gibberellin a3 on callus growth and organ formation inLimnophila chinensis tissue culture

The action of exogenously applied hormones in the induction of morphogenesis inLimnophila chinensis (Osb.) Merr. tissue culture has been demonstrated. Stem expiants were grown on Murashige and Skoog’s medium containing various levels of kinetin, gibberellic acid and indole-3-acetic acid. Formation of roots, shoots (normal or abnormal), plantlets and friable, hard or nodulated calluses depended largely on the hormone levels used. The formation of normal shoots and roots were stimulated by treatment with kinetin. GA₃ treatment stimulated the bud differentiation but inhibited the root initiation. A combination of kinetin and GA₃ gave variable results.     

Effect of growth regulators onVicia faba plants irrigated by sea water Leaf area,pigment content and photosynthetic activity

The antagonistic effects of some growth regulators [i.e. indol-3-yl-acetic acid (IAA), gibberellic acid (GA₃) or kinetin] on stress imposed by sea water on leaf area, pigment and photosynthetic activity in leaves of broad bean plants at different stages of development were investigated. Seed priming with GA₃ alleviated either partially or completely the effects induced by the two levels of sea water (10 and 25 %) used on leaf area at all experimental stages. However, IAA, GA₃ and kinetin inhibited leaf growth by themselves in almost all measurements. Seed pretreatment with kinetin alleviated the inhibition of pigment production in sea water-irrigated plants. Furthermore, GA₃ or kinetin nullified the deleterious effects imposed by irrigation with sea water particularly the high level (25 %) on photosynthetic¹⁴CO₂ fixation.     

THE EFFECTS OF HORMONES UPON THE DEVELOPMENT OF EXCISED FLORAL BUDS OF AQUILEGIA

Young excised floral buds of Aquilegia were grown on defined medium containing kinetin, indoleacetic acid (IAA), or gibberellic acid (GA₃). Only when 10⁻⁶ or 10⁻⁷ m kinetin was added to the basal medium was there a significant increase in the number of initiated whorls of primordia. Buds on the basal medium or on medium with IAA or GA₃ failed to initiate carpels. On medium with 10⁻⁶ or 10⁻⁷ m kinetin, buds successfully initiated a normal whorl of five carpels. A high level of inorganic nitrogen was also required for the initiation of carpels. With 10⁻⁵ m kinetin, individual buds initiated from 6–18 carpels. Staminodial primordia of these buds were replaced with carpels, or the floral apex enlarged to accommodate a single whorl of many carpels. Kinetin did not support the further differentiation of the floral organs. Sepals, petals, and carpels did differentiate on medium with GA₃, but stamens aborted. However, on medium with GA₃ and kinetin, stamen primordia differentiated into short filaments and anthers. Further unknown growth factors appear to be required for the complete differentiation of floral primordia into mature organs.     

Effects of phytohormones on some biochemical parameters during dark induced leaf senescence ofSechium edule on Darjeeling Hill of the Eastern Himalayas

All the concentrations (25-150 mgl-¹) of the phytohormones kinetin, IAA (indol-3-ylacetic acid) and GA₃ (gibberellic acid) increased the activity of DCPIP (2,6 dichlorophenolindophenol)-Hill reaction, chlorophyll and protein contents over the control data in leaves ofSechium edule Sw. on Darjeeling hill of the Eastern Himalayas; while ethrel (2-chloroethylphosphonic acid) treatments decreased these parameters in the hilly species. The most effective concentrations in increasing these parameters were 50 mg 1-¹ of kinetin, 50 mg 1-¹ of IAA and 100 mg 1-¹ of GA₃; whereas 50 mg 1-¹ of ethrel was most effective in decreasing these parameters during the induction of senescence in the hilly vegetable crop. The increase in these parameters was greatest with kinetin, followed by IAA and least with GA₃ in the hilly plant species studied.     

Responses of <Emphasis Type="Italic">Nigella sativa</Emphasis> to foliar application of gibberellic acid and kinetin

Foliar sprays of water or 1, 10 and 100 μM aqueous solutions of gibberellic acid (GA₃) or kinetin (KIN) were applied to 40-d-old plants of Nigella sativa (L.) to study their effects on net photosynthetic rate, nitrogen metabolism, and the seed yield. 10 μM solutions of both the hormones, especially GA₃, appreciably increased the activities of nitrate reductase and carbonic anhydrase, chlorophyll and total protein contents and net photosynthetic rate in the leaves, along with capsule number and seed yield plant⁻¹, at harvest.     

Effect of gibberellin and kinetin on the regeneration ability ofFucus vesiculosus L.

The influence of gibberellic acid (GA³) and kinetin (6-furfurylaminopurine) on the regeneration ability of the basal and apical thallus fragments ofFucus vesiculosus L. was examined. The naturally occurring gibberellin and kinetin-like substances in these thallus fragments were also studied. It was found that exogenously applied GA³ markedly increased the number of adventitious branches formed on the cut surface of the thallus fragments taken from the apical parts of plants. The concentration of 0.001 mg GA³ I-1 proved to be the most effective. The growth promoting effect of GA³ was increased by simultaneous action with kinetin. In experiments in which the fragments of the basal parts of the thallus were treated with GA³, as a rule a slight growth inhibition was observed. The growth responses of the investigated plant tissues to gibberellin and kinetin varied according to season. Usually their susceptibility to the applied plant hormones was greater in spring than is summer. The shifts in growth reaction were related to the seasonal changes in the content of endogenous gibberellin and kinetin-like substances in the investigated parts of the thallus. It is suggested that growth regulators of the gibberellin and cytokinin type are involved in the regeneration processes inFucus.     

The Uptake of Cytokinins by Protoplasts and Root Sections of Brassica campestris

The effect of cytokinins was studied on the incorporation of ¹⁴C-labelled precursors into the nucleic acid fraction of protoplasts isolated from callus or roots of Brassica campestris. Protoplasts from callus and roots took up ¹⁴C-uridine from the incubation medium and incorporated this precursor into the ribonucleic acid fraction during the experimental period of 16 h. Low concentrations of kinetin (10^?8-5 × 10^?6M) did not stimulate the incorporation, and kinetin inhibited this process at higher concentrations (5 × 10^?5M). This result led to an investigation on the uptake of cytokinins by protoplasts of roots. In contrast to a rapid uptake of radio-actively labelled adenine and uridine. protoplasts from roots took up only small amounts of labelled kinetin. zeatin, zeatin riboside and zeatin nucleotides from the incubation medium. Root sections took up far more adenine and kinetin than protoplasts from roots. The ratio between the amount of kinetin taken up and applied was much higher for the sections than for protoplasts, indicating that intact root cells took up kinetin far more rapidly than protoplasts. It is suggested that the plasmalemma and cell wall play an essential role in the uptake of cytokinins or that the differences in the uptake rates are related to differences between the rates of metabolism of cytokinins in root sections and in protoplasts.     

Phase Change in Hedera helix: Induction of the Mature to Juvenile Phase Change by Gibberellin A3

A sensitive and reproducible method to obtain GA₃ induced morphological reversion of mature Hedera helix to the juvenile form has been developed. Dose response experiments indicate that GA₃ stimulates reversion over a 50–100 fold range with a half maximal response at approximately 0.5 μg GA₃ per plant. The individual characteristics involved in phase change revert to the juvenile form in a sequential manner as GA₃ dose is increased. Variations in light intensity from 1.2–3.6 × 10⁴ lux and temperature from 15 to 26°C do not affect this hormonal response. Other growth regulators including indoleacetic acid, kinetin, abscisic acid and (2-chloroethyl)phosphonic acid (Ethephon) are inactive but other gibberellins (GA₁ and a mixture of A₄–A₇) are active in stimulating reversion. Therefore, the response is specific for gibberellins as a class of hormones but non-specific for a particular form of gibberellin. The significance of this response in relation to juvenility in woody plants is discussed.     

INFLUENCE OF HORMONAL TREATMENTS ON THE GROWTH OF TWO HALOPHYTIC SPECIES OF SUAEDA

Experiments were made to determine the interaction between the growth regulating substances, gibberellic acid and kinetin, and salinity on the growth and development of two species of Suaeda. We found that the growth of Suaeda maritima var. macrocarpa, an obligate halophyte, was greatly stimulated with treatments by both of these hormones in controls not treated with sodium chloride. These results suggest that halophytes grow poorly under nonsaline conditions because of some type of hormonal imbalance. GA₃ was found to stimulate growth at all salinities for both S. maritima var. macrocarpa and S. depressa, while kinetin proved to be inhibitory to growth and elongation of plants at higher salinities. Chlorophyll content decreased with salt treatments and was not significantly influenced by hormonal treatments. GA₃ had little influence on water content in roots and shoots, whereas plants treated with kinetin generally had reduced water contents.     

Kinetin-promoted transport of assimilates in stems of Phaseolus vulgaris L.

Kinetin, applied as a dispersion in aqueous lanolin to the stumps of decapitated stems of P. vulgaris plants with their roots removed, was found to promote the transport of ¹⁴C- and ³²P-labelled assimilates to the site of hormone application. Measurement of photosynthetic rate of, and assimilate export rate from the source leaves, indicated that kinetin was not acting to promote assimilate transport by stimulating these processes. Moreover, it was found that the time between kinetin application and detection of an enhanced transport flux was independent of the distance over which kinetin would need to move to be present throughout the length of the transport pathway. These observations, together with the finding that lateral applications of kinetin to the stems resulted in an enhanced localized accumulation of assimilates, provided evidence that kinetin acted locally at its point of application to stimulate assimilate transfer.Abbreviations GA₃ gibberellic acid - IAA indol-3yl-acetic acid     

Glyceride metabolism and gluconeogenesis in barley endosperm

Endosperms of quiescent barley grains contained, on average, 54.5 μg of neutral glyceride-glycerol, equivalent to ca 480 μg glyceride. Of this probably 90% was located in the aleurone layer. During germination the level of glyceride-glycerol declined. It also declined in degermed grains and aleurone layers incubated in vitro. The fall was accelerated by GA₃, but indoleacetic acid, kinetin and glutamine were without effect. Increases in the levels of malate synthase and isocitrate lyase from very low initial values, and the results of incorporation studies with [¹⁴C]-labelled substrates, indicate that the glyoxylate cycle functions to convert glycerides to sucrose in germinating grains and degermed grains incubated with GA₃, but not in degermed grains without the hormone. In the absence of GA₃ the glyceride could be a respiratory substrate in degermed grains. The aleurone layers converted exogenous glucose to sucrose. Little label from [¹⁴C]-amino acids appeared in sucrose but in some cases considerable incorporation occurred into glutamine.