The Effect of CCC and Amo-1618 on Growth, Catalase, Peroxidase and Indoleacetic Acid Oxidase Activity of Young Barley Seedlings

Barley seedlings were grown in darkness on filter paper saturated with phosphate buffer or CCC and Amo-1618 buffered solutions. The effects of CCC and Amo-1618 on growth, catalase, peroxidase and lAA-oxidase were studied on coleoptile and primary leaves. Both growth-retarding chemicals cause an inhibition of growth, Amo-1618 being more effective than CCC. They have no effect on catalase activity. Increasing concentrations of CCC and Amo-1618 progressively stimulate peroxidase and IAA-oxidase activities (Amo-1618 more than CCC). The enzymatic activity in short-treated plants is higher than in the corresponding control plants of the same height. It is proposed that CCC and Amo-1618 exert their effect on the growth of barley by acting on auxin catabolism.     

Kinetics of growth retardant and hormone interactions in affecting cucumber hypocotyl elongation

The capacities of indole-3-acetic acid (IAA) and gibberellin A₃ (GA₃) to counteract the inhibitory effects of (2-chloroethyl) trimethylammonium chloride (CCC), 2-isopropyl-4-dimethylamino-5-methylphenyl-1-piperidinecarboxylate methyl chloride (Amo-1618), and N,N-dimethylaminosuccinamic acid (B-995) on hypocotyl elongation in light-grown cucumber (Cucumis sativus L.) seedlings were investigated. One μg of GA₃ applied to the shoot tip was sufficient to completely nullify the effect of 10 μg of Amo-1618 or 25 μg of B-995 applied simultaneously to the shoot tip, and 10 μg of GA₃ completely counteracted the effect of 10⁻³ m CCC added to the root medium. One μg of IAA counteracted the effect of 10⁻³ m CCC in the root medium, but IAA did not nullify the action of either Amo-1618 or B-995. Experiments were conducted using 2 growth retardants simultaneously, which indicated that Amo-1618 and CCC inhibit a common process, namely GA biosynthesis, essential to hypocotyl elongation. However, since the effect of CCC was overcome by applications of both GA and IAA, growth retardation resulting from treatment with CCC apparently is not due solely to inhibition of GA biosynthesis. B-995 did not interact additively with either Amo-1618 or CCC, which suggests that B-995 affects a process different from those affected by the other 2 retardants. Thus, while inhibition evoked by B-995 is reversible by applied GA, the action of B-995 does not appear to be inhibition of GA biosynthesis.     

Some Effects of Amo-1618 on Growth, Peroxidase and α-Amylase Which Cannot Be Easily Explained by Inhibition of Gibberellin Biosynthesis

Growth and peroxidase activity of roots and stems of lentil seedlings were compared after treatment with Amo-1618, alone or in combination with gibberellic acid (GA) at varying concentrations. The peroxidase enhancement in Amo-1618 treated stems could not be attributed to a decrease in the gibberellin content since GA alone had no effect on this enzyme. In other experiments, AMO, at low concentrations, was able to induce α-amylase production in barley aleurone layers; the lag period needed for this induction, was longer than for GA. These facts seem to indicate that some growth retardants might act at least in some cases by mechanisms other than inhibition of gibberellin biosynthesis and reversal of GA action.     

Effects of Growth Retardants on Norway Spruce (Picea abies)

Young seedlings of Picea abies Karst, grown in nutrient solution were treated with the growth retardants Amo-1618, B-995, and CCC. These were added to the nutrient medium. B-995 and CCC retarded root and shoot growth in the concentrations 100, 10, and 1 mg/l. Growth was almost entirely inhibited by 300 mg/l, obviously due to toxicity. The effects of Amo-1618 were similar but more varying. GA counteracted the effects of all the retardants on shoot growth, but not on root growth.     

The effect of cotyledons on hypocotyl elongation in light-grown bean seedlings: Comparison between dwarf and tall varieties

Hypocotyl sections with and without the cotyledons were cutfrom bean seedlings and incubated under white light of 6000lux. The cotyledons had an inhibitory effect as well as a promotiveeffect on hypocotyl growth. The former effect was more strikingin the dwarf variety, and the latter in the tall variety. Whenthe hypocotyl units were exposed to light for shorter times(6 hr or less) or incubated under weaker light (1600 and 50lux), the inhibitory effect of the cotyledons decreased greatly,and in the tall variety the presence of cotyledons producedno inhibition, but a promotion of hypocotyl growth. GA treatmentenhanced hypocotyl growth and counteracted the growth inhibitioncaused by the cotyledons. On the whole, the GA effect was moremarked in the tall variety than in the dwarf. The elongation of bean hypocotyls may be controlled by a balancebetween the inhibitory and promotive effects of cotyledons,and the predominance of the former over the latter may be oneof the causes for expressing dwarfing. (Received November 13, 1976; )     

Increase in indoleacetic Acid oxidase activity of winter wheat by cold treatment and gibberellic Acid

The activity of indoleacetic acid oxidase increased 10-fold during 40 days of cold treatment of winter wheat seedlings. Puromycin and 6-methyl purine inhibited indoleacetic acid oxidase development in the cold. Addition of gibberellic acid stimulated indoleacetic acid oxidase development during germination at room temperature and during cold treatment. Amo-1618 inhibited indoleacetic acid oxidase development before and during cold treatment. Indoleacetic acid treatment increased indoleacetic acid oxidase activity during germination at room temperature while no significant effect on activity was observed during cold treatment.     

胚轴对萌发豌豆子叶中淀粉酶活性表达的影响

萌发豌豆的上、下肢轴均能诱导子叶中淀粉酶活性,外源GA和6—BA具有类似胚轴的作用。离体子叶的淀粉酶凝胶电泳只有一条活性极低的酶带,连生子叶中有两条酶带,其中由胚轴诱导新出现了一条活性很高的同工酶带,它的活性受亚胺环己酮的强烈抑制,而受放线菌素D影响不大。推测豌豆子叶中存在淀粉酶的长寿命mRN—A,胚轴和外源激素的作用在于促进mRNA的翻译。     

cDNA cloning and differential gene expression of three catalases in pumpkin

Three cDNA clones (cat1, cat2, cat3) for catalase (EC 1.11.1.6) were isolated from a cDNA library of pumpkin (Cucurbita sp.) cotyledons. In northern blotting using the cDNA-specific probe, the cat1 mRNA levels were high in seeds and early seedlings of pumpkin. The expression pattern of cat1 was similar to that of malate synthase, a characteristic enzyme of glyoxysomes. These data suggest that cat1 might encode a catalase associated with glyoxysomal functions. Furthermore, immunocytochemical analysis using cat1-specific anti-peptide antibody directly showed that cat1 encoding catalase is located in glyoxysomes. The cat2 mRNA was present at high levels in green cotyledons, mature leaf, stem and green hypocotyl of light-grown pumpkin plant, and correlated with chlorophyll content in the tissues. The tissue-specific expression of cat2 had a strong resemblance to that of glycolate oxidase, a characteristic enzyme of leaf peroxisomes. During germination of pumpkin seeds, cat2 mRNA levels increased in response to light, although the increase in cat2 mRNA by light was less than that of glycolate oxidase. cat3 mRNA was abundant in green cotyledons, etiolated cotyledons, green hypocotyl and root, but not in young leaf. cat3 mRNA expression was not dependent on light, but was constitutive in mature tissues. Interestingly, cat1 mRNA levels increased during senescence of pumpkin cotyledons, whereas cat2 and cat3 mRNAs disappeared during senescence, suggesting that cat1 encoding catalase may be involved in the senescence process. Thus, in pumpkin, three catalase genes are differentially regulated and may exhibit different functions.     

Chromatin RNA polymerase activity from soybean hypocotyls treated with gibberellic acid and AMO-1618

Chromatin isolated from control, AMO-1618 [2′-isopropyl-4′-(trimethylammonium chloride)-5′-methylphenyl piperidine carboxylate] and gibberellic acid (GA) treated soybean hypocotyl tissue incorporates labeled nucleoside triphosphates into acid-insoluble RNA. Gibberellic acid, sprayed on intact soybean hypocotyls, is shown to have enhanced the level of chromatin RNA polymerase activity while chromatin isolated from hypocotyls pretreated with AMO-1618 exhibits a lower polymerase activity relative to the control. Chromatin extracted from the treated or untreated seedlings are all sensitive to the inhibition (in varying degrees) by the presence of actinomycin D, pyrophosphate, or ribonuclease. Thus enhanced (or decreased) RNA-synthesizing capacity of chromatin in response to chemical treatments may be due to enhanced (or decreased) synthesis of RNA polymerase.     

Relationship of IAA-oxidase Activity to Gibberellinand IAA-induced Elongation of Light-grown Cucumber Seedlings

The growth and IAA-oxidase activity of light-grown cucumber seedlings (cv. Aonagajibae) were investigated in response to GA₃ and IAA. Both GA₃ and IAA induced significant elongation of the hypocotyl. The fresh and dry weights of the hypocotyl increased due to GA₃ or IAA treatment, whereas no significant change was observed in the cotyledons of GA₃-treated seedlings as compared with the controls. The fresh and dry weights of IAA-treated cotyledons were both lower than those of controls. Treatment with GA₃ or IAA resulted in retardation of IAA-oxidase activity in the hypocotyl and cotyledons. The degree of retardation was less in the cotyledons than in the hypocotyl. An inverse relationship was recognized between GA₃- or IAA-induced elongation and IAA-oxidase activity in the hypocotyl. The auxin-mediated mechanism for gibberellin action was discussed.     

EFFECTS OF GIBBERELLIN AND AMO-1618 ON GROWTH,DRY-MATTER ACCUMULATION,CHLOROPHYLL CONTENT AND PEROXIDASE ACTIVITY OF CITRUS SEEDLINGS

Monselise , S. P., and A. H. Halevy . (Hebrew U., Rehovot, Israel.) Effects of gibberellin and AMO–1618 on growth, dry-matter accumulation, chlorophyll content and peroxidase activity of citrus seedlings. Amer. Jour. Bot. 49(4): 405–412. Illus. 1962.—Sweet-lime seedlings, 6 months old, were sprayed with gibberellic acid (GA) and a growth retardant, AMO–1618, alone and in combination, at concentrations ranging between SO and 1600 ppm. Increasing concentrations of GA progressively increased shoot and internode length, did not influence number of leaves, and decreased leaf area. Dry weight of shoots was progressively increased up to 400 ppm, while dry weights of leaves and roots were decreased over all GA concentrations. Total dry weight of plants was increased by GA when related to leaf area or weight and to total chlorophyll content, which indicates a higher synthetic efficiency of leaves. This could not be detected by manometric determinations using leaf discs. It is suggested that a short determination period and/or work with detached leaves are responsible for failures to detect increased photosynthetic activity of GA-treated leaves. Chlorophyll content of leaves was decreased by increasing GA concentrations; it is shown that this is not due to “dilution” over a larger area of leaves. Peroxidase activity of leaves was only slightly reduced by GA, while it was increased by AMC–1618, acting as antagonist to GA. This is remarkable, since AMO–1618 did not clearly affect other procperties of citrus seedlings which are only slightly responsive to this chemical.     

Studies on energy status and mitochondrial respiration during growth and senescence of mung bean cotyledons

Several characteristics of mitochondrial respiration and energy status have been studied during growth and senescence of mung bean ( Phaseolus radiatus L.) cotyledons. The results showed that mitochondrial oxygen consumption, respiratory control, ADP:O ratios, and energy charge changed in the cotyledons during germination and growth of the seedlings. The respiration rate of intact cotyledons approximately reflected the trend of the oxidative activities of the isolated mitochondria. An increase was observed in both whole cotyledon respiration and mitochondrial oxygen uptake at the onset of senescence of mung bean cotyledons (day 3 after germination), which thereafter declined gradually. The capacity and activity of the alternative pathway increased markedly in mitochondria isolated from senescent cotyledons. After the onset of senescence, the mung bean cotyledon mitochondria exhibited a decrease both in the respiratory control ratios and ADP:O ratios, and the cotyledons exhibited a gradual decline in energy charge. All these results showed an irreversible deterioration of energy conservation in mung bean cotyledons. The role(s) of the alternative pathway in senescent mung bean cotyledons is discussed.     

Inhibition of Gibberellin Production in the Fungi Gibberella fujikuroi and Sphaceloma manihoticola by Plant Growth Retardants

The effect of different types of plant growth retardants on fungal gibberellin (GA) formation has been studied in cultures of Gibberella fujikuroi and Sphaceloma manihoticola. Quaternary ammonium compounds (chlormequat chloride, mepiquat chloride, Amo-1618), triazoles (uniconazole and several experimental compounds), and the norbornanodiazetine tetcyclacis inhibited GA biosynthesis in both fungal species. Concentrations between 2 × 10⁻⁴ and 10⁻⁹m were required for a 50% inhibition of the production of gibberellin A₃ in Gibberella fujikuroi and of giberellin A₄ in Sphaceloma manihoticola. The formation of other prominent GAs was affected at a similar degree of intensity. Tetcyclacis was the most active compound in both fungi. Compared to the growth retardants mentioned above, the biological activity of chlorphonium chloride was low. The acylcyclohexanediones prohexadione and LAB 198 999 had virtually no activity. Most likely, this lack of activity is due to a rapid metabolism of the compounds in the cultures. For the triazole-type compounds and tetcyclacis, a relatively distinct correlation exists in their ability to inhibit GA formation in fungal cultures, to block ent-kaurene oxygenase in a cell-free system, and to reduce shoot growth of rice seedlings. Due to differences in their metabolic fate and species specificities, such conclusions cannot be made for the other compounds.     

Comparative effects of some Amo-1618 analogs on gibberellin production in Fusarium moniliforme and on growth in higher plants

Summary The plant growth retardant 2-isopropyl-4-dimethylamino-5-methylphenyl-1-piperidinecarboxylate methyl chloride (Amo-1618) and three analogs (Carvadan, XII, XIII) were tested for ability to inhibit gibberellin production in Fusarium moniliforme and to suppress stem elongation in three species of higher plants.Amo-1618 and compound XII were highly effective in suppressing GA production in Fusarium whereas Carvadan and XIII were inactive. These inactive analogs were not degraded to any appreciable extent by Fusarium cultures.All four compounds suppressed stem growth although the relative effectiveness varied with the species tested. Carvadan was most active in dwarfing Phaseolus vulgaris and Pharbitis nil; compound XII was most active in Helianthus annuus.The lack of correlation between the ability to inhibit gibberellin production in Fusarium and to cause dwarfed growth in higher plants is interpreted to indicate that enzymes involved in gibberellin biosynthesis in different organisms exhibit certain variations which make them more or less sensitive to inhibitors such as the four growth retardants tested.     

Respiratory pathways in germinating maize radicles correlated with desiccation tolerance and soluble sugars

We have assessed the activities of the cytochrome and alternative pathways in total respiration and their role in each stage of germination of Zea mays L. radicles. Throughout imbibition, the salicylhydroxamic acid (SHAM) concentration needed to inhibit the cyanide-resistant pathway, without any side effects, decreased from 15 m M in quiescent embryos to 5 m M at 72 h after imbibition. Electrons predominantly flowed through the cytochrome pathway although the alternative pathway was already present at early imbibition. The capacity of the alternative path was about 70% of the control rate of respiration. Its engagement progressively increased from 18% after 10 min of imbibition to 70% at the radicle emergence and then decreased to 50% at 96 h after imbibition, concomitant with the onset of radicle growth. The alternative pathway was, however, not essential for germination. The observed activity of the alternative path correlated with the monosaccharide (glucose + fructose) content, suggesting that the alternative pathway could be acting according to the 'energy overflow model'. On the other hand, up to 24 h after imbibition at 16°C, maize radicles tolerate a severe desiccation, becoming intolerant at 72 h. On reimbibition of tolerant radicles, respiration increased immediately and the alternative pathway was rapidly engaged. At 72 h, no respiration was measured, indicating a total loss of the respiratory systems. The possible correlation between carbohydrate content, loss of desiccation tolerance and activity of the two respiratory pathways is discussed.     

Effect of salicylic acid on the alternative pathway of yellow lupine respiration

The effects of salicylic acid (SA) on the rate of respiration and the activity of cyanide-resistant sensitive to salicylhydroxamic acid oxidation pathway in detached etiolated cotyledons of yellow lupine (Lupinus luteus L.) and mitochondria isolated from these cotyledons were studied. Cotyledon treatment with 1 mM SA for 12 h increased the rate of oxygen uptake predominantly due to the activation of cyanide-resistant respiration (CRR) and alternative pathway of mitochondrial oxidation. It was established that the lupine genome encodes at least two isoforms of alternative oxidase (AO), LuAOX1 and LuAOX2, with the mol wt of about 35 kD. These proteins are always present in the mitochondria of etiolated lupine cotyledons, but their level increased rapidly after cotyledon treatment with SA, probably by increasing the mRNA content of the corresponding genes. SA-induced expression of Aox genes was correlated with the activation of CRR and an increase in the maximal activity (capacity) of AO in both detached yellow lupine cotyledons and mitochondria isolated from them.     

The embryonic axis controls lipid catabolism in cotyledons of apple seeds during germination

Activities of alkaline lipase (AlkL, EC 3.1.1.3), isocitrate lyase (ICL, EC 4.1.3.1), glucose 6-phosphate dehydrogenase (G6PDH, EC 1.1.1.49) and pyruvate kinase (PK, EC 2.7.1.40) were determined in embryos of apple ( Malus domestica Borb. cv. Antonówka) during culture in darkness or at 12 h photoperiod; in both cases either in the presence of gibberellin A₃ (GA₃) or AMO 1618 (inhibitor of GA synthesis). AlkL and ICL were stimulated by light and GA₃; light stimulation was reversed by AMO. G6PDH and PK were not affected by culture conditions. Almost all the activity of all enzymes was found in the cotyledons; only PK was distributed between axis and cotyledons. GA-like activity was found almost exclusively in the embryo axis. Cultured isolated cotyledons lost their sensitivity to light and AMO, but AlkL and ICL were still stimulated by GA₃. Translocation of GA from axis to cotyledons during the culture of embryos is postulated.     

Effects of Growth Retardants on the Growth of Excised Roots of Dolichos lablab L. in Culture

The effects of various growth-retarding chemicals on the growthof excised roots of Dolichos lablab in sterile culture are investigated.Application of a range of concentrations of CCC, phosfon, andB-995 inhibited growth in length of the roots by reducing thefrequency of cell division, with little or no effect on cellelongation. Treated roots had generally lower DNA, RNA, andprotein contents than controls, although CCC-treatment significantlyenhanced the soluble nitrogen content of the roots. The inhibitoryeffects of growth retardants were not reversed by IAA or GA,but substances like choline chloride and pyridoxine hydrochloridewere partially effective in reinstating normal growth in rootsgrowing in concentrations of CCC producing about 50 percentinhibition of growth in length.     

The level of flavonoids and amines in de-etiolated and methyl jasmonate treated seedling of common buckwheat

The effects of atmospheric methyl jasmonate on the level of flavonoids and biogenic amines in de-etiolated seedlings of common buckwheat (Fagopyrum esculentum Moench) were investigated. In cotyledons and hypocotyls of etiolated seedlings, some traces of anthocyanins were found, with no flavones and flavonols identified. A measurable content of flavones and flavonols was, however, determined in roots. De-etiolation process stimulated the accumulation of all flavonoid types. Methyl jasmonate clearly decreased the content of anthocyanins in the hypocotyl, not affecting their level in cotyledons. In case of roots, the content of anthocyanins increased after a 4-day treatment. In general, reduction in the level of flavones and flavonols was recorded only in the hypocotyl, however it was not always significant. Cotyledons of the seedlings treated with methyl jasmonate responded by a slight increase in flavonoids level. Methyl jasmonate considerably induced the accumulation of 2-phenylethylamine in all the seedling organs, increasing the content of putrescine and tryptamine in cotyledons, and decreasing the level of tryptamine in roots.     

Effects of Fluoride on Mitochondrial Activity in Higher Plants

The effects of fluoride on respiration of plant tissue and mitochondria were investigated. Fumigation of young soybean plants (Glycine max Merr. cv. Hawkeye) with 9–12 μg × m^?3 HF caused a stimulation of respiration at about 2 days of treatment followed by inhibition 2 days later. Mitochondria isolated from the stimulated tissue had higher respiration rates, greater ATPase activity, and lower P/O ratios, while in mitochondria from inhibited tissue, all three were reduced. Treatment of etiolated soybean hypocotyl sections in Hoagland's solution containing KF for 3 to 10 h only resulted in inhibition of respiration. Mitochondria isolated from this tissue elicited increased respiration rates with malate as substrate and inhibited respiration with succinate. With both substrates respiratory control and ADP/O ratios were decreased. Direct treatment of mitochondria from the etiolated soybean hypocotyl tissue with fluoride resulted in inhibition of state 3 respiration and lower ADP/O ratios with the substrates succinate, malate, and NADH. Fluoride was also found to increase the amount of osmotically induced swelling and cause a more rapid leakage of protein with mitochondria isolated from etiolated corn shoots (Zea mays L. cv. Golden Cross Bantam). The results are discussed with respect to possible effects of fluoride on mitochondrial membranes.