Arginase,Arginine Decarboxylase,Ornithine Decarboxylase,and Polyamines in Tomato Ovaries (Changes in Unpollinated Ovaries and Parthenocarpic Fruits Induced by Auxin or Gibberellin)

Arginase (EC 3.5.3.1) activity has been found in the ovaries and Young fruits of tomato (Lycopersicon esculentum Mill. cv Rutgers).Changes in arginase, arginine decarboxylase (EC 4.1.1.19), and ornithine decarboxylase activity (EC 4.1.1.17) and levels of free and conjugated putrescine, spermidine, and spermine were determined in unpollinated ovaries and in parthenocarpic fruits during the early stages of development induced by 2,4-dichlorophenoxyacetic acid (2,4-D) or gibberellic acid (GA3). Levels of arginase, free spermine, and conjugates of the three polyamines were constant in unpollinated ovaries and characteristic of a presenescent step. A marked decrease in arginase activity, free spermine, and polyamine conjugates was associated with the initiation of fruit growth due to cell division, and when cell expansion was initiated, the absence of arginase indicated a redirection of nitrogen metabolism to the synthesis of arginine. A transient increase in arginine decarboxylase and ornithine decarboxylase was also observed in 2,4-D-induced fruits. In general, 2,4-D treatments produced faster changes than GA3, and without treatment, unpollinated ovaries developed only slightly and senescence was hardly visible. Sensitivity to 2,4-D and GA3 treatment remained for at least 2 weeks postanthesis.     

Sucrose‐cleaving enzymes and carbohydrate pools in Lilium longiflorum floral organs

The activities of soluble invertase (EC 3.2.1.26), cell wall invertase (EC 3.2.1.26) and sucrose synthase (EC 2.4.1.13) were determined in Easter lily ( Lilium longiflorum Thunb. cv. Nellie White) floral organs during flower development. These enzyme activities were correlated with dry weight gains and carbohydrate pools to investigate the importance of their expression in maintaining sink strength of floral organs. In the early stages of flower bud development, anthers exhibited the highest rates of dry weight gain and activity of sucrolytic enzymes. Once anther growth was completed, the dry weight gain of tepal, filament, stigma and style increased with a concomitant increase in hexose concentrations and invertase activity. Although all three enzymes capable of catalyzing sucrose cleavage were present in every flower organ of L. longiflorum , soluble invertase was the predominant enzyme in all flower organs except stigma where cell wall invertase dominated. Soluble invertase activity was highly correlated with dry weight gain in most of the flower organs.     

Amelioration in secretion of hyperthermostable and Ca2+ -independent alpha-amylase of Geobacillus thermoleovorans by some polyamines and their biosynthesis inhibitor methylglyoxal-bis-guanylhydrazone

AIM: Effect of polyamines and their biosynthesis inhibitors on the production of hyperthermostable and Ca2+ -independent alpha-amylase by Geobacillus thermoleovorans MTCC 4220. METHODS AND RESULTS: The alpha-amylase was produced in starch-yeast extract-tryptone (SYT) broth with different polyamines (PA) and polyamine biosynthesis inhibitors, methylglyoxal-bis-guanylhydrazone (MGBG) and cyclohexylammonium sulphate (CHA) at 70 degrees C. The bacterial pellets were obtained after growing G. thermoleovorans at different temperatures, and used in determining total PA. The cell-free culture filtrates were used in alpha-amylase assays. During growth, total polyamines in biomass increased till 2 h, and thereafter, decreased gradually. The total polyamine content was very high in the biomass cultivated at 55 degrees C when compared with that of higher temperatures. Enzyme titre enhanced up to 70 degrees C, and thereafter declined. Extracellular enzyme and protein levels declined in the presence of exogenously added PA. The intracellular enzyme titres, however, were higher in putrescine (put) and spermidine (spd) than in spermine (spm). Polyamine biosynthesis inhibitor, MGBG enhanced secretion of alpha-amylase in a laboratory fermentor as well as shake flasks, although CHA did not affect it. CONCLUSIONS: The intracellular accumulation of put in the presence of MGBG appeared to enhance synthesis and secretion of alpha-amylase. Extracellular enzyme and protein levels were low in the presence of exogenously added PA, but their intracellular levels, however, were higher in put and spd than in spm. SIGNIFICANCE AND IMPACT OF THE STUDY: A substantial increase in the synthesis and secretion of alpha-amylase was attained in G. thermoleovorans in the presence of polyamine biosynthesis inhibitor MGBG.     

Involvement of polyamines in the control of fruitlet physiological abscission in grapevine (Vitis vinifera)

The potential contribution of polyamines (PAs) in the regulation of physiologically induced fruitlet abscission was investigated in cuttings from two cultivars of Vitis vinifera L., Pinot noir (PN) and Merlot (MRT). Abscission was higher in MRT than in PN and was preceded by a decrease in free PA levels. This decline was more pronounced in inflorescences than in leaves of the sensitive cultivar. Soluble conjugated PA showed an opposite trend in both cultivars. This suggests a cause-effect relationship between free and/or conjugated PA levels in floral organs and susceptibility to abscission. Spermidine (Spd), but not putrescine (Put) or diaminopropane, supplied at 0.5–1 m M to the nutritive medium prior to the anthesis, increased free and conjugated PA levels in the inflorescences and markedly inhibited abscission. α -Difluoromethylarginine, an inhibitor of arginine decarboxylase, but not α -difluoromethylornithine, an inhibitor of ornithine decarboxylase, lowered PA levels and increased abscission. Treatment with cyclohexylamine or β -hydroxyethylhydrazine as potent inhibitors of Spd synthase and PA oxidases, respectively, reduced the Spd and/or spermine levels and enhanced free Put in the inflorescences, inducing an increased abscission of floral organs shortly after anthesis. These data suggest that PAs, particularly Spd, could be involved in the regulation of grapevine fruitlet physiological abscission.     

Role of gibberellins in parthenocarpic fruit development induced by the genetic system pat-3/pat-4 in tomato

The role of gibberellins (GAs) in the induction of parthenocarpic fruit-set and growth by the pat-3/pat-4 genetic system in tomato ( Lycopersicon esculentum Mill.) was investigated using wild type (WT; Cuarenteno) and a near-isogenic line derived from the German line RP75/59 (the source of pat-3/pat-4 parthenocarpy). Unpollinated WT ovaries degenerated but GA₃ application induced parthenocarpic fruit growth. On the contrary, parthenocarpic growth of pat-3/pat-4 fruits, which occurs in the absence of pollination and hormone treatment, was not affected by applied GA₃. Unpollinated pat-3/pat-4 fruit growth was negated by paclobutrazol, an inhibitor of ent -kaurene oxidase, and this inhibitory effect was negated by GA₃. The quantification of the main GAs of the early 13-hydroxylation pathway (GA₁, GA₈, GA₁₉, GA₂₀, GA₂₉ and GA₄₄) in unpollinated ovaries at 3 developmental stages (flower bud, FB; pre-anthesis, PR; and anthesis, AN), by gas chromatography-selected ion monitoring, showed that the concentration of most of them was higher in pat-3/pat-4 than in WT ovaries at PR and AN stages. The concentration of GA₁, suggested previously to be the active GA in tomate, was 2–4 times higher. Unpollinated pat-3/pat-4 ovaries at FB, PR and AN stages also contained relatively high amounts (5–12 ng g⁻¹) of GA₃, a GA found at less than 0.5 ng g⁻¹ in WT ovaries. It is concluded that the mutations pat-3/pat-4 may induce natural facultative parthenocarpy capacity in tomato by increasing the concentration of GA₁ and GA₃ in the ovaries before pollination.     

Polyamine content as a marker of radiation injury in the rat spleen

The modifications of the polyamines putrescine (put), spermidine (spd) and spermine (spm) in rat spleen after 3 Gy whole-body irradiation were studied. Rats were irradiated at four different times of the day (00.00, 06.00, 12.00 and 18.00) and sacrificed between 12 h and 62 days after irradiation. Control animals, sacrificed at the same times of the day, showed higher levels of the spd/spm ratio during the hours of light. After irradiation the polyamine content was rapidly and significantly reduced over a period of 20 days. The modification of the amount of spm lasted for a longer period of time. Normal values of polyamine content were reached at later times when the mitotic activity was restored. The results show a close correlation between polyamine concentration and [3H]thymidine uptake.     

Polyamines and Flower Development in the Male Sterile Stamenless-2 Mutant of Tomato (Lycopersicon esculentum Mill.) : I. Level of Polyamines and Their Biosynthesis in Normal and Mutant Flowers

The levels of free putrescine, spermidine, and spermine, and the activities of ornithine decarboxylase and s-adenosylmethionine decarboxylase were determined in the floral organs of the normal and a male sterile stamenless-2 (sl-2/sl-2) mutant of tomato (Lycopersicon esculentum Mill.). Under the intermediate temperature regime, all mutant floral organs possessed significantly higher levels of polyamines and enzyme activities than their normal counterparts. In the low temperature-reverted mutant stamens, the polyamine levels and the activity of PA biosynthetic enzymes were not significantly different from the normal. It is suggested that the abnormal stamen development in the sl-2/sl-2 mutant is, in part, related to elevated levels of endogenous PAs.     

Effects of nutrient contents and defense compounds on herbivory in reproductive organs and leaves of Iris gracilipes

Optimal defense theory (ODT) states that the plant tissue with the highest value to fitness will receive the most protection compared with other plant parts. ODT can be applied to the differences in defenses among floral organs, although most studies have concentrated on the comparison between leaves and flowers. Using Iris gracilipes, we investigated whether ODT is supported when primary and accessory floral organs and leaves are distinguished. We found that anthers and perianths tended to be attacked more severely than ovaries and leaves in the bud and flower stages and that anthers contained the highest nitrogen and phosphorus concentrations. Although ovaries were also found to contain high nitrogen and phosphorus concentrations, they were less severely attacked by herbivores than anthers, perhaps because ovaries contained the highest condensed tannins concentrations among the floral organs except for perianths in the flower stage. Thus, noting that the number of ovules is very much smaller than that of pollen grains, we concluded that ovaries are the most intensively protected, consistent with the prediction of ODT as applied to floral organs. ODT is applicable to the difference in defense allocation among floral organs.     

Polyamine Content and Action in Roots of Zea mays L. in Relation to Aerenchyma Development

Roots of Zea mays L. developed more aerenchyma (cortical gas-filledspace) when partially deficient in oxygen (3 kPa) than whensupplied with air (20·8 kPa oxygen) in association withfaster production of ethylene (ethene). The possibility wastested that the additional ethylene production resulted fromdecreases in spermidine (spd) and spermine (spm) which share,with ethylene, a common precursor, S-adenosylmethionine. However,no decreases in spd and spm were seen in root tissue up to 4d-old. Removing oxygen completely also had little effect onspd and spm, but strongly suppressed both ethylene productionand aerenchyma formation. Partial oxygen shortage (3 kPa) increased the concentrationof putrescine (put), the precursor of spd and spm. This increasewas not a response to the extra ethylene formed by such rootssince ethylene treatment did no reproduce the effect. Applicationof inhibitors of put biosynthesis, difluoromethylarginine anddifluoromethylornithine, led to increased aerenchyma formation.Exogenous put inhibited the development of aerenchyma whilestimulating rather than inhibiting ethylene production, whentested in either air or 3 kPa oxygen. Thus, put appears to limitaerenchyma formation by suppressing ethylene action rather thanits production.Copyright 1993, 1999 Academic Press Ethylene, ethene, roots, aerenchyma, polyamines, oxygen shortage, anaerobiosis, environmental stress, Zea mays     

Polyamine metabolism during the growth cycle of tobacco BY-2 cells.

We studied polyamine (PA) biosynthesis, oxidation and conjugation in asynchronously dividing cells of tobacco BY-2 cell suspension culture (Nicotiana tabacum L.) during 7-day growth cycle. We analyzed the levels of free and conjugated PAs and the activities of biosynthetic and catabolic enzymes during the subculture interval. The contents of free spermidine and spermine started to increase after the inoculation into the fresh medium, positively correlated with the mitotic activity of BY-2 cells and reached their maxima at the beginning of exponential phase on day 3. On the contrary, the endogenous level of free Put showed a transient decline in the lag-phase, and then increased till the end of exponential phase (day 5). The time-course of the content of PCA-soluble conjugates showed a trend similar to that of the free PAs. The inoculation of BY-2 cells into the fresh medium resulted in a sharp increase in the activities of ornithine decarboxylase (ODC, EC 4.1.1.17) and S-adenosylmethionine decarboxylase (SAMDC; EC 4.1.1.50). Arginine decarboxylase (ADC; EC 4.1.1.19) activity remained low during the whole subculture interval. The rise of diamine oxidase (DAO; EC 1.4.3.6) in the first day after subculture coincided with the decrease in free Put level. De novo synthesis of PAs in BY-2 cells after inoculation into the fresh medium and the participation of both PA conjugation with hydroxycinnamic acids and Put oxidative degradation in maintaining of free PA levels during the growth cycle are discussed.     

Changes in Polyamine Biosynthesis Associated with Postfertilization Growth and Development in Tobacco Ovary Tissues

Polyamine (PA) titers and the activities of arginine decarboxylase (ADC, EC 4.1.1.19) and ornithine decarboxylase (ODC, EC 4.1.1.17), enzymes which catalyze rate-limiting steps in PA biosynthesis, were monitored during tobacco ovary maturation. In the period between anthesis and fertilization, the protein content of ovary tissues rapidly increased by about 40% and was accompanied by approximately a 3-fold increase in ODC activity, while ADC activity remained nearly constant. PA titers also remained relatively unchanged until fertilization, at which time they increased dramatically and the DNA content of ovary tissues doubled. This increase in PA biosynthesis was correlated with a further 3-fold increase in ODC activity, reaching a maximum 3 to 4 days after fertilization. During this time, ADC activity increased only slightly and accounted for approximately 1% of the total decarboxylase activity when ODC activity peaked. The postfertilization burst of biosynthetic activities slightly preceded a period of rapid ovary enlargement, presumably due to new cell division. During later stages of ovary development, DNA levels fell precipitously, while PA titers and decarboxylase activities decreased to preanthesis levels more slowly. In this period, growth producing a 300% increase in ovary fresh weight appears to be the result of cell enlargement.

Synchronous changes in PA titers and in the rates of PA biosynthesis, macromolecular synthesis, and growth in the tobacco ovary suggest that PAs may play a role in the regulation of postfertilization growth and development of this reproductive organ.

     

A yeast mitotic activator sensitises the shoot apical meristem to become floral in day-neutral tobacco

True day-neutral (DN) plants flower regardless of day-length and yet they flower at characteristic stages. DN Nicotiana tabacum cv. Samsun, makes about forty nodes before flowering. The question still persists whether flowering starts because leaves become physiologically able to export sufficient floral stimulus or the shoot apical meristem (SAM) acquires developmental competence to interpret its arrival. This question was addressed using tobacco expressing the Schizosaccharomyces pombe cell cycle gene, Spcdc25, as a tool. Spcdc25 expression induces early flowering and we tested a hypothesis that this phenotype arises because of premature floral competence of the SAM. Scions of vegetative Spcdc25 plants were grafted onto stocks of vegetative WT together with converse grafts and flowering onset followed (as the time since sowing and number of leaves formed till flowering). Spcdc25 plants flowered significantly earlier with fewer leaves, and, unlike WT, also formed flowers from axillary buds. Scions from vegetative Spcdc25 plants also flowered precociously when grafted to vegetative WT stocks. However, in a WT scion to Spcdc25 stock, the plants flowered at the same time as WT. SAMs from young vegetative Spcdc25 plants were elongated (increase in SAM convexity determined by tracing a circumference of SAM sections) with a pronounced meristem surface cell layers compared with WT. Presumably, Spcdc25 SAMs were competent for flowering earlier than WT and responded to florigenic signal produced even in young vegetative WT plants. Precocious reproductive competence in Spcdc25 SAMs comprised a pronounced mantle, a trait of prefloral SAMs. Hence, we propose that true DN plants export florigenic signal since early developmental stages but the SAM has to acquire competence to respond to the floral stimulus.     

In vivo inhibition of polyamine biosynthesis and growth in tobacco ovary tissues

Post fertilization growth of tobacco ovary tissues treated with inhibitors of polyamine (PA) biosynthesis was examined in relation to endogenous PA titers and the activities of arginine decarboxylase (ADC, EC 4.1.1.19) and ornithine decarboxylase (ODC, EC 4.1.1.17). DL-alpha-Difluoromethylornithine (DFMO) and DL-alpha-difluoromethylarginine (DFMA), specific, irreversible ("suicide") inhibitors of ODC and ADC in vitro, were used to modulate PA biosynthesis in excised flowers. ODC represented >99% of the total decarboxylase activity in tobacco ovaries. In vivo inhibition of ODC with DFMO resulted in a significant decrease in PA titers, ovary fresh weight and protein content. Simultaneous inhibition of both decarboxylases by DFMO and DFMA produced only a marginally greater depression in growth and PA titers, indicating that ODC activity is rate-limiting for PA biosynthesis in these tissues. Paradoxically, DFMA alone inhibited PA biosynthesis, not as a result of a specific inhibition of ADC, but primarily through the inactivation of ODC. In vivo inhibition of ODC by DFMA appears to result from arginase-mediated hydrolysis of this inhibitor to urea and DFMO, the suicide substrate for ODC. Putrescine conjugates in tobacco appear to function as a storage form of this amine which, upon hydrolysis, may contribute to Put homeostasis during growth.     

Phosphatidic acid is a major phospholipid class in reproductive organs of Arabidopsis thaliana

Phospholipids are the crucial components of biological membranes and signal transduction. Among different tissues, flower phospholipids are one of the least characterized features of plant lipidome. Here, we report that floral reproductive organs of Arabidopsis thaliana contain high levels of phosphatidic acid (PA), a known lipid second messenger. By using floral homeotic mutants enriched with specific floral organs, lipidomics study showed increased levels of PA species in ap3-3 mutant with enriched pistils. Accompanied gene expression study for 7 diacylglycerol kinases and 11 PA phosphatases revealed distinct floral organ specificity, suggesting an active phosphorylation/dephosphorylation between PA and diacylglycerol in flowers. Our results suggest that PA is a major phospholipid class in floral reproductive organs of A. thaliana.     

RETARDED PALEA1 controls palea development and floral zygomorphy in rice

Poaceae, one of the largest flowering plant families in angiosperms, evolved distinct inflorescence and flower morphology diverging from eudicots and other monocots. However, the mechanism underlying the specification of flower morphology in grasses remains unclear. Here we show that floral zygomorphy along the lemma-palea axis in rice (Oryza sativa) is partially or indirectly determined by the CYCLOIDEA (CYC)-like homolog RETARDED PALEA1 (REP1), which regulates palea identity and development. The REP1 gene is only expressed in palea primordium during early flower development, but during later floral stages is radially dispersed in stamens and the vascular bundles of the lemma and palea. The development of palea is significantly retarded in the rep1 mutant and its palea has five vascular bundles, which is similar to the vascular pattern of the wild-type lemma. Furthermore, ectopic expression of REP1 caused the asymmetrical overdifferentiation of the palea cells, altering their floral asymmetry. This work therefore extends the function of the TCP gene family members in defining the diversification of floral morphology in grasses and suggests that a common conserved mechanism controlling floral zygomorphy by CYC-like genes exists in both eudicots and the grasses.     

Polyamines and Plant Morphogenesis

Alterations in free and conjugated polyamines (PAs) and the enzymes involved in their biosynthesis, namely arginine decarboxylase, ornithine decarboxylase and S-adenosyl methionine decarboxylase have been reported to occur during cell division, growth, embryogenesis and rhizogenesis in an array of plant materials. However, the relationship, if any, between them and all these processes has not yet been established. It seems that specific PAs at specific concentration ranges are required during critical stages of growth and morphogenetic events. Furthermore, the effects of PA biosynthesis inhibitors vary considerably at different developmental stages of the same tissue. The present review deals with the available information about the possible role of PAs in aforesaid physiological processes.     

Expression study of five genes involved in floral organ development in multiple seeded rice

Rice cultivar Jugal is a unique floral organ mutant from South Bengal and Odisa, the two sister states of Eastern India, carries more than one kernels in most of its spikelet. Most of the mature florets of this line possess more than one carpal which later developed into more than one kernels within a single grain on maturity. In order to study the role of floral organ development genes commonly involved, expression study of five selected floral organ developmental genes (OsMADS3, OsMADS13, OsMADS21, OsMADS58, and DL) were studied through real time based quantitative PCR for three consecutive flower organ developmental stages (Sp5, Sp6, and Sp7) with reference to a normal rice line (IR36). All the studied genes showed differential relative expression in respect to the reference gene both in mutant and normal rice lines for the studied genes and stages and individual distinct pattern except DL gene which was almost similar in both Jugal and IR36 at early stage of floral organ development viz Sp5 and Sp6 stage. However, after Sp6 stage the expression is reduced in the normal rice (IR36) but in case of the mutant rice (Jugal) the expression started to increase and at Sp7 the expression level was much higher in the mutant line. The information resulted from the investigation form the basic idea on regulatory aspects of floral organ development in rice.