The relationship between the activities of the pentose phosphate pathway and glycolysis during early stages of floral induction in spinach

A quantitative cytochemical study was made of fructokinase, glucokinase, and fructokinase (both PFK-ATP and PFK-PP + F-2:6-P) activities in shoot apices of 4-week old Spinacia oleracea. The rates of activity of these enzymes in the central zone of the shoot apex of plants kept on a short day regime were compared with those from plants transferred from a range of timing up to 24 h to a continuous light regime when floral induction occurred. A mechanism is suggested explaining how no measurable change in activities of the enzymes assayed could still account for the availability of adequate levels G-6-P as substrate for pentose pathway activity which is almost doubled early on in cells of the central zone of shoot apices induced to flower.     

Enzymes of sucrose and hexose metabolism in developing kernels of two inbreds of maize

Tissue distribution and activity of enzymes involved in sucrose and hexose metabolism were examined in kernels of two inbreds of maize (Zea mays L.) at progressive stages of development. Levels of sugars and starch were also quantitated throughout development. Enzyme activities studied were: ATP-linked fructokinase, UTP-linked fructokinase, ATP-linked glucokinase, sucrose synthase, UDP-Glc pyrophosphorylase, UDP-Glc dehydrogenase, PPi-linked phosphofructokinase, ATP-linked phosphofructokinase, NAD-dependent sorbitol dehydrogenase, NADP-dependent 6-P-gluconate dehydrogenase, NADP-dependent Glc-6-P dehydrogenase, aldolase, phosphoglucoisomerase, and phosphoglucomutase. Distribution of invertase activity was examined histochemically. Hexokinase and ATP-linked phosphofructokinase activities were the lowest among these enzymes and it is likely that these enzymes may regulate the utilization of sucrose in developing maize kernels. Most of the hexokinase activity was found in the endosperm, but the embryo had high activity on a dry weight basis. The endosperm, which stores primarily starch, contained high PPi-linked phosphofructokinase and low ATP-linked phosphofructokinase activities, whereas the embryo, which stores primarily lipids, had much higher ATP-linked phosphofructokinase activity than did the endosperm. It is suggested that PPi required by UDP-Glc pyrophosphorylase and PPi-linked phosphofructokinase in the endosperm may be supplied by starch synthesis. Sorbitol dehydrogenase activity was largely restricted to the endosperm, whereas 6-P-gluconate and Glc-6-P dehydrogenase activities were highest in the base and pericarp. A possible metabolic pathway by which sucrose is converted into starch is proposed.     

A close temporal and spatial correlation between cell growth,cell wall synthesis and the activity of enzymes of mannan synthesis in Acetabularia mediterranea

The synthesis of cell wall mannan and the activities of guanosine-diphosphate-mannose-pyrophosphorylase (EC2.7.7.13) and mannan synthetase were studied during the development of nucleate and enucleated cells of the alga Acetabularia mediterranea. The activities of both enzymes are relatively high as long as the cells grow and synthesize mannans. With termination of growth and mannan synthesis, the activities of both enzymes, but especially of mannan synthetase, drop to a low value. Furthermore, the activities of both enzymes are distributed in the cell along an apical-basal gradient. High activities are present in the apical regions of the cell where growth and mannan synthesis mainly occur, whereas in the basal region, growth, mannan synthesis and the activity of the two enzymes are slight. Since the in vitro activity of GDP-Man-pyr is at least 100 times higher than that of mannan synthetase, it was concluded that mannan synthetase activity is the limiting factor in mannan synthesis. This conclusion is supported by the determined pool sizes of Fru 6-P, Man 6-P, Man 1-P and GDP-Man during the development of the cells. The control of mannan synthesis and with it cell wall formation and growth through the regulation of mannan synthetase activity is discussed.Abbreviations DD dark-dark regime - Fru 6-P fructose-6-phosphate - GDP-Man guanosine-diphosphate-mannose - GDP-Manpyr GDP-diphosphate-mannose-pyrophosphorylase - GTP guanosine-triphosphate - LD light-dark regime - Man 1-P mannose-1-phosphate - Man 6-P mannose-6-phosphate - TCA trichloracetic acid     

THE CONCEPT OF INCIPIENT VASCULAR TISSUE IN FERN APICES

A comparison of shoot apices of runners and rosettes of Nephrolepis with shoot apices of Adiantum indicates that a uniform concept of apical organization can be applied throughout. The region directly below the apical initials is designated a “central zone,” by analogy with that found in Lycopodium. The central zone is regarded as undifferentiated meristematic tissue rather than incipient vascular tissue.     

The anatomy of the shoot apex ofHyoscyamus niger L. reversed from the generative to the vegetative state

A study was made of the anatomical structure of the shoot apices ofHyoscyamus niger L. in plants which were transferred from a long-day to a short-day regime after the initiation of the inflorescence. After a certain time these plants are reverted to the vegetative stage with the inhibition of the development of further flower buds and the renewed production of rosette leaves. The inflorescence apex consisted of a few superficial layers of cells and a corpus composed of slightly elongated cells. The anatomical structure of the apices which were reverted into the vegetative state resembled that of shoot apices in the intermediate stage. The apex had several layers of small cells, under which there was a group of small but irregularly arranged cells which passed into the rib meristem. The shoot apices of plants transferred from a long to a short-day regime at different time intervals after fulfilling the requirements of minimal photoperiodic induction thus, on the short day, display morphological and anatomical characteristics of various degrees of transition from generative to vegetative stage.     

A MORPHOMETRY STUDY OF THE ULTRASTRUCTURE OF ECHINOCEREUS ENGELMANNII (CACTACEAE). II. THE MATURE,ZONATE SHOOT APICAL MERISTEM

The shoot apical meristems of adult Echinocereus engelmannii plants are zonate and have a tunica, central mother cells, a peripheral zone, and a pith-rib meristem. An ultrastructural, stereological study showed that each zone has its own distinct ultrastructure, but that the differences between the zones are quite small, both on a protoplasmic basis and on a cytoplasmic basis. Furthermore, the ultrastructure present in the adult apices differed only slightly from that which had been found in seedling apices, demonstrating a long-term stability of structure. The standard deviations found in the sample were small, indicating little variability from one plant to the next and suggesting that there are little or no cyclic changes during the plastochron or a 24-hr photoperiod. The ultrastructures found in the shoot apical meristems differed significantly and markedly from mature tissues of the same plants.     

The effect of 2,4-dichlorophenoxyacetic acid on the metabolic utilization of free carbohydrates in cucumber mosaic virus infected cucumber plants

The application of 2,4-dichlorophenoxyacetic acid at a concentration of 1.0 × 10⁻³ M toCucumis sativus brings about a decrease in the activity of carbohydrate catabolizing enzymes in the leaves of experimental plants. On the contrary, in CMV infected plants the activity of sucrase, glucokinase, fructokinase, phosphoglucoisomerase and glucose-6-phosphate dehydrogenase are enhanced at the same time. The application of 2,4-D to virus infected plants promotes this effect further, so that the activities of the enzymes investigated are twice as high as those in the control.     

Floral transition as a sequence of growth changes in different components of the shoot apical meristem ofChenopodium rubrum

The changes in cell division rate were studied in different components of the shoot apex ofChenopodium rubrum during short-day photoperiodic induction and after the inductive treatments. Induced and vegetative apices were compared. Accumulation of metaphases by colchicine treatment was used to compare the mean cell cycle duration in different components of the apex. A direct method of evaluating the increase in cell number obtained by anticlinal or periclinal divisions was applied if the corresponding components of induced and non-induced apices had to be compared. The short-day treatment prolonged the cell cycle more in the peripheral zone than in the central zone and still more in the leaf primordia. The importance of changing growth relations for floral transition was shown particularly if the induced plants were compared with the vegetative control with interrupted dark periods. Induced plants transferred to continuous light showed further changes in the rates of cell division. The cell cycle was shortened more in the central zone than in the peripheral zone,i.e. there was a further shift in growth relations within the apical dome. The cell cycle in the leaf and bud primordia was also shortened if compared with the vegetative control, the acceleration being stronger in the bud primordia. There was a subsequent retardation in cell division in the leaf primordia formed during and after the inductive treatment if the plants were fully induced. An inhibition of the oldest bud primordia was observed in fully induced apices, as well.     

HISTOCHEMICAL STUDY OF ENZYME ACTIVITY IN THE SHOOT APICAL MERISTEM OF BRASSICA CAMPESTRIS DURING TRANSITION TO FLOWERING. IV. GLUCOSE-6-PHOSPHATASE

Glucose-6-phosphatase (G6P) activity was determined in fresh-frozen, cryostat sections in the shoot apical meristem of Brassica campestris L. Enzymatic activity was differentially distributed in a zonate pattern in the vegetative meristem, but not in the transition and floral meristem. Vegetative apices showed a heterogenous localization with the highest activity in the central zone and the pith-rib meristem zone. At the early transition stage of development, G6P activity in the peripheral zone increased slightly. At the late transitional (prefloral) stage, G6P activity was not localized within the peripheral zone in island-like areas of activity. This is the first demonstration of G6P in shoot apical meristem at the vegetative, transition, and floral stage. The results indicate that G6P activity 1) is an accompanying event of evocation, but 2) does not mark incipient floral primordia. G6P may play an important role in the maintenance of glucose-6-phosphate homeostasis in an evoked shoot apical meristem.     

Restricted cell/cell communication in the shoot apex ofSilene coeli-rosa during the transition to flowering is associated with a high mitotic index rather than with evocation

Summary Plants ofSilene coeli-rosa given 5 or more long days (LDs) flowered, even when the LDs were followed by 48 hours of darkness before their return to short days (SDs). The mitotic indices of shoot apices from induced plants shortly after induction were significantly higher than the indices of shoot apices from vegetative plants. Two major mitotic peaks were observed in the shoot apices of plants given 7 long days (LDs) on day 8. One coincided with that reported byFrancis andLyndon (1979).Cell to cell movement was tested in the shoot apices of vegetative and LD treated plants using probes with a molecular size of 749 daltons (fluorescein-hexaglycine) and 847 daltons (fluorescein-leucyl diglutamyl leucine). These probes showed some movement in the shoot apices of both short day (SD) and LD treated plants, but fluorescein-leucyl diglutamyl leucine was immobile in the induced apices of 7 LD plants on day 8 at time intervals which coincided with major mitotic activity in the shoot apex. Symplasmic restriction in the shoot apex was also observed in plants given 8 LDs (i.e., plants not returned to SDs on day 7).In plants that were placed in 48 hours of darkness after the 7 LD treatment or in plants given 5 LDs, there was no strong peak in the mitotic index, even though all these LD treatments resulted in 100% flowering. In such plants no symplasmic restriction was found in the shoot. Thus the symplasmic restriction on day 8 of 7 LD plants is associated with the high mitotic index, but neither of these phenomena is an essential part of the evocation process.Abbreviations F(Glu)₂ L-glutamylglutamic acid conjugated to fluorescein isothiocyanate isomer I (F-) - F(Gly)₆ F-hexaglycine - FLGGL F-leucyl-diglutamyl-leucine - F(PPG)₅ F-the pentamer (propyl-propyl glycine) - LD long day - LDs long days - SD short day - SDs short days     

Histochemical investigation of dehydrogenases in shoot apices of wheat plants at different ontogenetic stages

The activity and localization of alcohol (AD), lactic (LD), malic (MD), isocitric (ICD) and succinic (SD) dehydrogenase, resp. was histochemically determined in shoot apices of plants in the vegetative condition during transition to flowering and at the reproductive state. The enzymes were determined in freehand sections, as well as in shoot apices incubatedin toto according toNachlas et al. (1958) using Nitro-BT. The characteristic localization of the enzymes depending on the ontogenic stage of the shoot apex is described. Different parts, layers and cell groups of the apex showed differences in enzyme activity. SD could not be detected in shoot apices. The histochemical findings are discussed in relation to former results obtained from a biochemical investigation of respiration in the same material.     

Pathways of d-fructose and d-glucose catabolism in marine species of Alcaligenes,Pseudomonas marina,and Alteromonas communis

Cell-free extracts of d-fructose grown cells of marine species of Alcaligenes as well as Pseudomonas marina contained an activity which catalyzed a P-enolpyruvate-dependent phosphorylation of d-fructose in the 1-position as well as activities of the following enzymes: 1-P-fructokinase, fructose-1,6-P₂ aldolase, PPi-dependent 6-P-fructokinase, fructokinase, glucokinase, P-hexose isomerase, glucose-6-P dehydrogenase, 6-P-gluconate dehydrase, and 2-keto-3-deoxy-6-P-gluconate aldolase. The presence of these enzyme activities would allow d-fructose to be degraded by the Embden-Meyerhof pathway and/or the Entner-Doudoroff pathway. In cell-free extracts of d-glucose grown cells, the activity catalyzing a P-enolpyruvate-dependent phosphorylation of d-fructose as well as 1-P-fructokinase activity were reduced or absent while the remaining enzymes were present at levels similar to those found in d-fructose grown cells. Radiolabeling experiments suggested that both d-fructose and d-glucose were utilized primarily via the Entner-Doudoroff pathway. Alteromonas communis, a marine species lacking 1-P-fructokinase and the PPi-dependent 6-P-fructokinase, contained all the enzyme activities necessary for the catabolism of d-fructose and d-glucose by the Entner-Doudoroff pathway; the involvement of this pathway was also consistent with the results of the radiolabeling experiments.Non-Standard Abbreviations EDP Entner-Doudoroff pathway - EMP Embden-Meyerhof pathway - FDP fructose-1,6-P₂ - FDPase FDP phosphatase - F-1-P fructose-1-P - F-6-P fructose-6-P - FPTS PEP: d-fructose phosphotransferase system - PPi-6-PFK PPi dependent 6-PFK - G-6-P glucose-6-P - KDPG 2-keto-3-deoxy-6-P-gluconate - PEP P-enolpyruvate - 1-PFK 1-P-fructokinase - 6-PFK 6-P-fructokinase - 6-PGA 6-P-gluconate     

Mechanism of preventive effects of exendin-4 and des-fluoro-sitagliptin in a murine model of fructose-induced prediabetes

Protective effects of exendin-4 (glucagon-like peptide-1 -GLP-1- receptor agonist) and des-fluoro-sitagliptin (dipeptidyl peptidase-4 inhibitor) on fructose-induced hepatic disturbances were evaluated in prediabetic rats. Complementary, a possible direct effect of exendin-4 in human hepatoblastoma-derived cell line HepG2 incubated with fructose in presence/absence of exendin-9-39 (GLP-1 receptor antagonist) was investigated. In vivo, after 21 days of fructose rich diet, we determined: glycemia, insulinemia, and triglyceridemia; hepatic fructokinase, AMP-deaminase, and G-6-P dehydrogenase (G-6-P DH) activities; carbohydrate-responsive element-binding protein (ChREBP) expression; triglyceride content and lipogenic gene expression (glycerol-3-phosphate acyltransferase -GPAT-, fatty acid synthase -FAS-, sterol regulatory element-binding protein-1c -SREBP-1c); oxidative stress and inflammatory markers expression. In HepG2 cells we measured fructokinase activity and triglyceride content. Hypertriglyceridemia, hyperinsulinemia, enhanced liver fructokinase, AMP-deaminase, and G-6-P DH activities, increased ChREBP and lipogenic genes expression, enhanced triglyceride level, oxidative stress and inflammatory markers recorded in fructose fed animals, were prevented by co-administration of either exendin-4 or des-fluoro-sitagliptin. Exendin-4 prevented fructose-induced increase in fructokinase activity and triglyceride contain in HepG2 cells. These effects were blunted co-incubating with exendin-9-39. The results demonstrated for the first time that exendin-4/des-fluro-sitagliptin prevented fructose-induced endocrine-metabolic oxidative stress and inflammatory changes probably acting on the purine degradation pathway. Exendin 9–39 blunted in vitro protective exendin-4 effects, thereby suggesting a direct effect of this compound on hepatocytes through GLP-1 receptor. Direct effect on fructokinase and AMP-deaminase activities, with a key role in the pathogenesis of liver dysfunction induced by fructose, suggests purine degradation pathway constitute a potential therapeutic objective for GLP-1 receptor agonists.     

T-DNA integration into genomic DNA of rice following Agrobacterium inoculation of isolated shoot apices

This paper establishes that the isolated shoot meristem of monocotyledons can be infected and transformed using Agrobacterium. Since this explant from nearly any cereal cultivar can rapidly regenerate into a plant, using this explant effectively eliminates the genotype regeneration restrictions to cereal crop transformation allowing direct transformation of elite germplasm. Shoot apices of Oryza sativa L. Tropical Japonica, cv. Maybelle were explants used for cocultivation, and gene transfer was accomplished using Agrobacterium containing plasmids for the bar gene expression driven by the CaMV 35S promoter or by the rice actin 1 promoter. Experiments to determine the survival rates of isolated shoot apices on media containing the herbicide, glufosinate-ammonium (PPT), established that no shoot apices survived on 0.5 or 1.0 mg/l PPT. After shoot apices were cocultivated with Agrobacterium, 2.8% (overall 20 out of 721 shoot apices) survived on 0.5 mg/l PPT. Results demonstrated that the use of the actin 1 promoter-based expression vector and an extra-wounding treatment of the meristematic cells appeared to be most effective in promoting transformation. Integration, expression and transmission of the transferred foreign genes in primary, R1 and R2 generation plants were confirmed by molecular analyses and herbicide application tests. A germination test of R2 progeny from one of the transgenic plants (R1) established a phenotype segregation ratio showing a non-Mendelian inheritance pattern. Inactivation of the transferred foreign gene in R2 progeny appeared to result from transgene methylation.     

Zinc inhibition of hepatic fructose metabolism in rats

The ability of Zn to modulate key metabolic processes was investigated in a study of gluconeogenesis in isolated hepatocytes from fasted rats. Zn (100 μM) inhibited glucose production from fructose by 41%, sorbitol by 28%; glycerol by 17%, and glyceraldehyde by 26%. Maximum inhibition of gluconeogenesis from fructose occurred at 25 μM Zn. Zn inhibited the rate of lactate production from fructose by 24% but not from sorbitol, glycerol, or glyceraldehyde. Fructose uptake by hepatocytes was not affected by Zn. A positive linear relationship (r=0.994) was obtained between inhibition by Zn of glucose and lactate production, indicating that a common step in both pathways is inhibited by Zn. The effect of Zn on fructokinase, aldolase-B, and triokinase activities was determined on semipurified rat liver enzyme preparations. Zn had no affect on triokinase activity but inhibited the two other enzymes in a dose-dependent manner, with the inhibition of aldolase-B being much greater than of fructokinase for concentrations of Zn between 2.5 and 20 μM. Zn increased the intracellular concentration of fructose-1-P in hepatocytes incubated with fructose, indicating a more potent Zn inhibition of aldolase-B than fructokinase. In addition, hepatocytes treated with Zn had decreased ATP and ADP concentrations, but had normal energy charge, suggesting an effect of Zn on adenine nucleotide degradation or synthesis. The demonstration that Zn inhibits two enzymes in fructose metabolism adds to the growing list of metabolic pathways that are catalyzed by enzymes that are sensitive to Zn.     

Efficient <Emphasis Type="Italic">in vitro</Emphasis> plant regeneration from shoot apices and gene transfer by particle bombardment in <Emphasis Type="Italic">Jatropha curcas</Emphasis>

An efficient and reproducible in vitro plant regeneration system from shoot apices was developed in Jatropha curcas. Benzylaminopurine (BAP; 2.5 μM) was most effective in inducing an average of 6.2 shoots per shoot apex. Incorporation of gibberellic acid (GA3; 0.5 μM) to basal medium was found essential for elongation of shoots. The BAP-habituated mother explants continuously produced shoots during successive subculture without any loss of morphogenic potential. The shoots rooted efficiently on half-strength MS medium. The rooted plantlets were acclimatized with more than 98 % success and the plants transferred to soil:compost in nursery showed no sign of variation compared to the seed-grown plants. The whole process of culture initiation to plant establishment was accomplished within 5–6 weeks. A genetic transformation system in J. curcas was established for the first time, using bombardment of particles coated with plasmid pBI426 with a GUS-NPT II fusion protein under the control of a double 35S cauliflower mosaic virus (CaMV) promoter. The β-glucuronidase (GUS) activity in J. curcas shoot apices was significantly affected by the gold particle size, bombardment pressure, target distance, macrocarrier travel distance, number of bombardments, and type and duration of osmotic pre-treatment. The proliferating bombarded shoot apices were screened on medium supplemented with 25 mg dm⁻³ kanamycin and surviving shoots were rooted on medium devoid of kanamycin. The integration of the transgene into genomic DNA of transgenic plants was confirmed by PCR and Southern blot hybridization. The transgenic plants showed insertion of single to multiple copies of the transgene.     

THE EFFECTS ON CHRONIC GAMMA RADIATION ON THE INTERNAL APICAL CONFIGURATIONS OF THE VEGETATIVE SHOOT APEX OF COLEUS BLUMEI

Clonal plants of a bushy variety ‘Trailing Queen’ of Coleus blumei Benth. were exposed to 600–700 R per day for 28 days. Shoot apices of control and irradiated plants were studied microscopically. The normal vegetative shoot apex has two tunica layers and a corpus which is partly layered and partly composed of randomly divided cells. Five cytohistological zones were observed, including a cup-shaped zone, imposed upon the tunica-corpus relationships. Response to gamma radiation resulted in abnormal internal apical configurations. All irradiated shoots was found to have the same internal configurations. Only a single tunica layer broken by occasional periclinal divisions was observed. The cytohistological organization of irradiated apices was different from that seen in the normal shoot tips in that two zones were found. Evidence of a radiosensitivity gradient within the vegetative shoot apex was also observed. The possible significance of the results is discussed.     

Amelioration of salinity tolerance in <Emphasis Type="Italic">Solanum tuberosum</Emphasis> L. by exogenous application of ascorbic acid

The present investigation envisaged revealing the role of exogenous application of ascorbic acid in increasing resistance against NaCl stress. Shoot apices from 60-d-old, in vitro-grown plants of two commercially important cultivars of Solanum tuberosum L., cvs. Desiree and Cardinal, were inoculated on Murashige and Skoog (MS) medium supplemented with 0.5 mM ascorbic acid for 72 h as a pretreatment. Pretreated and non-pretreated shoot apices were transferred to MS medium containing different concentrations of NaCl (0–140 mM; eight treatments). Results were recorded for morphological (shoot length, shoot number, root length, root number, and number of nodes) and biochemical features (protein, peroxidase, catalase, and superoxide dismutase activities) after 60 d of salt treatment. Similarly, 60-d-old, well-proliferated callus cultures were also pretreated with ascorbic acid for 24 h and transferred to an optimized callus proliferation medium containing different concentrations of salt. Results were recorded after 60 d of salt treatment for percentage relative fresh weight growth and biochemical parameters. Salinity severely inhibited all the growth parameters in both the cultivars. Pretreatment with ascorbic acid to both salt-treated plants and callus cultures showed significant differences with respect to almost all of the growth and biochemical parameters studied. Protein content as well as catalase and superoxide dismutase activities increased significantly in both the cultivars, although peroxidase activity showed a decreasing trend in ascorbic acid-pretreated plants as well as callus cultures.     

Auxin gradient along the root of the maize seedling

[5-³H]Indol-3yl-acetic acid (IAA) applied to the shoot apices of intact 6-day-old maize (Zea mays L.) plants moved into the primary root and accumulated at the root apex. IAA from the shoot could partially satisfy the requirement of the primary root for IAA for growth.Abbreviation IAA indol-3yl-acetic acid     

Growth correlations and rna synthesis in different parts of the shoot apical meristem ofChenopodium rubrunt L. induced to flowering

Uridine-³H incorporation and RNA concentration were investigated in different parts of the shoot apical meristem ofChenopodium rubrum using autoradiography and cytophotometry. A single inductive cycle was sufficient to bring about postinductive first events in the shoot apex but not for complete flower differentiation. The initial activation of RNA synthesis manifested itself in all zones of the apex. The first increase was more conspicuous in the peripheral than in the central zone. The indications of the first events in the apices after a single inductive cycle disappear prior to morphological reversal to the vegetative state. Induction by three short days led to rapid flower differentiation. The increase in RNA synthesis and concentration was most conspicuous in the central zone in this case. The ratio of RNA synthesis and content between bud and leaf primordia (B/L) also change in relation to photoperiodic induction. In vegetative plants the B/L ratio was low while after induction it increased. The shifts in activity of RNA synthesis observed in the shoot apical meristem are related to the changes in growth activity of the different parts of the apex. The growth ratios in the apices bear the character of growth correlations. The change in the growth correlations following photoperiodic induction together with the total activation of RNA synthesis are considered to represent one of the first events of the transition to the reproductive state.