Mechanism of export of organic material from the developing fruits of pea

Pisum sativum L. fruits export a small quantity of radiolabeled substances to other plant parts after the fruits are allowed to photosynthesize in the presence of ¹⁴CO₂. Export was uninhibited by peduncle girdling suggesting an apoplastic route for transport of material, presumably by `reverse' flow in the peduncle xylem. To determine if any diurnal water potential gradient formed between pea leaves and fruit might be responsible for the observed export, the water potentials of the various organs were monitored over 24 hours. Water potential differences of up to 7.5 bars existed between leaves and fruit in long photoperiods, and up to 2.5 bars in short photoperiods. Pulses of <sup>14</sup>CO<sub>2</sub> labeling indicated that initial delivery of exported label was to `transpirational sinks,' with subsequent redistribution of label to metabolic sinks. Export to the apical bud appeared to be direct via the xylem. Application of membrane-impermeable inulin to a surgically opened seed coat `cup' resulted in export mainly to the subtending leaf with little redistribution. Simultaneous application of sucrose to the seed coat resulted in more extensive distribution of the sucrose, consistent with reloading of the sucrose into mature leaf phloem. Thus, export of material from fruits appears to occur via a xylem pathway in response to transpirationally derived water potential gradients.     

The transport of substances out of developing fruits in relation to the induction of apical senescence in Pisum sativum line G2

Apical sencscence in G2 peas occurs only in long days in the presence of fruit. The effect of fruits could be caused by the export of a senescence hormone from the fruits to the shoot tip. Export of radiolabeled material from developing fruits of G2 peas grown in long days was therefore examined following injection of the pods with [¹⁴C]-sucrose, [¹⁴C]-acetate, or [¹⁴C]-mevalonate or after allowing the pods to photosynthesize in ¹⁴CO₂ for 48 h. In all cases a small amount (<1%) of radioactivity was exported, primarily to the younger fruits on the same side of the plant and the to the shoot apex. After feeding ¹⁴CO₂ to the fruit, the radiolabeled material partitioned into acidic ethyl acctate and possessed a carboxyl group. While this radioactivity had chromatographic properties similar to abscisic acid (ABA) in a number of solvent systems, it was not identical to either ABA, phascic acid or dihydrophaseic acid. The nature of the labeled material found in the apex was different in short days, in which senescence does not occur, or when the leaves were the source of the radioactive compounds. The labeled material in the apex was similar after feeding ¹⁴CO₂, [¹⁴C]-acctatc, or [¹⁴C]-sucrose, but different if the fruits were injected with[¹⁴C]-mevalonate.Identification of the chemical nature of the labeled material in the apex was not possible due to the small amount present. Parallel purification of an extract from treated fruits led to the identification of N-benzoylaspartate and N-phenylacetyl-aspartate. The radiolabeled substance from the apex was run with these two chemically synthesized compounds on several gas chromatogtaphic columns, and was also recrystallized together several times. The label and the pure material did not have identical retention times; neither did they co-purify so that, while similar, the material exported to the apex is not the above compounds.     

Effect of photoperiod on polyamine metabolism in apical buds of g2 peas in relation to the induction of apical senescence

Polyamine content and arginine decarboxylase activity of apical buds were measured to determine whether polyamines are required to prevent apical senescence in pea. Polyamines were assayed as dansyl derivatives which were separated by reverse phase high performance liquid chromatography and detected by fluorescence spectrophotometry. High polyamine concentrations were found in the vigorous apices of plants grown under a short day photoperiod during which senescence is delayed. As the apex senesced in long days, the amounts of polyamines per organ declined in parallel with decreases in the size of the apical bud. However, a decrease in polyamine concentration, due mainly to a change in spermidine, occurred at the time of marked reduction in bud size and growth rate, but not before the onset of the early symptoms of senescence. No correlation was found with arginine decarboxylase. The results suggest polyamines may be required to support bud growth, but the photoperiodic mechanism which governs apical senescence of G2 peas does not exert control through polyamine metabolism.     

Photoperiodic and genetic control of carbon partitioning in peas and its relationship to apical senescence

Apical senescence but not flower initiation is delayed by short days (SD) compared to long days (LD) in pea plants (Pisum sativum L.) of genotype E Sn Hr. We recently reported that delay of senescence correlated with slower reproductive development, suggesting that fruits are weaker sinks for assimilates under delayed senescence conditions. Thus, we have examined assimilate partitioning in peas to determine if genotype and photoperiod regulate relative sink strength. Assimilate diversion by developing fruit has been implicated in senescence induction. A greater percentage of leaf-exported ¹⁴C was transported to fruits and a smaller percentage to the apical bud of G2 peas (genotype E Sn Hr) in LD than in SD. Relatively more of the ¹⁴C delivered to the apical bud of G2 peas was transported to flower buds than to young leaves in LD as compared to SD. There was no striking photoperiodic difference in carbon partitioning in genetic lines without the Sn Hr allele combination. The Sn Hr allele combination and photoperiod may regulate the relative strength of reproductive and vegetative sinks. Photoperiodic differences in sink strength early in reproduction suggest that these genes regulate sink strength by affecting the physiology of the whole plant. High vegetative sink strength in SD may maintain assimilate supply to the apical bud, delaying senescence.     

Serotonin; its possible relation to allergic disease

Increased amounts of serotonin as well as histamine have been found in the blood of animals during anaphylactic shock. Certain animals, particularly those in which antihistamines do not prevent anaphylaxis, have been found to have increased quantities of serotonin in the lung tissue during anaphylactic shock. Serotonin is a chemical derived from the amino acid tryptophan, which is widely distributed. It is excreted in the urine as the metabolite 5-hydroxyindoleacetic acid. Serotonin has been found in increased amounts in the blood of patients with carcinoids. The increase of serotonin in the blood and the finding of the excretory product in the urine has become a corroborative sign of the disease. The involvement of serotonin in the production of mental disease is evidenced by the effect of serotonin antagonists, which appear to influence mental behavior.That serotonin antagonists may be of ultimate value in the treatment of allergic disease is a possibility to be considered.     

Changes in developing pea (Pisum sativum) seeds in relation to their ability to withstand desiccation

In two separate experiments in 1970 and 1971 the germination ability of seeds that had been rapidly desiccated following harvesting from glasshouse-grown pea plants improved with time after fertilization. In 1970 the germination of seeds directly after harvesting also improved with time. In both years the readiness with which electrolytes were leached from dried seeds decreased with seed age, measured as time after fertilization. The germination ability of desiccated seeds, and of undried seeds in 1970, improved after the cessation of seed moisture increase on the parent plant. This improvement coincided, in 1970, with a fall in the readiness with which electrolytes could be leached from seeds and with a decline in the rate of oxygen uptake of seeds directly after harvest. Vital staining of dried seeds from harvest samples indicated the viability of the samples as determined by germination tests. In both years, when the cotyledons first showed complete staining, the staining was very intense; in 1970 this occurred in a harvest sample with a greater than 50% germination level, but in 1971 it occurred in a sample that showed incomplete staining in the embryo axis and less than 5 % germination. In 1971 the comparative rates of moisture loss from seeds, measured with a sensor element diffusion porometer, were very low at 25 days after fertilization, increased up to a peak at 35 days and fell to a rate at the final harvest (44 days) which was nearly three times faster than the initial rate. Thus, during the period of dehydration, after 37 days, the seeds were able to lose moisture. When seeds were allowed to lose moisture slowly after harvest, before being desiccated rapidly, their eventual condition as measured by leaching improved. Also, the faster the initial water loss preceding rapid desiccation the more readily were the dried seeds leached. It is suggested that seeds can only withstand rapid desiccation after the cessation of moisture increase and after some slow dehydration, which is accompanied by a slowing down in physiological activity.     

The capacity of plastids from developing pea cotyledons to synthesise acetyl CoA

In order to determine whether the enzymes required to convert triose phosphate to acetyl CoA were present in pea (Pisum sativum L.) seed plastids, a rapid, mechanical technique was used to isolate plastids from developing cotyledons. The plastids were intact and the extraplastidial contamination was low. The following glycolytic enzymes, though predominantly cytosolic, were found to be present in plastids: glyceraldehyde 3-phosphate dehydrogenase (EC 1.2.1.12), phosphoglycerate kinase (EC 2.7.2.3), and pyruvate kinase(EC 2.7.1.40). Evidence is presented which indicates that plastids also contained low activities of enolase (EC 4.2.1.11) and phosphoglycerate mutase (EC 2.7.5.3). Pyruvate dehydrogenase, although predominantly mitochondrial, was also present in plastids. The plastidial activities of the above enzymes were high enough to account for the rate of lipid synthesis observed in vivo.Abbreviations FPLC fast protein liquid chromatography - PPi pyrophosphate     

Nitrogen mineralization in relation to C:N ratio and decomposability of organic materials

A desk study was made on N mineralization of various organic materials. Data were obtained from the literature containing information on mineralized organic N, C/N of the substrate and some index for decomposability. Per class of substrate decomposability, linear regression lines between N mineralization and substrate C/N were established, from which apparent initial age and humification coefficient were calculated to indicate decomposability. For quantitative grading, a resistance index was formulated comprising the concentrations of lignin and polyphenols. The results confirmed that the fraction of mineralized organic N is linearly related to substrate C/N for equally decomposable materials. The nature of the organic materials and the processes to which they had been subjected were reflected in the ranking by decomposability.     

The apical nutations of pea seedlings in relationship to growth regulators

The apical hook curvatures of pea seedlings are associated with a higher level of endogenous auxin on the lower (ventral) side of the curvature in comparison with the upper (dorsal) side. For this reason this curvature may be reversed by applying IAA to the dorsal side of the hook. On the other hand the analyses did not reveal any differences in the content of endogenous gibberellin between the dorsal and ventral side of the hook. Endogenous gibberellin if applied either to the dorsal or to the ventral side of the hook curvature closes the hook. Exogenous cytokinin benzyladenine closes the curvature only if applied to the ventral side of the hook, but it opens it if applied to the dorsal one.     

Transcriptional profiling of the pea shoot apical meristem reveals processes underlying its function and maintenance

Background  

Despite the importance of the shoot apical meristem (SAM) in plant development and organ formation, our understanding of the molecular mechanisms controlling its function is limited. Genomic tools have the potential to unravel the molecular mysteries of the SAM, and legume systems are increasingly being used in plant-development studies owing to their unique characteristics such as nitrogen fixation, secondary metabolism, and pod development. Garden pea (Pisum sativum) is a well-established classic model species for genetics studies that has been used since the Mendel era. In addition, the availability of a plethora of developmental mutants makes pea an ideal crop legume for genomics studies. This study aims to utilise genomics tools in isolating genes that play potential roles in the regulation of SAM activity.     

Iron deficiency in rice shoots: identification of novel induced genes using RDA and possible relation to leaf senescence

Rice plants are highly susceptible to Fe-deficiency. Under nutrient deprivation, plant cells undergo extensive metabolic changes for their continued survival. To provide further insight into the pathways induced during Fe-deficiency, rice seedlings were grown for 3, 6 and 9 days in the presence or absence of Fe. Using RDA (Representational Difference Analysis), sequences of 32 induced genes in rice shoots under Fe-deficiency were identified. About 30% of the sequences found have been previously reported as responsive to other abiotic and even biotic stresses. However, this is the first report that indicates their relation to Fe deprivation. Differential expression of selected genes was confirmed by semi-quantitative RT-PCR analysis. The identification of classical senescence-related sequences, such as lipase EC 3.1.1.-, ubiquitin-conjugating enzyme EC 6.3.2.19, beta-Glucosidase EC 3.2.1.21 and cysteine synthase EC 2.5.1.47, besides the higher accumulation of total soluble sugars prior to the decrease of total chlorophyll content in Fe-deficient leaves, indicate that sugar accumulation may be one of the factors leading to premature leaf senescence induced by Fe-deficiency.     

Release of lateral buds from apical dominance by glyphosate in soybean and pea seedlings

Application of a sublethal dose of glyphosate (N-[phosphonomethyl]glycine) to the seedlings of soybean (Glycine max L. Merr. cv. Evans) and pea (Pisum sativum L. cv. Alaska) promoted growth of the cotyledonary and other lateral buds. The pattern of the glyphosate-induced lateral bud growth was different from that induced by decapitation. Under the experimental condition, glyphosate did not kill the apical buds. Feeding stem sections of the seedlings with radiolabeled indole-3-acetic acid ([2¹⁴C]IAA) and subsequent analysis of free [2-¹⁴C]IAA and metabolite fractions revealed that the glyphosate-treated plants had higher rates of IAA metabolism than the control plants. The treated pea plants metabolized 75% of [2-¹⁴C]IAA taken up in the 4-h incubation period compared to 46.5% for the control, an increase of 61%. The increase was small but consistent in soybean seedlings. As a result, the glyphosate-treated plants had less free IAA and ethylene than the control plants. The increase of IAA metabolism induced by glyphosate is likely to change the auxin-cytokinin balance and contribute to the release of lateral buds from apical dominance in these plants.     

Release of lateral buds from apical dominance by glyphosate in soybean and pea seedlings

Application of a sublethal dose of glyphosate (N-[phosphonomethyl]glycine) to the seedlings of soybean (Glycine max L. Merr. cv. Evans) and pea (Pisum sativum L. cv. Alaska) promoted growth of the cotyledonary and other lateral buds. The pattern of the glyphosate-induced lateral bud growth was different from that induced by decapitation. Under the experimental condition, glyphosate did not kill the apical buds. Feeding stem sections of the seedlings with radiolabeled indole-3-acetic acid ([2¹⁴C]IAA) and subsequent analysis of free [2-¹⁴C]IAA and metabolite fractions revealed that the glyphosate-treated plants had higher rates of IAA metabolism than the control plants. The treated pea plants metabolized 75% of [2-¹⁴C]IAA taken up in the 4-h incubation period compared to 46.5% for the control, an increase of 61%. The increase was small but consistent in soybean seedlings. As a result, the glyphosate-treated plants had less free IAA and ethylene than the control plants. The increase of IAA metabolism induced by glyphosate is likely to change the auxin-cytokinin balance and contribute to the release of lateral buds from apical dominance in these plants.     

A cytokinin complex from the developing fruits of Aegle marmelos

Following ion-exchange, Sephadex LH-20 and paper chromatography of the methanolic extracts of young, developing fruits of Aegle marmelos Correa, cytokinin-like activity in the soybean callus assay was detected in six fractions. Of the four butanol-soluble compounds, two were tentatively identified as zeatin and zeatin riboside, and the others as zeatin glucoside and zeatin riboside glucoside. The major cytokinino of the butanol-insoluble fraction is probably zeatin nucleotide. The levels of compounds with cytokinin-like activity were high during the early phase, and low in the subsequent period of fruit growth. The activity resembling that of cytokinin glucoside increased with maturation of the fruit. The content of free cytokinins in the fruits was more than that released from the tRNA.     

Isolation of prolegumin from developing pea seeds: its binding to endomembranes and assembly into prolegumin hexamers in the protein storage vacuole

A fraction enriched in endoplasmic reticulum and Golgi membranesfrom developing cotyledons of Pisum sativum L. has proved tobe a convenient source for the isolation of prolegumin, theprecursor of the major 11S storage globulin of pea seeds. Twopro-proteins were isolated with molecular masses of 60 kDa and75 kDa, respectively. A monoclonal antibody, designated 2B1,against prolegumin was raised using the in vitro immunizationtechnique. This antibody recognizes the 60 kDa precursor polypeptide,but only the 20 kDa ß-subunit of mature legumin. Prolegumin,like the ß-subunit of the mature legumin, is a hydrophobicprotein. After import into the protein storage vacuole, andafter formation of the protein bodies trimeric 9S proleguminassembles into 12S hexamers without prior processing of theprecursor. Since prolegumin in vitro does not oligomerize intomore than 9S tnmers these results suggest that a protein-mediatedassembly of 9S prolegumin trimers into 12S prolegumin hexamersprobably occurs in the lumen of the protein storage vacuole.Prolegumin, but not mature legumin, binds very tightly to membranes.This property points to a possible way of identifying a putativeprolegumin receptor. Key words: Calcium, Endoplasmic reticulum, Golgi apparatus, legumim, monoclonal antibody, pea cotyledons