Jaleh Daie

Sucrose uptake in isolated phloem of celery is a single saturable transport system

The uptake of different sugars was studied in segments of isolated phloem from petioles of celery (Apium graveolens L.) in order to determine the kinetics and specificity of phloem loading in this highly uniform conductive tissue. The uptake kinetics of sucrose and the sugar alcohol, mannitol, which are both phloem-translocated, indicated presence of a single saturable system, while uptake of non-phloem sugars (glucose and 3-O-methylglucose) exhibited biphasic kinetics with lower uptake rates than those for sucrose and mannitol. The presence of unlabeled mannitol, 3-O-methylglucose and maltose in the incubation solution did not cause inhibition of labeled-sucrose uptake, indicating high carrier specificity and lack of sucrose hydrolysis in vivo. The pH optimum for sucrose uptake was 5–6. Furthermore, a rapid and transient alkalinization of the external media by sucrose indicated a sugar/H+-cotransport mechanism. Dual-labeling experiments showed that sucrose influx continued at a constant rate (Vmax=15 μmol·h-1·(g FW)-1), whereas sucrose efflux was low and insensitive to external concentration. Therefore, the saturable uptake kinetics for sucrose did not appear to be the result of an equilibrium between rates of sucrose influx and efflux.

End-Product Control of Carbon Metabolism in Culture-Grown Sugar Beet Plants (Molecular and Physiological Evidence on Accelerated Leaf Development and Enhanced Gene Expression)

Sugar beet (Beta vulgaris L.) seedlings were grown on media containing 90 to 300 mM sucrose or glucose. Compared to controls, sugar-grown plants had higher growth rate, photosynthesis, and leaf sugar levels. The steady-state level of transcripts increased significantly for the small subunit of ribulose-1,5-bisphosphate carboxylase/oxygenase (Rubisco) (rbcS) and the cytosolic fructose-1,6-bisphosphatase and moderately for the Rubisco large subunit (rbcL). The transcript level of sucrose phosphate synthase remained unchanged. Fructose-1,6-bisphosphatase and Rubisco activities did not change in the presence of sugars, but that of sucrose phosphate synthase increased (44 and 90% under selective and nonselective assay conditions, respectively). Accelerated leaf development was indicated by (a) autoradiograms of leaves that showed that sucrose loading occurred earlier, (b) export capacity that also occurred earlier but, after about 2 weeks, differences were not detectable, and (c) sucrose synthase activity that declined significantly. Several conclusions emerged: (a) response was nonosmotic and gene and sugar specific, (b) sugars caused accelerated leaf development and sink-to-source transition, (c) enhanced gene expression was due to advanced leaf development, and (d) whereas Rubisco and cytosolic fructose-1,6-bisphosphatase genes were sugar repressed in mature leaves of greenhouse-grown plants, they were unaffected in mature, culture-grown leaves. To our knowledge, these data provide the first evidence in higher plants that, depending on the physiological/developmental context of leaves, sugars lead to differential regulation of the same gene.

Purification and light-dependent molecular modulation of the cytosolic fructose-1,6-bisphosphatase in sugarbeet leaves

Cytosolic fructose-1,6-bisphosphatase (FBPase) was purified 472-fold from sugarbeet (Beta vulgaris L.) leaves by ammonium sulfate fractionation, anion-exchange chromatography (DEAE Sepharose), cation-exchange chromatography (S-Sepharose), gel filtration (Sephacryl S-300), and hydrophobic interaction chromatography (Phenyl Sepharose). The dissociated polypeptide (molecular mass of 37 kD) was used to generate polyclonal antibodies. Western blot analysis revealed a single band that was identified as the cytosolic FBPase. Enzyme activity and protein and transcript levels were measured under various light and dark conditions in growth chamber-grown plants. FBPase protein level remained unchanged during a diurnal cycle, but enzyme activity and transcript levels were highest and lowest at the end of the light and dark periods, respectively. Light-dependent increase in the enzyme activity and transcript level was gradual, occurring several hours after the onset of light. At the end of an extended dark period (48 h), FBPase activity was negligible, protein level was unchanged, and transcript level had declined (but considerable amounts of transcript remained). Neither activity nor protein and transcript were detected in etiolated leaves. Nearly 24 h of continuous exposure to light was required before the FBPase protein and activity reached maximal levels. Unlike the chloroplastic FBPase, which is light activated (direct regulation), changes in the cytosolic FBPase activity and transcription appear to be light dependent in an indirect manner. The data provide first evidence on the coarse control of this enzyme via a light-dependent modulation of transcription and posttranslational modification.

Phloem loading of sucrose: Update and opportunities in molecular biology

Sucrose accumulates in the phloem against a concentration gradient via a presumed sucrose-specific carrier protein located at the plasmalemma of the sieve elements/companion cells. Recent evidence suggests that sucrose carrier in soybean is a 62-kDa protein. Immunocytochemical localization has shown the protein to be exclusively at the plasmalemma, which is also the site of sucrose transport. To enhance our understanding of the phenomenon, the structural gene of the sucrose carrier must be cloned and sequenced. Furthermore, development of appropriate probes should help answer long-standing questions relative to the molecular nature of sugar transport and phloem loading, the mechanism of induction/activation of sugar carriers, and developmental regulation of expression of genes encoding such carriers.

Differential rates of sucrose and hexose transport by asparagus cell cultures

Abstract Cell cultures of asparagus ( Asparagus officinalis L.) grew well on media containing 3% (w/v) sucrose, glucose or fructose. However, optimal growth occurred with sucrose. The objective of this work was to characterize sugar transport and to determine the presence of sugar carriers in these cells. Cell cultures preferentially transported fructose, glucose, 3- O -methyl glucose (3-OMG), sucrose and l -glucose, in that order. Both sucrose and 3-OMG uptake was inhibited by carbonylcyanide chlorophenyl hydrazone (CCCP) and p -chloromercuribenzene sulfonic acid (PCMBS). Due to high cell wall invertase activity, sucrose was efficiently hydrolyzed so that no sucrose was detected in the culture medium after 5 days and cells accumulated more hexose than sucrose. Furthermore [ 14 C]sucrose uptake was partially inhibited in the presence of unlabeled 3-OMG. Cell cultures transported 1′-fluorosucrose, a sucrose analog not subject to invertase hydrolysis, at rates similar to that of sucrose. The evidence indicated that despite the presence of high cell wall invertase activity and efficient hexose transport, a sucrose carrier is present at the plasmalemma of asparagus cell cultures. Therefore, sucrose hydrolysis by invertase is not essential for transport into these cells.

Hormone-mediated enzyme activity in source leaves

The initial step in carbon allocation occurs in leaves and is the chemical partitioning of carbon between sucrose and starch. Sucorse phosphate synthase is one of the enzymes belived to regulate rate of sucrose synthesis. In this study, the effects of indoleacetic acid, gibberellic acid, and abscisic acid on the activity of this enzyme were investigated in source leaves of mature sugar beets. Preliminary evidence is presented that, concurrent with a modification of sucrose uptake rates, i.e., phloem loading, these plant growth substances modify the activity of sucrose phosphate synthase resulting in altered partitioning of carbon between sucrose and starch.

Water relations, photosynthesis and assimilate partitioning in leaves of pepper (Capsicum annuum) transplants: Effect of water stress after transplanting

SummaryWhen pepper (Capsicum annuum L., cv. Maor) transplants, grown in Speedling type trays, were transplanted into wet soil but not watered for up to 48 h (day/night temperature 30/25°C), their leaf water potential was reduced from —0.5 to nearly —2.0 MPa. This decrease was mainly due to loss of leaf tugor. At such low water potential, photosynthesis of the source leaves was completely inhibited. The leaf water potential returned to the control values 24 h after rewatering but the photosynthetic rate of the stressed plants did not fully recover and was dependent on the extent to which the water potential had decreased during the stress. Translocation of l4C-assimilates, following l4C02 feeding, was inhibited in stressed transplants with the upward flow towards the young leaves being more affected that the downward flow towards the roots. After rewatering, transport of 14C-assimilates from the source leaf of the stressed transplants to the root was recovered but transport to the young developing leaves r...

Effect of heat stress on the growth, root sugars, acid invertase and protein profile of pepper seedlings following transplanting

The development of pepper transplants is sensitive to the prevailing temperature in the field. High temperatures immediately after transplanting pepper seedlings delay their development in the field. During nine days of post-transplanting heat stress (30/25°C or 35/25°C, day/night temperature regimes), root and shoot growth were inhibited compared to the control transplants (25/18°C) with a significant decline in total protein and soluble acid invertase activity in the roots of the heat stressed plants. Acid invertase activity was less sensitive to the heat stress in the leaves than in the roots. A significant reduction in the root invertase activity occurred as early as two days after the beginning of the stress. Transplanting caused a 72% inhibition in the carbon allocation to the roots, compared with the non-transplanted seedlings. Exposure to heat after transplanting did not further reduce carbon allocation to the roots. The protein profile on SDS-PAGE from leaves shows that during the heat stress, so...

Bioregulator enhancement of sink activity in sugar beet

The sink mobilizing abillity is partially determined by sugar uptake rates of storage cells. Two synthetic growth regulators (Pix and BAS 106W) were tested for their effect on sucrose uptake in root tissue discs or glucose uptake in cell cultures of sugar beet. In tissue discs, uptake at the plasmalemma was determined by incubating the discs for 1 h in the presence of 5 mM sucrose and at the tonoplast for 4 h in the presence of 40 mM sucrose. Cell cultures were incubated for 1 h in the presence of 1 mM glucose. Pix (10 mg l−1) caused a 20% stimulation of active sucrose uptake at the plasmalemma. Active sucrose uptake at the tonoplast was increased 67% by 100 mg l−1 Pix. No effect of BAS 106W was observed on sucrose uptake in tissue discs. In cell cultures, a 65% enhancement of active glucose uptake was observed with both Pix and BAs 106W. When the bioregulators were applied to the root medium of seedlings, Pix but not BAS 106W resulted in increased root/shoot ratio, translocation of 14C-assimilates, and allocation of more biomass to the root sink. The data suggested that sugar transport and translocation may be used as biochemical criteria for rapid screening of effective yield enhancing bioregulators.