Absorption of Iron by Enzymically Isolated Leaf Cells

Iron uptake was studied using cells enzymically isolated from green tobacco leaves. Absorption was increased both by light and succinate as probable energy sources. Bicarbonate in the incubation mixture was inhibitory, and citrate also reduced absorption presumably by chelation with the metal. Absorption of iron was temperature sensitive and optimal at 25°C. Temperature coefficients and activation energies suggested that absorption was energy mediated. NaN₃ and DNP inhibited uptake at concentrations of 10-³M and 10^?4M, respectively. The inhibition caused by DNP was not negated by an external supply of ATP. The results suggest that iron absorption is an active metabolic process in cells enzymically isolated from green tobacco leaves. Cells from Fe-chlorotic leaves of PI 54619–5–1 soybean absorbed less iron than those derived from healthy leaves of the same variety, while leaf cells from the variety Hawkeye showed no such differences.     

Absorption,translocation, and metabolism of14C-clomazone in soybean (Glycine max) and threeAmaranthus weed species

The patterns of clomazone (2-[(2-chlorophenyl) methyl-4,4-dimethyl-3-isoxazolidinone) absorption, translocation, and metabolism and their contribution to the plant selectivity of this herbicide were studied in tolerant soybean [Glycine max (L.) Merr.] andAmaranthus hybridus and in susceptibleA. retroflexus andA. lividus. Differential root absorption appeared to play a significant role in the differential response of these four plant species to clomazone. Absorption of root-applied¹⁴C-clomazone was greater by the two sensitiveAmaranthus weeds than by the tolerant soybean andA. hybri-dus. Following application of¹⁴C-clomazone to roots, most of the absorbed radioactivity was translocated to the leaves of all four species. Approximately 50% of the absorbed¹⁴C-clomazone was metabolized by all four plant species as early as 12 h after treatment. Thin layer Chromatographic (TLC) analysis of plant tissue extracts from all four species revealed the formation of two major metabolites of clomazone. These unidentified metabolites had Rf values of 0.4 and 0.8, respectively, in a butanol∶acetic acid∶water (12∶3∶5, vol/vol/vol) developing system. The Rf value of unaltered clomazone in this system was 0.95. Differential metabolism or differential rate of metabolism of clomazone was not observed in this study and did not seem to account for the tolerance of soybean andA. hybridus or the suceptibility ofA. retroflexus andA. lividus to this herbicide.     

Sodium absorption by barley roots: role of the dual mechanisms of alkali cation transport

Radioactively labeled Na(+) absorbed by barley roots was sequestered in an intracellular compartment or compartments ("inner" spaces) in which it was only very slowly exchangeable with exogenous Na(+). Absorption of this fraction proceeded at a constant rate for at least 1 hour.When the rate of Na(+) absorption was examined over the range of concentrations, 0.005 to 50 mm, the isotherm depicting the relation showed dual kinetics as follows. Over the range, 0.005 to 0.2 mm, a single Michaelis-Menten term describes the relation between the concentration of Na(+) and the rate of its absorption. The mechanism of Na(+) absorption operating over this range of concentrations, mechanism 1 of alkali cation transport, is severely inhibited in the presence of Ca(2+) and virtually rendered inoperative for Na(+) transport by the combined presence of Ca(2+) and K(+). The mechanism is equally effective in Na(+) transport whether Cl(-) or F(-) is the anion, but is somewhat inhibited when the anion is SO(4) (2-).Over the high range of concentrations, 0.5 to 50 mm Na(+), a second, low-affinity mechanism of Na(+) absorption comes into play. In the presence of Ca(2+) and K(+), this mechanism 2 is the only one to transport Na(+) effectively, since Na(+) absorption via mechanism 1 is virtually abolished under these conditions.Anaerobic conditions, low temperature, and the uncoupler, 2,4-dinitrophenol, inhibit Na(+) absorption both at low and high Na(+) concentrations.     

Absorption,translocation, and metabolism of 14C-thifensulfuron in soybean (Glycine max), spurred anoda (Anoda cristata), and velvetleaf (Abutilon theophrasti)

The absorption, translocation, and metabolism of thifensulfuron-methyl {methyl 3-[[[[(4-methoxy)-6-methyl-1,3,5-triazin-2-yl]-amino]-carbonyl] amino]sulfonyl]-2-thiophenecarboxylate} were investigated in tolerant Essex soybean [Glycine max (L.) Merr.], moderately tolerant Vance soybean, and spurred anoda [Anoda cristata (L.) Schlecht.], and susceptible velvetleaf (Abutilon theophrasti Medic.). Radiolabeled (thiophene-2-¹⁴C) thifensulfuron-methyl was absorbed readily by young seedlings of all species following a foliar spray with the herbicide. Spot-applied ¹⁴C-thifensulfuron-methyl was absorbed by the treated leaf of all species. Absorption of thifensulfuron-methyl was limited when excised stems of all species were dipped into the herbicide solution for 2 h. Translocation of absorbed thifensulfuron-methyl to other plant parts was limited in all species, regardless of the method of its application. Root exudation of leaf-applied thifensulfuron-methyl was observed in all species and it was higher in seedlings of spurred anoda and velvetleaf. The two soybean cultivars metabolized 62–70% of absorbed thifensulfuron-methyl at 3 days after treatment with spot-applied ¹⁴C-thifensulfuron. Velvetleaf and tolerant spurred anoda metabolized about 50% of the absorbed herbicide. The major metabolite formed in all species appeared to be deesterified thifensulfuron acid. Differential metabolism seems to be a contributing factor in the selectivity of thifensulfuron-methyl between the two soybean cultivars and velvetleaf. The metabolic basis for the moderate tolerance of spurred anoda to thifensulfuron-methyl is not understood at the present time.Plant Pathology, Physiology, and Weed Science Department, Contribution no. 628.     

ABSORPTION OF HERBICIDES BY ROOTS

Crafts , A. S., and S. Yamaguchi . (U. California, Davis.) Absorption of herbicides by roots. Amer. Jour. Bot. 47(4): 248—255. Illus. 1960.—Many herbicides are used through soil. When 2,4-D*² was applied to culture-solution barley, bean, cotton, and Zebrina plants, there was evidence that the herbicide is held at high concentration by the roots. Very little of the labeled compound moved into the tops of barley, Zebrina and bean; a fair quantity was found in cotton foliage. Barley seedlings allowed to absorb 2,4-D*, ATA*, MH*, urea*, monuron*, dalapon*, simazin*, P³² and IAA* showed interesting differences. All chemicals were highly sorbed by roots, 2,4-D* was moved to tops in least amount, urea was next; the other seven were moved in larger quantities. ATA*, MH*, IAA*, P³², and dalapon* seemed readily phloem mobile; monuron* and simazin* seemed limited to xylem movement. Results are discussed in relation to the mechanism of root uptake.     

Accumulation of (2,4-dichlorophenoxy) acetic acid and (2,4,5-trichlorophenoxy) acetic acid by Parenchyma Tissue as Influenced by Metabolic Inhibitors and Lecithin

Parenchyma tissue from potato (Solanum tuberosum L. cv. Russet) tubers was treated with inhibitors to the release of metabolic energy in order to determine the importance of an active transport system for (2,4-dichlorophenoxy)acetic acid (2,4-D) and (2,4,5-trichlorophenoxy)acetic acid (2,4,5-T) accumulation. Results from treatments with carbonylcyanide m-chlorophenylhydrazone, an inhibitor of oxidative phosphorylation, and N,N'-dicyclohexylcarbodiimide, an inhibitor of membrane-bound adenosine triphosphatase, indicated 2,4-D and 2,4,5-T accumulation to be independent of available energy as influenced by these metabolic inhibitors. Lecithin treated parenchyma tissue accumulated more 2,4-D and 2,4,5-T than untreated tissue indicating possible binding of the herbicide to the lecithin moiety.     

Sodium absorption by barley roots: its mediation by mechanism 2 of alkali cation transport

Radioactively labeled Na⁺ absorbed by barley roots was sequestered in an intracellular compartment or compartments (“inner” spaces) in which it was only very slowly exchangeable with exogenous Na⁺. Absorption of this fraction proceeded at a constant rate for at least 1 hour.     

The absorption of potassium and several organic compounds by barley roots: Effect of siduron

Absorption of ⁴²K by excised roots of barley (Hordeum vulgareL.) grown in 0 or 5 ppm siduron (l-(2-methylcyclohexyl)-3-phenylurea)was a linear function of time for at least 60 minutes with transportbeing unidirectional. Absorption of siduron was a function ofthe external concentration to the limits of its solubility (0.09mM). However, the siduron- ¹⁴C absorbed by roots grown in either0 or 5 ppm siduron was in a readily exchangeable form and desorptionfor 4 hr exchanged 80 % of the label. Glucose-¹⁴C, adenine-8¹⁴Cand leucine¹⁴C were actively absorbed with 70 to 85 % of thelabel being absorbed in 24 hr. Although roots grown in iduronabsorbed less ⁴²K, glucose-¹⁴C, adenine-¹⁴C and siduron-¹⁴C,and more leucine-¹⁴C than similar roots grown in water culture,it is probable that these differences were not large enoughto account for the noted reduction (60%) in root growth. (Received January 9, 1968; )     

The absorption and translocation of diclofop-methyl and amitrole in wheat and oat roots

The absorption and translocation of diclofop-methyl (methyl 2-[4(2',4'-dichlorophenoxy)phenoxy]propanoate) was examined by using a specially designed treatment apparatus that separated excised roots or roots of seedlings into four zones. [¹⁴C]-Diclofop-methyl was absorbed along the entire root length of both wheat ( Triticum aestivum L.) and oat ( Avena sativa L.). In both species, absorption was greatest in the apical region of the root. Absorption by the apical region of wheat roots was more than three times greater than the basal portions, and more than twice as great as the apical region of oat roots. Less than 5% of the absorbed diclofop-methyl was translocated in both wheat and oat roots. Diclofop-methyl and diclofop(2-[4(2',4'-dichlorophenoxy)phenoxy]propanoic acid) were the predominant translocated forms. The absorption and translocation of amitrole (3-amino-1,2,4-triazole) were also examined. Amitrole was absorbed along the entire length of wheat roots and translocated primatily in the basipetal direction. The usefulness of the specially designed apparatus for biochemical and physiological studies is discussed.     

Effects and Absorption of Sethoxydim in Cell Cycle Progression of Corn (Zea mays) and Pea (Pisum sativum)

The mechanisms of selective herbicidal action of sethoxydim were investigated by using cultured root tips of corn (Zea mays L. cv Goldencrossbantam) and pea (Pisum sativum L. cv Alaska). Meristematic cells in the cultured roots were arrested in G₁ and G₂ of the cell division cycle by sucrose starvation and resumed growth and cell division (proliferation) when sucrose was provided. Corn root growth after sucrose addition was inhibited by sethoxydim at concentrations of 0.01 micromolar and greater when roots were treated in the presence of sucrose but was not inhibited at 10 micromolar sethoxydim when they were treated during sucrose starvation. Greater absorption of [¹⁴C]sethoxydim into the meristematic region of corn roots was observed when cells were in proliferative condition but not when they were arrested by sucrose starvation, whereas no greater absorption of the herbicide into pea meristems was observed in either growth condition. In the cell cycle study, greater absorption of [¹⁴C]sethoxydim into the corn root meristem was observed at a certain limited time before S (DNA synthesis) stage. The physiological effects and the greater absorption of sethoxydim clearly depended on cell cycle progression of corn root meristem, whereas fatty acid synthesis, as well as its inhibition by sethoxydim, was not associated with either cell cycle progression or greater absorption of the herbicide.     

Effects of pH Changes on Ion and 2,4-D Uptake of Wheat Roots

The quantitative relationships between pH-dependent ion and 2,4-D uptake in winter wheat seedlings (Triticum aestivum L. cv. Yubileynaya 50) have been investigated. The movement of various ions (potassium, phosphate, nitrate and ammonium) and 2,4-D across the root membranes was monitored with radioactive and stable isotope tracer methods. It was found that the H₊ ion concentration of the absorption solution strongly influences the 2,4-D uptake of the roots. Simultaneously, the 2,4-D uptake stimulates secretion of H⁺ into the absorption solution, that is, a H⁺ efflux can accompany the uptake of 2,4-D. This finding is consistent with the acid secretion theory of auxin and fusicoccin action. At pH 4 the 2,4-D uptake was much higher than at pH 6, thereby inhibiting the ion uptake and increasing the phytotoxicity in the plant. The results indicate that 2,4-D enters the root cells rapidly at the lower pH, mostly as undissociated molecules. With reference to the 2,4-D concentration in the roots at pH 4, a possible transport mechanism of the auxin herbicide is briefly discussed.     

A Role for Glutathione Transferase Omega 1 (GSTO1-1) in the Glutathionylation Cycle

The glutathionylation of intracellular protein thiols can protect against irreversible oxidation and can act as a redox switch regulating metabolic pathways. In this study we discovered that the Omega class glutathione transferase GSTO1-1 plays a significant role in the glutathionylation cycle. The catalytic activity of GSTO1-1 was determined in vitro by assaying the deglutathionylation of a synthetic peptide by tryptophan fluorescence quenching and in T47-D epithelial breast cancer cells by both immunoblotting and the direct determination of total glutathionylation. Mutating the active site cysteine residue (Cys-32) ablated the deglutathionylating activity of GSTO1-1. Furthermore, we demonstrate that the expression of GSTO1-1 in T47-D cells that are devoid of endogenous GSTO1-1 resulted in a 50% reduction in total glutathionylation levels. Mass spectrometry and immunoprecipitation identified β-actin as a protein that is specifically deglutathionylated by GSTO1-1 in T47-D cells. In contrast to the deglutathionylation activity, we also found that GSTO1-1 is associated with the rapid glutathionylation of cellular proteins when the cells are exposed to S-nitrosoglutathione. The common A140D genetic polymorphism in GSTO1 was found to have significant effects on the kinetics of both the deglutathionylation and glutathionylation reactions. Genetic variation in GSTO1-1 has been associated with a range of diseases, and the discovery that a frequent GSTO1-1 polymorphism affects glutathionylation cycle reactions reveals a common mechanism where it can act on multiple proteins and pathways.     

Uptake and accumulation of the herbicide bentazon by cultured plant cells

Cellular absorption of the herbicide bentazon, a weak acid with pK_a 3.45, was investigated using suspension-cultured cells of velvetleaf (Abutilon theophrasti Medic.). Bentazon accumulated rapidly to concentrations approximately four times that of the external medium. Bentazon accumulation against a concentration gradient was not due to its conversion to metabolites, partitioning into lipids, or binding onto cellular constituents. Bentazon uptake was related linearly to the external bentazon concentration, implying that movement of the herbicide into cells was not carrier-mediated. Bentazon was able to diffuse freely and extensively out of the cells, indicating that bentazon can readily diffuse across cell membranes. Potassium cyanide and carbonyl cyanide m-chlorophenyl hydrazone inhibited bentazon accumulation as did nitrogen gas when bubbled through the uptake medium. Absorption was pH-dependent with the greatest amount of bentazon accumulating at acidic external pH. Calculations indicated that conversion of uncharged bentazon to bentazon anion in the cytoplasm accounts for cellular accumulation of bentazon. These results provide evidence that bentazon is absorbed across membranes via simple diffusion and that bentazon accumulates in plant cells via an energy-dependent, ion-trapping mechanism which results in bentazon accumulation in the cytoplasm.     

Delftia acidovorans MC1 resists high herbicide concentrations--a study of nutristat growth on (RS)-2-(2,4-Dichlorophenoxy)propionate and 2,4-dichlorophenoxyacetate

Delftia acidovorans MC1 was continuously cultivated under nutristat conditions with elevated concentrations of the herbicides (RS)-2-(2,4-dichlorophenoxy)propionate [(RS)-2,4-DP] and 2,4-dichlorophenoxyacetate (2,4-D). The presence of 1-5 mM of either of these compounds did not essentially inhibit growth. Moreover, substrate consumption was not essentially affected at pH values of 7.0-9.0 selected by reason of alkaline in situ conditions found e.g. on contaminated building rubble but was decreased at pH 9.3. The adenylate energy charge declined to some degree as the herbicide concentration rose, the extent of this increasing as the pH rose. This was caused by an increase in the concentration of ADP and in particular AMP, in contrast to the fairly constant ATP level of around 4 nmol/mg dry mass with (RS)-2,4-DP and 2 nmol/mg with 2,4-D. Comparison of the individual growth parameters with theoretical data taking into account maintenance coefficients of 0.48 mmol (RS)-2,4-DP/g*h and 0.6 mmol 2,4-D/g*h revealed that the culture followed purely kinetic rules. This excludes the necessity of using substrate to a significant extent to satisfy extra efforts in energy for homeostasic work under these accentuated conditions.     

Amitrole Absorption by Bean (Phaseolus vulgaris L. cv ;Red Kidney') Roots : Mechanism of Absorption

The mechanism of transport of the herbicide 3-amino-1,2,4-triazole (amitrole) into Phaseolus vulgaris roots appears to be passive, as judged by the effect of temperature (Q₁₀ = 1.3 between 15 and 25°C) and the lack of sensitivity to metabolic inhibition afforded by 2,4-dinitrophenol and NaN₃. Amitrole absorption is a linear function of external concentration over several orders of magnitude and, thus, is not facilitated by a carrier mechanism. The absorption of amitrole is sensitive to external pH, being stimulated under acid conditions. This stimulation of amitrole absorption is seen at low (≤1 millimolar) amitrole concentrations, but not at high (50 millimolar) amitrole levels. While the apparent octanol-water partition coefficient varies with the pH of the aqueous phase, there is no clear correspondence between absorption and the apparent partition coefficient. Roots do not accumulate amitrole above concentration equilibrium; however, at a time when the net amitrole content of the root tissue begins to saturate, amitrole can be detected in the xylem stream. On a fresh-weight basis, amitrole absorption by roots is equal to that accomplished by trifoliate-leaf tissue. An estimate of the permeability coefficient (according to the analysis of Tyree et al. 1979 Plant Physiol 63: 367-374) suggests that amitrole possesses near-optimal permeability for an ambimobile solute, on the order of 2.12 (± 0.47) × 10⁻⁹ meters per second.     

Differential Absorption, Translocation and Metabolism of Metribuzin [4-amino-6-tert-butyl-3-(methylthio)-as-triazine-5(4H)one] by Soybean Cultivars

Soybean (Glycine max (L.) Merr.) cultivars have been reported to range in tolerance to injury by 4-amino-6-tert-butyl-3-(methylthio)-as-triazine-5(4H)one (metribuzin), from tolerant (e.g.‘Bragg’) to susceptible (e.g.‘Coker 102’ and ‘Semmes’). ‘Bragg,’‘Coker 102’, and ‘Semmes’ soybeans were grown in sand subirrigated with nutrient solution containing labelled (¹⁴C-carbonyl metribuzin) and nonlabelled metribuzin to determine cultivar variability in absorption, translocation, and metabolism of metribuzin. Plants were periodically harvested, autoradiographed, and radioactivity in tissue extracts quantified. Data indicated that all 3 cultivars readily absorbed and translocated metribuzin. However, ‘Bragg’ tissues accumulated greater quantities of metribuzin metabolites than the other two cultivars. The major ¹⁴C-containing metabolite in ‘Semmes’ and ‘Coker’ roots and stems was 6-tert-butyl-as-triazine-3-5-(2H,4H)-dione, whereas the major ¹⁴C-metabolite isolated from‘Bragg’ roots and stems was a glucose conjugate. Results indicated that differential-intraspecific responses to metribuzin may result from differential capacities for herbicide detoxification by conjugation.     

Cadmium uptake kinetics in intact soybean plants

The absorption characteristics of Cd²⁺ by 10- to 12-day-old soybean plants (Glycine max cv Williams) were investigated with respect to influence of Cd concentration on adsorption to root surfaces, root absorption, transport kinetics and interaction with the nutrient cations Cu²⁺, Fe²⁺, Mn²⁺, and Zn²⁺. The fraction of nonexchangeable Cd bound to roots remained relatively constant at 20 to 25% of the absorbed fraction at solution concentration of 0.0025 to 0.5 micromolar, and increased to 45% at solution concentration in excess of 0.5 micromolar. The exchangeable fraction represented 1.4 to 32% of the absorbed fraction, and was concentration dependent. Using dinitrophenol as a metabolic inhibitor, the `metabolically absorbed' fraction was shown to represent 75 to 80% of the absorbed fraction at concentration less than 0.5 micromolar, and decreased to 55% at 5 micromolar. At comparatively low Cd concentrations, 0.0025 to micromolar 0.3, root absorption exhibited two isotherms with K₂ values of 0.08 and 1.2 micromolar. Root absorption and transfer from root to shoot of Cd²⁺ was inhibited by Cu²⁺, Fe²⁺, Mn²⁺, and Zn²⁺. Analyses of kinetic interaction of these nutrient cations with Cd²⁺ indicated that Cu²⁺, Fe²⁺, Zn²⁺, and possibly Mn²⁺ inhibited Cd absorption competitively suggesting an involvement of a common transport site or process.     

Mechanism of cholesterol absorption by the hydatid cysts of Echinococcus granulosus (Cestoda)

• 1.1. The mechanism of cholesterol absorption by hydatid cysts of Echinococcus granulosus was studied in function of time of incubation, size of cysts, temperature of incubation, concentration of cholesterol substrate and presence of metabolic inhibitor.
• 2.2. The results indicated that the uptake of cholesterol continuously increased with time of incubation, that small cysts absorbed cholesterol better than large ones, that the optimum temperature for uptake was 25°C and that uptake increased with increase in concentration of substrate.
• 3.3. The addition of the metabolic inhibitor 2, 4-dinitrophenol did not affect the process of absorption thus eliminating the participation of the mechanisms of active transport or facilitated diffusion.
• 4.4. It was suggested that mechanisms of simple diffusion and possibly exchange diffusion are operating in cholesterol absorption by hydatid cysts.

     

The effect of 3,5-diiodo-4-hydroxyphenylmethylsulphone (ioxysone) on Phaseolus vulgaris: absorption and translocation studies

The herbicide 3,5-diiodo-4-hydroxyphenylmethylsulphone (ioxysone) is shown to be absorbed by the roots of dwarf bean ( Phaseolus vulgaris ) and readily translocated. No evidence was obtained that iodide becomes released from the herbicide within the plant. This behaviour is discussed and contrasted with that of the corresponding nitrile (ioxynil), a widely used contact herbicide which is not readily translocated in the plant and whose mode of action is different from that of ioxysone.     

Rapid phytochrome-mediated changes in the uptake by bean roots of sodium acetate [1-14C] and their modification by cholinergic drugs

Summary 4 min of red light increases the uptake of sodium acetate[1-¹⁴C] by excised, etiolated secondary roots of Phaseolus aureus Roxb. 4 min of far-red light reveres this effect. AMO-1618, which inhibits acetylcholinesterase activity, enhances the red-light effect, while d-tubocurarine, which blocks the animal acetylcholine receptor, inhibits it. Red light also increases basipetal translocation of the label. When the metabolic fate of the label was determined in dark-held roots, 36% of the label remained as acetate, 48% evolved as [¹⁴C]CO₂, 3% partitioned with acetylcholine, and 3% effluxed from the roots. The rest of the label was associated with the coarse residue left after extraction. The major effect of red light was to increase the uptake of the label in the acetate fraction.We interpret these observations to mean that the phytochrome mechanism immediately causes an increase in uptake of the label during brief irradiation with red light. Because of our previous demonstration that both red light and acetylcholine increase respiration, it is probable that the increased absorption of the label is a process requiring respiratory energy. These data support the concept of phytochrome as a membrane-bound functional system that in bean roots is mediated by the acetylcholine mechanism.Abbreviations ACh Acetylcholine - AChE acetylcholinesterase - ATP adenosine triphosphate - AMO-1618 2-isopropyl-4-dimethylamino-5-methylphenyl-1-piperidine carboxylate methyl chloride - TPB tetraphenyl boron - D darkness - FR far-red - R red