Transport of the Herbicide 3-Amino-1,2,4-Triazole by Cultured Tobacco Cells and Leaf Protoplasts

The chanism of transport of the herbicide 3-amino-l,2,4-triazole (amitrole) into Phaswolw vulgaris roots appears to be passive, as judged by the effect of temperature (Qio = 13 between 15 and 25°C) and the lack of sensitivity to metabolic inhibition afforded by 2,4dinitrophenol 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 The mechanism(s) by which polar solutes negotiate the barrier of cell wall and plasmalemma are topics of continued interest to plant physiologists. In particular, knowledge of the means by which xenobiotics enter the symplast is of both theoretical and practical concern. The water-soluble herbicide, 3-amino-1,2,4-triazole (amitrole), is a compound which shows extensive xylem and phloem mobility in a variety of plant species and is relatively nonselective in its phytotoxicity. Amitrole has been proposed to exert its herbicidal activity by several modes of action, including interference in Chi biosynthesis and disruption ofhistidine metabolism (see 1 and 2). Regardless of the actual mechanism of action, amitrole must clearly penetrate one or more membrane systems in order to exert its effect. An understanding of the factors which contribute to its penetrability may therefore allow one to increase efficacy while limiting the amount of material released into the environment. This study was conducted with the aim of elucidating the mechanism by which amitrole is absorbed. Root Absorption Studies. Root uptake studies were performed in Plexiglas chambers of two or more compartments placed in a thermostated bath. In most experiments, roots were sealed through holes in compartment dividers with silicone vacuum grease (Dow Corning Corporation). Thus, the distal 0.5 cm (compartment A), the more proximal 2.5 cm (compartment B), and the remaining portion of the primary root up to the region of emergence from the seed (compartment C) were effectively isolated from each other. Treatment with radioactive herbicide could be accomplished in any of these three compartments, and uptake by and transfer from the treated segments determined by excising the treated and untreated portions and sampling the solution contained in the other compartments. In most cases, ["C]amitrole was added to compartment B; over a 30-min uptake period, no counts above background were found in either the tissue or solution in compartments A or C. Mixing and aeration in the compartment containing the label was accomplished by a stream of humidified air delivered by a polyethylene tube. For experiments involving uptake by roots and transfer of the label to the xylem (Fig. 4), the proximal 0.2 cm of root protruded into compartment A; the divider between sections B and C was removed.Radioactivity was assayed by liquid scintillation counting in a mixture of toluene: Triton X-100 (2:1, v/v) containing 4 g/L Omnifluor (New England Nuclear) on a Beckman LS 8000 LScounter at efficiencies ranging from 85 to 96%. Resul...

Section: Discussionsupporting

confidence: 57%

Amitrole Absorption by Bean (Phaseolus vulgaris L. cv `Red Kidney') Roots

Lichtner

1983

“…The by Pi. On the other hand, nonsaturable absorption of the d to new bentazon-free herbicides amitrole and atrazine indicated these herbicides e Fisher's LSD value for moved across cell membranes by diffusion (14,18). The significance.…”

Section: Metabolic Inhibitorsmentioning

confidence: 99%

Uptake and Accumulation of the Herbicide Bentazon by Cultured Plant Cells

Sterling¹,

Balke²,

Silverman³

1990

Cellular absorption of the herbicide bentazon, a weak acid with pK. 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 extemal bentazon concentration, implying that movement of the herbicide into cells was not carriermediated. 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 extemal 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.A herbicide must be absorbed by plant cells and accumulate at the site of action before it causes phytotoxicity. In general, nonfacilitated diffusion is the primary mechanism by which herbicides move across plant cell membranes (10), although active or carrier-mediated transport of a few herbicides into plant cells has been documented (5, 17). Natural, plant growth regulating chemicals (1) and secondary plant products (13) are transported across membranes by a combination of passive and active processes. Knowledge of the mechanisms involved in the assymetric distribution of herbicides across membranes may explain relationships between herbicide concentration and phytotoxicity.

“…From these studies, most herbicides have been shown to be transported passively across the lipid bilayer into protoplasts, with diffusion as the likely driving force. Amitrole uptake into tobacco protoplasts was reported to be via an energy-independent, noncamer-mediated process (Singer and McDaniel, 1982). Studies of protoplasts from maize roots showed that triazines moved rapidly into protoplasts with maximal accumulation occurring after 10 s of incubation (Darmstadt et al, 1983(Darmstadt et al, , 1984.…”

mentioning

confidence: 99%

Characterization of Paraquat Transport in Protoplasts from Maize (Zea mays L.) Suspension Cells

Hart¹,

DiTomaso²,

Kochian³

1993

Protoplasts isolated from maize (Zea mays L.) suspension cells were used to study transport of paraquat.[14C]Paraquat uptake was measured in 400-pL centrifuge tubes using silicon oil centrifugation techniques. Approximately 50% of accumulation from a 100 p~ paraquat solution occurred in the first 10 s, and net accumulation reached a maximum after about 10 min. Membrane binding accounted for about 30% of apparent accumulation. Concentration-dependent uptake kinetics were characterized by a nonsaturating curve, which was resolved into a linear and a saturable component. The K,,, of the saturable component was 132 p~, and the V,, was 0.512 nmol pL of protoplasts-' min-'. In the absence of sucrose, the Vmax of the saturable component was reduced by 52%, suggesting that paraquat uptake across the plasmalemma is energy dependent. Measurement of concentration-dependent binding of paraquat to burst protoplasts showed a linear response. This suggests that the linear component from intact protoplast concentration kinetics represented paraquat binding to the plasmalemma surface. Calcium inhibited the saturable component, and this inhibition was shown by Lineweaver-Burk analysis to be noncompetitive. Putrescine, a divalent cationic polyamine with a charge distribution similar to that of paraquat, competitively inhibited paraquat uptake. These results show that paraquat transport characteristics at the plasmalemma of maize protoplasts are similar to those reported earlier for paraquat transport in roots of intact maize seedlings.