Effect of the Defoliant Thidiazuron on Ethylene Evolution from Mung Bean Hypocotyl Segments

Abstract

The effect of the defoliant thidiazuron (N-phenyl-N'1,2,3-thiadiazo1-5- (2,18), it is important to understand the biochemical mechanism(s) by which TDZ enhances its production. Etiolated mung bean hypocotyls have been used extensively as a model system for evaluating the effects ofmany compounds that either enhance ' Mention of trademark or proprietary product does not constitute a guarantee or warranty of the product by the United States Department of Agriculture and does not imply its approval to the exclusion of other products that may also be suitable.2 Abbreviations: TDZ, thidiazuron; AVG, aminoethoxyvinylglycine; PCIB, 2-p-chlorophenoxy-2-methyl propionic acid; ACC, I-aminocyclopropane-l-carboxylic acid; AIBA, a-aminoisobutyric acid; AOA, aminooxyacetic acid.or reduce the rate of ethylene production. As a first step toward understanding/defining the physiological basis(es) of the defoliating activity of TDZ, its effect on ethylene evolution was examined. This paper describes the effects of TDZ on ethylene evolution from mung bean hypocotyls and attempts to define the biochemical basis for this interaction. A preliminary report of this research has been presented (22). MATERIALS AND METHODSPlant Material and Experimental Procedure. Mung bean (Vigna radiata L. Wilczek) seeds were surface-sterilized by soaking in 1% (v/v) NaOCI (1:5 dilution of commercial bleach) for 5 min. The seeds were then rinsed in running distilled H20 for 4 h and were sown in flats containing vermiculite. Etiolated seedlings were raised in an incubator (25 ± 1C) for 6 d. Hypocotyl segments (1.5 cm) were prepared by excising a portion of the hypocotyl immediately below the closed hook. All manipulations were conducted under a low fluence, green safelight in a darkroom.All experiments described in this paper were repeated at least three times. Whenever possible, each treatment within an experiment was replicated (n = 3). Due to the nature of some of the experiments, replication within an experiment was not feasible. Data from a typical experiment are presented.Chemicals. Technical-grade TDZ was a gift from E. Pieters of Nor-Am Agricultural Products, Inc. Stock solutions of thidiazuron were prepared in DMSO. A DMSO concentration of 0.1% (v/v) was used in all studies (including controls). AVG was a gift from Dr. R. W. Bagley of HLR Sciences, Inc. AOA, chloramphenicol, a-amino-isobutyric acid, BSA, DL-DTT, Hepes, pyridoxal-5-phosphate, sucrose, and PCIB were purchased from Sigma Chemical Co. ACC and Mes were purchased from Calbiochem. (NH4)2 SO4 was purchased from Schwartz-Mann. Dose-Response Studies. Excised hypocotyl segments were placed in 25 ml flasks that contained 4 ml of treatment solution (10 segments/flask). The treatment solution consisted of 10 mM Mes/KOH buffer (pH 5.7) containing 2% (w/v) sucrose, 5 mM CaCl2, 50 gg/ml chloramphenicol ± various concentrations of TDZ. The flasks were sealed and were incubated in the dark (25C) for 24 h. At that time the headspace was sampled and the ethylene content determined by GC usi...

Cytokinin-Induced Ethylene Biosynthesis in Nonsenescing Cotton Leaves

“…In etiolated mung bean hypocotyl segments, a marked synergism was found between kinetin and calcium ions (7). Other studies using this same tissue found only a marginal effect of calcium ions on TDZ-induced ethylene production (13). This type of marginal response to external calcium was also found in the present study (Table II).…”

“…studies found a 6 h lag period in kinetin-induced ethylene production in etiolated mung bean hypocotyl segments (7). Previous studies in this laboratory have found a readily measurable increase in ethylene production from etiolated mung bean hypocotyl segments within 2 h of TDZ treatment (13). In cotton leaves, no increase in ethylene evolution could be detected until 7 to 8 h following TDZ treatment (Table III).…”

“…As is the case with auxins (5), the increase in ethylene production following TDZ treatment of etiolated tissue segments was attributed to an increase in ACC synthase activity (13). No effect of TDZ treatment on EFE was observed.…”

“…In cotton, TDZ was far more effective than the auxin 2,4-D in stimulating ethylene production while in sunflower the reverse was true. Cytokinins have been shown to enhance ethylene production in root (12), epicotyl (3), hypocotyl (7,13) and leaf tissues (2,9,14). In these cases the cytokinins studied were much less effective than auxins and the maximum stimulation observed was generally small (less than 10 x control).…”

Morphoregulatory Role of Thidiazuron

“…Similarly, TDZ has been shown to promote shoot regeneration with an efficiency comparable to or greater than that of cytokinins (7,9). The effectiveness of TDZ to inhibit stomatal closure and to stimulate ethylene production in bean is also consistent with the properties of a cytokinin (6), although the pattern of the lag period in elevation of ethylene production by TDZ-treated mung bean tissue was similar to that induced by an auxin (19). However, in our study, both exogenous BAP and IAA were replaced by TDZ.…”