Stress - induced Ethylene Production in the Ethylene - requiring Tomato Mutant

Ethylene synthesis in vegetative tisues is thought to be controlled by indoleacetic acid (IAA). However, ethylene synthesis in the diageotropka (dgt) mutant of tomato (Ly mkoaesc MIlL) was much less sensitive to IAA than In the norml variety (VFNS). Yet, mehanial wounding stimuted ethyne production by the mutant. The dgt tomato provides an opportunity to study the regulation of stress ethylene independent of IAA effects. Waterlogging (i.e anaerobic stress) stimulted production of the ethylene precursor, 1-aminocyclopropane-1-carboxylic acid (ACC), in the roots. The ACC was transported to the shoot where it was converted to ethylene. The dgt mutant efficiently utilzed ACC for ethylene synthesis under aerobic conditions. The results confirm that the genetic lesion in dgrt is located at a step prior to the formation of ACC. Furthermore, induction of ethylene synthesis by anaerobic or mechanical stresses in this mutant is independent of IAA actio The diageotropica (dgt) mutant of tomato is characterized by its diageotropic habit in both shoots and roots, dark green hyponastic leaves, and lack of lateral roots. These phenotypic characters are pleiotropic effects of a single gene mutation in the parent variety VFN8 (13). Zobel (14) reported that ethylene concentrations as low as 5 nl 1-1 would completely normalize the mutant. IAA would also restore the normal morphology, but dgt was 10 times less sensitive to the auxin than was VFN8. Zobel (14) suggested that the aberrant phenotype of dgt might be due to this reduction in auxin-induced ethylene synthesis. When wounded by excision, tissues of both dgt and VFN8 produced a burst of "stress ethylene." These observations suggest that IAA is not involved in the production of wound ethylene. Waterlogging is also known to increase ethylene synthesis in tomato (2, 6). It has recently been demonstrated that the stimulation of ethylene synthesis in waterlogged plants is due to export of an ethylene precursor, ACC2, from anaerobic roots (3). Anaerobic stress accelerates synthesis and accumulation ofACC in the roots, and ACC is transported in the xylem to the shoot where it is rapidly converted to ethylene. It has been proposed that IAA induces the synthesis of the enzyme which converts SAM to ACC in mung bean hypocotyls (11, 12). Little is known, however, about the regulation of stress ethylene synthesis (10). In this paper, ethylene production rates in response to waterlogging, wounding, ACC, and IAA treatments by dgt and VFN8 tomato plants are compared. ' This work was supported in part by National Science Foundation Grant PCM 78-98278. 2Abbreviations: ACC: I-aminocyclopropane-l-carboxylic acid; SAM: S-adenosylmethionine. MATERIALS AND METHODS Seeds ofboth the dgt and VFN8 isolines oftomato (Lycopersicon esculentum Mill.) were obtained from Dr. C. M. Rick of the University of California at Davis. Plants from the original seeds were grown in a greenhouse and the selfed progeny seeds were used in these experiments. The dgt plants displayed the characteristic phenotypic syndrome described by Zobel (15). The growth conditions, waterlogging treatments, ethylene measurements, xylem sap collection, and ACC feeding, identification, and assay techniques have been described in detail previously (3). Briefly, plants of both dgt and VFN8 were grown for 5 weeks in a growth chamber for waterlogging studies, or in a greenhouse for ACC or IAA feeding experiments. Ethylene production was estimated by sealing excised petioles in test tubes and periodically sampling the gas phase (5). Data are expressed on a fresh weight basis. Petiole angles were measured with a transparent protractor. Xylem sap from detopped root systems was collected for 3 h under a vacuum (50mm Hg). ACC was assayed by the method of Lizada and Yang (8). ACC or IAA was fed through the transpiration stream to shoot cuttings. VFN8 and dgt plants were generally compared in the same factorial experiment, allowing single degree offreedom tests in the analysis of variance. Since ACC levels in xylem sap of control plants were generally below detection, error bars indicating ± 1 SE have been used on ACC curves for flooded plants to give an estimate of experimental error. RESULTS The responses of dgt and VFN8 tomato plants to 48-h flooding were qualitatively identical (Table I). In both varieties, epinasty, ethylene production, and ACC export from the root increased dramatically due to flooding. The appearance of ACC in the xylem sap preceded the increase in ethylene synthesis (Fig. 1). Development of epinasty in flooded plants showed a time course similar to that for ethylene production (data not shown). When dgt plants were flooded for 30 h and then drained, ACC export from the root system fell from 0.75 nmol h-' to below detection (<0.02 nmol h-) within 6 h (Fig. 2). Petiolar ethylene production also fell rapidly and eventually returned to the control rate (Fig. 2). The time courses ofACC transport and ethylene production in VFN8 were virtually identical to those of dgt shown in Figures I and 2 (3). Evidence for a quantitative difference in ethylene production between the two isolines is shown in Figure 3. In this case, ethylene production by petioles from control and flooded plants was measured for 3.5 h following excision. Both VFN8 and dgt showed a typical increase in ethylene synthesis due to excision (5, 10). However, the magnitude of the increase is much greater in dgt than in VFN8. Flooding and wounding appear to have additive effects on ethylene production between 0.5 to 1.5 h following excision. Subsequently, the production rates for both flooded and 327 www.plantphysiol.org on July 15, 2017 Published by Downloaded from Copyright © 1980 American Society of Plant Biologists. All rights reserved. 328 BRADFORD AND YANG Table I. Effects of 48-h Flooding on Epinasty, C2,H4 Production, andACC Flux in VFN8 and dgt Tomato Plants Epinasty indicates the increase in petiole angle of the third leaf from zero time. Ethylene production refers to the rate by petioles during the first 30 min following excision. ACC refers to the concentration of the compound found in the xylem sap. ACC flux is calculated from the concentration in the sap times the exudation rate. Values are means of four plants per treatment. Variety Treatment nasty C2H4 ACC ACC Flux degrees nl g ' h' PM nmol h-' VFN8 Control 6 0.17 < 0.02 < 0.02 Flooded 45*** 0.89* 2.3*** 1.8*** dgt Control 6 0.27 <0.02 < 0.02 Flooded 32*** 1.19** 3.3*** 0.8*** * Difference from control significant at P < 0.05; ** difference from control significant at P < 0.01; *** difference from control significant at P<0.001.