Planta
Planta (1984)162:342 344
9 Springer-Verlag 1984
Growth, graviresponsiveness and abscisic-acid content of Zea mays seedlings treated with Fluridone Randy Moore and James D. Smith Department of Biology, Baylor University, Waco, TX 76798, and Department of Soil and Crop Science, Texas A and M University, College Station, TX 77843, USA
Abstract. Ten-d-old seedlings o f Z e a mays L. cv. Tx 5855 treated with 1-methyl-3-phenyl-5-(3-[trifluoromethyl]phenyl)-4-(1H)-pyridinone (Fluridone) were analyzed for abscisic acid (ABA) content using high-performance liquid chromatography with an analysis sensitivity of 2.5 ng ABA g-l fresh weight (FW). Seedlings were divided into three portions: leaves, detipped roots, and root tips (terminal 1.5 ram). Control plants (water treatment only; no Fluridone) were characterized by the following amounts of ABA: leaves, 0.1144-0.024 (standard deviation) gg ABA g-1 FW; detipped roots, 0.2604-0.039+ gg ABA g-1 FW; root tips, no ABA detected. We did not detect any ABA in tissues of Fluridone-treated plants. Primary roots of treated and untreated seedlings were strongly graviresponsive, with no significant differences between the curvatures or the growth rates of primary roots of Fluridone-treated and control seedlings. These results indicate that 1) Fluridone completely inhibits ABA synthesis, and 2) ABA is not necessary for positive gravitropism by primary roots of Zea mays.
Key words: Abscisic acid and root growth - Fluffdone - Gravitropism (root) - Root growth - Zea (root growth).
Introduction
There is a large amount of evidence indicating that the root cap is the source of a growth-inhibiting influence on the root, and that an imbalance of inhibitor(s) originating in the cap is the basis for root gravicurvature (see reviews by Jackson and Barlow 1981; Moore 1984; Wilkins 1984). Transport of the inhibitor(s) from the cap to the elongating zone (at greater concentrations in the lower Abbreviations: ABA = abscisic acid; Fluridone = 1-methyl-3phenyl-5-(3-[trifluoromethyl]phenyl)-4-(1 H)-pyridinone; FW = fresh weight; SD = standard deviation
half than in the upper half) of the root presumably results in more rapid growth of the upper half of the root, thereby accounting for positive gravicurvature (see reviews by Audus 1983; Wilkins 1984). One inhibitor that many investigators believe to be involved in root gravitropism is abscisic acid (ABA) (Audus 1983; Wilkins 1984). Evidence supporting this conclusion includes the following observations: 1) Abscisic acid is present in caps and tips of roots. 2) Abscisic acid inhibits root elongation. 3) Asymmetrical application of ABA induces root curvature. 4) Light-mediated inhibition of root elongation is apparently a consequence of ABA synthesis in root caps. 5) Gravicurvature by roots of several cultivars of Z. mays that require light for graviresponsiveness correlates positively with increased levels of ABA in their root caps. 6) The onset of graviresponsiveness correlates positively with an increase in ABA content in the lower half and a concomitant decrease in ABA in the upper half of horizontally-oriented roots (see reviews by Jackson and Barlow 1981; Audus 1983; Wilkins 1984). Recently, however, several investigators have questioned the involvement of ABA in root graviresponsiveness, based on the following observations: 1) There is little evidence that intact roots can transport ABA from the root cap to the zone of elongation. 2) If the cap inhibitor is ABA, then placing root caps on growing coleoptiles would be expected to inhibit elongation. However, an auxinlike stimulation (instead of an ABA-like inhibition) of growth occurs in response to this treatment. 3) There is no significant lateral asymmetry of ABA in gravistimulated roots (Mertens and Weiler 1983). 4) Abscisic acid has been reported to have no effect on root elongation or to promote, at least initially, root elongation (Evans and Mulkey 1982; Mulkey et al. 1983; also see reviews by Jackson and Barlow 1981; Audus 1983). Several investigators have suggested that ABA is synthesized via the carotenoid pathway (see re-
R. Moore and J. D. Smith: ABA content and root graviresponsiveness of Fluridone-treated seedIings v i e w b y M i t b o r r o w 1983). I f t h i s is t r u e , t h e n i n h i biting carotenogenesis should also prevent the synthesis o f A B A . I n this s t u d y w e t r e a t e d s e e d a n d seedlings ofZ. mays with Fluridone, an inhibitor of c a r o t e n o i d s y n t h e s i s ( F o n g et al. 1 9 8 3 a , b). W e s u b s e q u e n t l y m e a s u r e d the g r o w t h a n d gray• s p o n s i v e n e s s o f t h e t r e a t e d r o o t s in o r d e r to d e t e r mine if ABA was necessary for gray• ness.
M a t e r i a l and methods Plant material and growth conditions. Caryopses of inbred lines of Zea mays L. cv. Tx 5855 were obtained from plants grown at the Texas A and M University Farm. They were soaked in Fluridone (100 nag 1-1) for 10 h and then planted in vermiculite in a controlled-environment growth chamber. Seedlings were watered regularly with Fluridone (100 mg l-l). Control seeds and plants were soaked and watered with water only. The plants were maintained at 30 ~ under a daily light regime of 16h light: 8 h dark. Light was provided by fluorescent lamps (cool white, F40CW; General Electric, Cleveland, O., USA) at a fluence of 500 W m -2. Ten d after planting the seedlings were gently removed from the vermiculite and divided into three parts: 1) leaves, 2) detipped roots, and 3) root tips (terminal 1.5 mm) which included the cap and the apex of the root proper. Fluridone was obtained through the courtesy of Dr. Jim Barrenfine, Lilly Research Laboratories, Greenfield, Ind., USA). Determination of ABA content. ABA was extracted for highperformance liquid chromatography (HPLC) quantification (sensitive to 2.5 ng ABA g-1 fresh weight [FW]) according to procedures similar to those described by Ciha et al. (1977). A minimum of three replications each utilizing at least 0.663 g fresh weight (FW) of tissue were made for each tissue. Tissues were homogenized in a tissue grinder in 80% ethanol (20 ml g - I FW). Completely homogenized tissue was placed in small flasks and shaken for 24 h at 4 ~ centrifuged at 2000g for 10 rain, and the supernatant filtered using a 0.22-lain syringetype filter (Mill• Corp., Bedford, Mass., USA). The filtrate was dried under nitrogen using a Fisher (Pittsburgh, Pa., USA) Model 190 sample concentrator, until all methanol had evaporated. The aqueous residue was reconstituted with 0.1M NaHCO 3 and partitioned against diethyl ether three times, saving the aqueous phase. The latter was adjusted to pH 2.8 with HC1, again partitioned against diethyl ether three times saving the ether phase, evaporated to dryness under nitrogen, and stored at -20 ~C. Preparative HPLC procedures were similar to those described by Ciha et al. (1977). Dried samples were reconstituted in 5% (v/v) methanol in 0.2 N acetic acid, centrifuged for 5 rain at 2500 g, and microfiltered. Known volumes of filtered sample were injected into a Laboratory Data Control HPLC system (Riviera Beach, Fla., USA) with an LDC microprocessor that controlled the solvent program, plotted separations detected by a dual-channel ultraviolet monitor (254 and 269 nm), calculated peak height and area, and compared peak area with a previously determined internal standard to estimate quantity. We standardized our preparative protocol using a spherisorb ODS 5-lain reverse phase column (Alltech Associates, Deerfield, Ill., USA) with a C18 pre-column cartridge (Alltech Associates) and a 30-min nonlinear gradient (10-100% methanol adjusted to pH 3 with 1 N acetic acid) at a flow rate of 1 ml min-1 at 22 ~C. Under these conditions the retention time for ABA was 21-23 min. The 21 23-min fraction was collected, evaporated to dryness under nitrogen gas, and stored at -20 ~C.
343
Quantification of ABA was accomplished using a 20-rain • run utilizing the colunan arrangement and HPLC system described above, using 30% acetonitrile (adjusted to pH 3 with 1 N phosphoric acid) at a flow rate of 1 ml m i n - 1 at 22 ~C. Under these conditions the retention times were 8.8-8.9 rain for trans, trans-ABA and 10.0-10.2 min for cis, trans-ABA. Measuring growth and gray• of roots. Primary roots (length = 15 • 2 ram) of treated and untreated plants were placed in a closed, humid (relative humidity=90• chamber at room temperature in light from fluorescent lamps (cool white, F20T12-CW; General Electric) having a fluence of 310 laW cm -2. Root growth and gravicurvature were measured by shadowgraphing the roots after 1, 2, 4 and 6 h. After determining the mean gray• and growth rates (• standard deviation, SD) for Fluridone-treated and control roots, their growth and gray• were compared. Student's t test (see Sokal and Rohlf 1969, pp. 220-223) was used to evaluate the significance levels of differences between means. Differences with probability levels greater than 5% were considered insignificant.
Results T h e a m o u n t s ( + S D ) o f A B A p r e s e n t in l e a v e s , detipped roots, and root tips of control and Flurid o n e - t r e a t e d s e e d l i n g s o f Z . m a y s a r e p r e s e n t e d in T a b l e 1. G r o w t h r a t e s a n d g r a v i c u r v a t u r e s ( • S D ) of Fluridone-treated and control roots are present e d in T a b l e 2. N e i t h e r t h e g r o w t h r a t e s n o r g r a v i c u r v a t u r e s o f t r e a t e d a n d u n t r e a t e d r o o t s w e r e significantly different.
Discussion A l t h o u g h t h e a m o u n t s o f A B A r e p o r t e d in T a b l e 1 f o r r o o t s o f Z . rnays a r e s o m e w h a t g r e a t e r t h a n Table 1. ABA content of control and Fluridone-treated seedlings of Zea mays cv. Tx 5855 ABA content (gg g - I FW)
Root tips Detipped roots Leaves
Control
Fluridone-treated
N.D. a 0.260 • 0.039 0.114 • 0.024
N.D. N.D. N.D.
a N.D. = none detected, with an analysis sensitivity of 2.5 ng ABA g 1 FW Table 2. Growth rates and gray• ( • SD) of Fluridonetreated and control roots of Zea mays cv. Tx 5855 Time (h)
1
2
Growth rate (ram h - l) Control 1.0--0.3 1.1• +Fluridone 1.2+0.2 1.2• Gray• (degrees) Control 12-1=4 +Fluridone 9•
27• 33•
8
4
6
1.1• 1.0•
1.0i0.2 1.1•
69• 74•
77• 79•
8
344
R. Moore and J. D. Smith: ABA content and root graviresponsiveness of Fluridone-treated seedlings
those reported for roots of some other plants, they nevertheless are well within the range of those reported in the literature for plant roots (see review by Audus 1983). Also, our results indicate that the amount of ABA in root tips ofZ. mays cv. Tx 5855 is less than that of mature regions of the root, an observation noted previously for certain other cultivars of Z. mays (Rivier and Pilet 1981). Finally, our inability to detect any ABA in root tips of Z. mays cv. Tx 5855 (Table 1) is consistent with a similar observation by Rivier and Pilet (1981), who did not detect any ABA in tips of primary roots of Z. mays cv. Orla 264. We did not detect any ABA in Fluridone-treated seedlings of Z. mays (Table 1). Consistent with this observation is the fact that Fluridone-treated seeds o f Z . mays are viviparous (Fong et al. 1983 a, b) a characteristic long associated with ABA content (see review by Taylorson and Hendricks 1977). Significantly, ABA treatment reverses this Fluffdone-induced vivipary ofZ. mays seeds (Fong et al. 1983b). Also, Fluridone-treated seedlings of Z. mays are hypersensitive to gibberellic acid, a hormone usually causing responses antagonist to those caused by ABA (Devlin et al. 1980). Taken together, these results indicate that Fluridone completely inhibits ABA synthesis in Z. mays seedlings. As far as we are aware, this is the first report of a treatment having such an effect on ABA synthesis in plant tissues. Fluridone inhibits carotenoid biosynthesis in Z. mays seedlings (Fong et al. 1983 a, b). Therefore, the absence of ABA in Fluridone-treated seedlings (Table 1) correlates positively with carotenoid deficiency, supporting the suggestion that carotenoids may be precursors to ABA (Fong etal. 1983b). Consistent with this conclusion are the observations that 1) carotenoid-deficient mutants of Z. mays are ABA deficient (Smith 1983), and 2) other herbicides that inhibit the biosynthesis of carotenoids (e.g., Norflurazon) also inhibit ABA accumulation in light-grown plants (Henson 1984). The rates of growth and gravicurvature of Fluridone-treated (i.e., ABA-lacking) roots were not significantly different from those of control (i.e., ABA-containing) roots (Table 2). These results indicate that ABA is not necessary for positive gravicurvature by primary roots of Z. mays. Consistent with this conclusion is the observation that although ABA is absent from tips of primary roots of Z. mays cv. Orla 264 (Rivier and Pilet 1981), these roots are nevertheless strongly graviresponsive (Pilet and Elliott 1981).
This research was supported by the Texas Agricultural Experiment Station Project No. 6371 and the University Research Committee of Baylor University. We thank Bill Kenyon for his helpful suggestions, and Ann Blakey for her excellent technical assistance. We are also most grateful to Dr. Jim Barrentine (Lilly Research Labs., Greenfield, Ind., USA) for providing us with Fluridone.
References Audus, L.J. (1983) Abscisic acid in root growth and geotropism. In: Abscisic acid, pp. 421 478, Addicott, F.T., ed. Praeger, New York Ciha, A.J., Brenner, M.L., Brun, W. A. (1977) Rapid separation and quantification of abscisic acid from plant tissues using high performance liquid chromatography. Plant Physiol. 59, 821 826 Devlin, R.M., Kisiel, M.J., Kostusiak, A. (1980) Enhancement of gibberellic acid sensitivity in corn (Zea mays) by fluridone and R-40244. Weed Sci. 28, 11-12 Evans, M.L., Mulkey, T.J. (1982) Comparative effects of auxin and abscisic acid in growth, hydrogen ion etflux and gravitropism in primary roots of maize. In: Plant growth substances 1982, pp. 33-42, Wareing, P.F., ed. Academic Press, London New York Fong, F., Koehler, D.E., Smith, J.D. (1983 a) Fluridone induction of vivipary during maize seed development. In: Ill Int. Syrup. on Pre-hmwest Sprouting in Cereals, pp. 188-196, Kruger, J.E., LaBerge, D.E., eds. Westview Press, Boulder, Colo. Fong, F., Smith, J.D., Koehler, D.E. (1983b) Early events in maize seed development. Plant Physiol. 73, 899-901 Henson, I.E. (1984) Inhibition of abscisic acid accumulation in seedling shoots of pearl millet (Pennisetum arnericanum (L.) Leeke) following induction of chlorosis by Norflurazon. Z. Pflanzenphysiol. 114, 35 43 Jackson, M.B., Barlow, P.W. (1981) Root geotropism and the role of growth regulators from the cap: a re-examination. Plant Cell Environ. 4, 107-123 Mertens, R., Weiler, E.W. (1983) Kinetic studies on the redistribution of endogenous growth regulators in gravireacting plants organs. Planta 158, 339-348 Milborrow, B.V. (1983) Pathways to and from abscisic acid. In: Abscisic acid, pp. 79-111, Addicott, F.T., ed. Praeger~ New York Moore, R. (1984) How roots perceive and respond to gravity. Am. Biol. Teach. 47, 257-265 Mulkey, T.J., Evans, M.L., Kuzmanoff, K.M. (1983) The kinetics of abscisic acid action on root growth and gravitropism. Planta 157, 150-157 Pilet, P.E., Elliott, M.C. (1981) Some aspects of the control of root growth and georeaction: the involvement of indoleacetic acid and abseisic acid. Plant Physiol. 6/, 1047 1050 Rivier, L., Pilet, P.E. (1981) Abscisic acid levels in the root tips of seven Zea mays varieties. Phytochemistry 20, 17 19 Smith, J.D. (1983) Regulation of seed development in Zea mays. (Abstr.) Am. J. Bot. 70 (5, 2), 63 Sokal, R.R., Rohlf, F.J. (1969) Biometry. Freeman, San Francisco Taylorson, R.B., Hendricks, S.B. (1977) Dormancy in seeds. Annu. Rev. Plant Physiol. 28, 331-354 Wilkins, M.B. (1984) Gravitropism. In: "Advanced plant physiology, pp. 163-185, Wilkins, M_B., ed. Pitman, London Received 20 March; accepted 5 June 1984
Planta (1984)162:342 344
9 Springer-Verlag 1984
Growth, graviresponsiveness and abscisic-acid content of Zea mays seedlings treated with Fluridone Randy Moore and James D. Smith Department of Biology, Baylor University, Waco, TX 76798, and Department of Soil and Crop Science, Texas A and M University, College Station, TX 77843, USA
Abstract. Ten-d-old seedlings o f Z e a mays L. cv. Tx 5855 treated with 1-methyl-3-phenyl-5-(3-[trifluoromethyl]phenyl)-4-(1H)-pyridinone (Fluridone) were analyzed for abscisic acid (ABA) content using high-performance liquid chromatography with an analysis sensitivity of 2.5 ng ABA g-l fresh weight (FW). Seedlings were divided into three portions: leaves, detipped roots, and root tips (terminal 1.5 ram). Control plants (water treatment only; no Fluridone) were characterized by the following amounts of ABA: leaves, 0.1144-0.024 (standard deviation) gg ABA g-1 FW; detipped roots, 0.2604-0.039+ gg ABA g-1 FW; root tips, no ABA detected. We did not detect any ABA in tissues of Fluridone-treated plants. Primary roots of treated and untreated seedlings were strongly graviresponsive, with no significant differences between the curvatures or the growth rates of primary roots of Fluridone-treated and control seedlings. These results indicate that 1) Fluridone completely inhibits ABA synthesis, and 2) ABA is not necessary for positive gravitropism by primary roots of Zea mays.
Key words: Abscisic acid and root growth - Fluffdone - Gravitropism (root) - Root growth - Zea (root growth).
Introduction
There is a large amount of evidence indicating that the root cap is the source of a growth-inhibiting influence on the root, and that an imbalance of inhibitor(s) originating in the cap is the basis for root gravicurvature (see reviews by Jackson and Barlow 1981; Moore 1984; Wilkins 1984). Transport of the inhibitor(s) from the cap to the elongating zone (at greater concentrations in the lower Abbreviations: ABA = abscisic acid; Fluridone = 1-methyl-3phenyl-5-(3-[trifluoromethyl]phenyl)-4-(1 H)-pyridinone; FW = fresh weight; SD = standard deviation
half than in the upper half) of the root presumably results in more rapid growth of the upper half of the root, thereby accounting for positive gravicurvature (see reviews by Audus 1983; Wilkins 1984). One inhibitor that many investigators believe to be involved in root gravitropism is abscisic acid (ABA) (Audus 1983; Wilkins 1984). Evidence supporting this conclusion includes the following observations: 1) Abscisic acid is present in caps and tips of roots. 2) Abscisic acid inhibits root elongation. 3) Asymmetrical application of ABA induces root curvature. 4) Light-mediated inhibition of root elongation is apparently a consequence of ABA synthesis in root caps. 5) Gravicurvature by roots of several cultivars of Z. mays that require light for graviresponsiveness correlates positively with increased levels of ABA in their root caps. 6) The onset of graviresponsiveness correlates positively with an increase in ABA content in the lower half and a concomitant decrease in ABA in the upper half of horizontally-oriented roots (see reviews by Jackson and Barlow 1981; Audus 1983; Wilkins 1984). Recently, however, several investigators have questioned the involvement of ABA in root graviresponsiveness, based on the following observations: 1) There is little evidence that intact roots can transport ABA from the root cap to the zone of elongation. 2) If the cap inhibitor is ABA, then placing root caps on growing coleoptiles would be expected to inhibit elongation. However, an auxinlike stimulation (instead of an ABA-like inhibition) of growth occurs in response to this treatment. 3) There is no significant lateral asymmetry of ABA in gravistimulated roots (Mertens and Weiler 1983). 4) Abscisic acid has been reported to have no effect on root elongation or to promote, at least initially, root elongation (Evans and Mulkey 1982; Mulkey et al. 1983; also see reviews by Jackson and Barlow 1981; Audus 1983). Several investigators have suggested that ABA is synthesized via the carotenoid pathway (see re-
R. Moore and J. D. Smith: ABA content and root graviresponsiveness of Fluridone-treated seedIings v i e w b y M i t b o r r o w 1983). I f t h i s is t r u e , t h e n i n h i biting carotenogenesis should also prevent the synthesis o f A B A . I n this s t u d y w e t r e a t e d s e e d a n d seedlings ofZ. mays with Fluridone, an inhibitor of c a r o t e n o i d s y n t h e s i s ( F o n g et al. 1 9 8 3 a , b). W e s u b s e q u e n t l y m e a s u r e d the g r o w t h a n d gray• s p o n s i v e n e s s o f t h e t r e a t e d r o o t s in o r d e r to d e t e r mine if ABA was necessary for gray• ness.
M a t e r i a l and methods Plant material and growth conditions. Caryopses of inbred lines of Zea mays L. cv. Tx 5855 were obtained from plants grown at the Texas A and M University Farm. They were soaked in Fluridone (100 nag 1-1) for 10 h and then planted in vermiculite in a controlled-environment growth chamber. Seedlings were watered regularly with Fluridone (100 mg l-l). Control seeds and plants were soaked and watered with water only. The plants were maintained at 30 ~ under a daily light regime of 16h light: 8 h dark. Light was provided by fluorescent lamps (cool white, F40CW; General Electric, Cleveland, O., USA) at a fluence of 500 W m -2. Ten d after planting the seedlings were gently removed from the vermiculite and divided into three parts: 1) leaves, 2) detipped roots, and 3) root tips (terminal 1.5 mm) which included the cap and the apex of the root proper. Fluridone was obtained through the courtesy of Dr. Jim Barrenfine, Lilly Research Laboratories, Greenfield, Ind., USA). Determination of ABA content. ABA was extracted for highperformance liquid chromatography (HPLC) quantification (sensitive to 2.5 ng ABA g-1 fresh weight [FW]) according to procedures similar to those described by Ciha et al. (1977). A minimum of three replications each utilizing at least 0.663 g fresh weight (FW) of tissue were made for each tissue. Tissues were homogenized in a tissue grinder in 80% ethanol (20 ml g - I FW). Completely homogenized tissue was placed in small flasks and shaken for 24 h at 4 ~ centrifuged at 2000g for 10 rain, and the supernatant filtered using a 0.22-lain syringetype filter (Mill• Corp., Bedford, Mass., USA). The filtrate was dried under nitrogen using a Fisher (Pittsburgh, Pa., USA) Model 190 sample concentrator, until all methanol had evaporated. The aqueous residue was reconstituted with 0.1M NaHCO 3 and partitioned against diethyl ether three times, saving the aqueous phase. The latter was adjusted to pH 2.8 with HC1, again partitioned against diethyl ether three times saving the ether phase, evaporated to dryness under nitrogen, and stored at -20 ~C. Preparative HPLC procedures were similar to those described by Ciha et al. (1977). Dried samples were reconstituted in 5% (v/v) methanol in 0.2 N acetic acid, centrifuged for 5 rain at 2500 g, and microfiltered. Known volumes of filtered sample were injected into a Laboratory Data Control HPLC system (Riviera Beach, Fla., USA) with an LDC microprocessor that controlled the solvent program, plotted separations detected by a dual-channel ultraviolet monitor (254 and 269 nm), calculated peak height and area, and compared peak area with a previously determined internal standard to estimate quantity. We standardized our preparative protocol using a spherisorb ODS 5-lain reverse phase column (Alltech Associates, Deerfield, Ill., USA) with a C18 pre-column cartridge (Alltech Associates) and a 30-min nonlinear gradient (10-100% methanol adjusted to pH 3 with 1 N acetic acid) at a flow rate of 1 ml min-1 at 22 ~C. Under these conditions the retention time for ABA was 21-23 min. The 21 23-min fraction was collected, evaporated to dryness under nitrogen gas, and stored at -20 ~C.
343
Quantification of ABA was accomplished using a 20-rain • run utilizing the colunan arrangement and HPLC system described above, using 30% acetonitrile (adjusted to pH 3 with 1 N phosphoric acid) at a flow rate of 1 ml m i n - 1 at 22 ~C. Under these conditions the retention times were 8.8-8.9 rain for trans, trans-ABA and 10.0-10.2 min for cis, trans-ABA. Measuring growth and gray• of roots. Primary roots (length = 15 • 2 ram) of treated and untreated plants were placed in a closed, humid (relative humidity=90• chamber at room temperature in light from fluorescent lamps (cool white, F20T12-CW; General Electric) having a fluence of 310 laW cm -2. Root growth and gravicurvature were measured by shadowgraphing the roots after 1, 2, 4 and 6 h. After determining the mean gray• and growth rates (• standard deviation, SD) for Fluridone-treated and control roots, their growth and gray• were compared. Student's t test (see Sokal and Rohlf 1969, pp. 220-223) was used to evaluate the significance levels of differences between means. Differences with probability levels greater than 5% were considered insignificant.
Results T h e a m o u n t s ( + S D ) o f A B A p r e s e n t in l e a v e s , detipped roots, and root tips of control and Flurid o n e - t r e a t e d s e e d l i n g s o f Z . m a y s a r e p r e s e n t e d in T a b l e 1. G r o w t h r a t e s a n d g r a v i c u r v a t u r e s ( • S D ) of Fluridone-treated and control roots are present e d in T a b l e 2. N e i t h e r t h e g r o w t h r a t e s n o r g r a v i c u r v a t u r e s o f t r e a t e d a n d u n t r e a t e d r o o t s w e r e significantly different.
Discussion A l t h o u g h t h e a m o u n t s o f A B A r e p o r t e d in T a b l e 1 f o r r o o t s o f Z . rnays a r e s o m e w h a t g r e a t e r t h a n Table 1. ABA content of control and Fluridone-treated seedlings of Zea mays cv. Tx 5855 ABA content (gg g - I FW)
Root tips Detipped roots Leaves
Control
Fluridone-treated
N.D. a 0.260 • 0.039 0.114 • 0.024
N.D. N.D. N.D.
a N.D. = none detected, with an analysis sensitivity of 2.5 ng ABA g 1 FW Table 2. Growth rates and gray• ( • SD) of Fluridonetreated and control roots of Zea mays cv. Tx 5855 Time (h)
1
2
Growth rate (ram h - l) Control 1.0--0.3 1.1• +Fluridone 1.2+0.2 1.2• Gray• (degrees) Control 12-1=4 +Fluridone 9•
27• 33•
8
4
6
1.1• 1.0•
1.0i0.2 1.1•
69• 74•
77• 79•
8
344
R. Moore and J. D. Smith: ABA content and root graviresponsiveness of Fluridone-treated seedlings
those reported for roots of some other plants, they nevertheless are well within the range of those reported in the literature for plant roots (see review by Audus 1983). Also, our results indicate that the amount of ABA in root tips ofZ. mays cv. Tx 5855 is less than that of mature regions of the root, an observation noted previously for certain other cultivars of Z. mays (Rivier and Pilet 1981). Finally, our inability to detect any ABA in root tips of Z. mays cv. Tx 5855 (Table 1) is consistent with a similar observation by Rivier and Pilet (1981), who did not detect any ABA in tips of primary roots of Z. mays cv. Orla 264. We did not detect any ABA in Fluridone-treated seedlings of Z. mays (Table 1). Consistent with this observation is the fact that Fluridone-treated seeds o f Z . mays are viviparous (Fong et al. 1983 a, b) a characteristic long associated with ABA content (see review by Taylorson and Hendricks 1977). Significantly, ABA treatment reverses this Fluffdone-induced vivipary ofZ. mays seeds (Fong et al. 1983b). Also, Fluridone-treated seedlings of Z. mays are hypersensitive to gibberellic acid, a hormone usually causing responses antagonist to those caused by ABA (Devlin et al. 1980). Taken together, these results indicate that Fluridone completely inhibits ABA synthesis in Z. mays seedlings. As far as we are aware, this is the first report of a treatment having such an effect on ABA synthesis in plant tissues. Fluridone inhibits carotenoid biosynthesis in Z. mays seedlings (Fong et al. 1983 a, b). Therefore, the absence of ABA in Fluridone-treated seedlings (Table 1) correlates positively with carotenoid deficiency, supporting the suggestion that carotenoids may be precursors to ABA (Fong etal. 1983b). Consistent with this conclusion are the observations that 1) carotenoid-deficient mutants of Z. mays are ABA deficient (Smith 1983), and 2) other herbicides that inhibit the biosynthesis of carotenoids (e.g., Norflurazon) also inhibit ABA accumulation in light-grown plants (Henson 1984). The rates of growth and gravicurvature of Fluridone-treated (i.e., ABA-lacking) roots were not significantly different from those of control (i.e., ABA-containing) roots (Table 2). These results indicate that ABA is not necessary for positive gravicurvature by primary roots of Z. mays. Consistent with this conclusion is the observation that although ABA is absent from tips of primary roots of Z. mays cv. Orla 264 (Rivier and Pilet 1981), these roots are nevertheless strongly graviresponsive (Pilet and Elliott 1981).
This research was supported by the Texas Agricultural Experiment Station Project No. 6371 and the University Research Committee of Baylor University. We thank Bill Kenyon for his helpful suggestions, and Ann Blakey for her excellent technical assistance. We are also most grateful to Dr. Jim Barrentine (Lilly Research Labs., Greenfield, Ind., USA) for providing us with Fluridone.
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