Transport of 14C-lableled indoleacetic acid in Vicia root segments

IAA transport in Vicia root segments was investigated for comparisonwith that in intact roots. Lanolin paste (1-mm-wide ring) oragar blocks (3?3?1.5mm), both containing IAA-2-¹⁴C were appliedto the surface or a cut end of the root segments, respectively;transported ¹⁴C was collected in receiver agar blocks placedon the cut end of the segments. When lanolin paste was appliedto 5-mm segments, basipetal transport of IAA predominated overacropetal transport. When agar blocks were applied to 1- and2-mm segments, the same was true; in longer segments (3 and5 mm long), however, basipetal movement occurred predominantlyat first but was surpassed by acropetal movement after 2–3hr. Among the segments tested (regions 2–4, 4–6and 8–10 mm from the tip), the most apical one showedthe distinctest predominancy of basipetal movement. The velocitiesof the acropetal and basipetal movement of the ¹⁴C were estimatedat 3–3.8 and 8–12 mm/hr, respectively. Autoradiographicstudy and the experiment in which wire was inserted longitudinallythrough the central part of the segments showed that basipetalmovement occurred mainly through the outer part of the rootsand acropetal movement mainly through the central cylinder.The present results were compatible with those obtained previouslywith intact roots. Some properties of polar movement, such asits specificity, inhibition by TIBA, and dependency on terneprature are described. (Received March 22, 1978; )     

Auxin Transport in Roots: VI. MOVEMENT OF IAA THROUGH DIFFERENT ZONES OF ZEA ROOTS

The movement of IAA through 6-mm segments excised 1 mm, 7 mm,and 13 mm behind the apex of the primary root of Zea mays seedlingshas been investigated at temperatures between 10 and 25°C. In all segments, and at all temperatures, the movement of IAAwas polarized acropetally, more IAA being found in apical receiverblocks than in basal ones after transport periods of up to 24h. The amounts of IAA which moved acropetally through a segmentdecreased as the segment was taken at an increasing distancebehind the root apex. Similarly, at least after transport periodsof 8 h, more IAA moved basipetally through the apical segmentthan through the basal ones. At 10°C the velocity of acropetal movement was similar inall three segments, but the acropetbut the acropetal flux wasgreatest in the apical segment and smallest in the most basalone. The same situation appears to exist at the other temperatures. The flux and velocity of the acropetal movement of IAA througha 6-mm segment taken 7 mm behind the apex of the root were similarto those previously reported for the acropetal movement througha 12-mm segment excised 1 mm behind the apex. The smaller amountsof IAA which move acropetally through longer root segments aretherefore attributable to a limitation of the flux in the mostbasal regions of the segment.     

Polar transport of kinetin in tissues of radish

Polar transport of kinetin-8-¹⁴C occurred in segments of petioles, hypocotyls, and roots of radish (Raphanus sativus L.). The polarity was basipetal in petioles and hypocotyls and acropetal in roots. In segments excised from seedlings with fully expanded cotyledons, indole-3-acetic acid was required for polarity to develop. In hypocotyl segments isolated at this stage, basipetal and acropetal movements were equal during the first 12 hours of auxin treatment after which time acropetal movement declined. Pretreatment with auxin eliminated this delay in the appearance of polarity. In hypocotyl segments excised from seedlings with expanding cotyledons, exogenous auxin was unnecessary for polarity. Potassium cyanide abolished polarity at both stages of growth by allowing increased acropetal movement. The rate of accumulation of kinetin in receiver blocks was greater than the in vivo increase in cytokinin content of developing radish roots.     

Auxin Transport in Tendril Segments of Passiflora caerulea

The movement of auxin through tendril segments of Passiflora caerulca L. has been investigated using IAA-2-¹⁴C. It has been shown that (1) flux of IAA through the segments is strongly polarized basipetally: (2) the amount of ¹⁴C recovered in the basal receiver blocks increases linearly within a transport period of 6 h; (3) velocity of basipetal transport is 14.5 mm h^?1; (4) at least 70% of the radioactivity in the receiver blocks is confined to the IAA molecule: approximately 55% of ¹⁴C from methanolic extracts of the segments is IAA: (5) at low temperatures (2–4°C) the basipetal transport is abolished; (6) white light promotes basipetal transport, and this effect is abolished in a CO₂-free atmosphere; (7) no difference could be detected in ¹⁴C content between dorsal and ventral halves of tendril segments nor among individual dorsal and ventral receiver blocks.     

Auxin transport in roots

Summary Light promotes the net acropetal movement of ¹⁴C through 6-mm subapical segments of dark-grown roots of Zea mays supplied at their basal ends with 1 M IAA-1-¹⁴C in agar blocks. This promotion occurs only when the segments are irradiated during the transport period, and both red and blue light appear to be as effective as white light at the radiant flux densities used in this investigation. The promotion is not found if the segments are pretreated with light and then returned to darkness before the trasport of IAA-1-¹⁴C is determined. The very slight basipetal movement of ¹⁴C through the segments supplied with an apical source of IAA-1-¹⁴C is unaffected by light.Only one radioactive substance is found in the apical receiver blocks. This substance has an R_f virtually identical to those of the stock solution of IAA incorporated into the donor block and of unlabelled IAA. The movement of radioactivity into the receiver blocks through, the illuminated segments therefore appears to reflect the movement of IAA. Light thus increases the acropetal movement of IAA through the Zea root segment.The primary roots of Zea mays var. Giant Horse Tooth seedlings grown in total darkness do not exhibit a positive geotropic response. When the seed is orientated with the embryo uppermost the radicle grows out horizontally. On exposure to light, however, the roots bend down. This reaction appears about 3–9 hours after the onset of illumination, and white, red and blue light appear to be equally effective at the flux densities employed in this study. Green light in the spectral band between 510–530 nm did not appear to induce this positive geotropic responsiveness.     

Maintenance of polar auxin transport in Zea coleoptiles by anaerobic metabolism

Summary The basipetal movement of IAA in 5-mm Zea coleoptile segments is drastically reduced under anaerobic conditions, but it remains greater than acropetal movement which is closely similar in the presence and absence of oxygen. The polarity of IAA movement has thus been confirmed in Zea coleoptile segments which have been deprived of oxygen. This net polar flux is dependent upon anaerobic metabolism since it is abolished in the presence of the metabolic inhibitiors sodium fluoride and iodoacetic acid.Acropetal movement of IAA is unaffected by the presence of sodium fluoride in air or anaerobic conditions. Uptake of IAA from a basal donor is not affected by sodium fluoride in air, but under anaerobic conditions the inhibitor decreased uptake by approximately 13%.Under anaerobic conditions both inhibitors reduce basipetal movement of IAA to the level of acropetal movement, and both decrease the total uptake of IAA from an apical donor by up to 30–45%. Under aerobic conditions sodium fluoride has no marked effect upon either the uptake of IAA from an apical donor or the basipetal movement of IAA by the segments. On the other hand, iodoacetic acid greatly decreased the uptake of IAA by the segments in air, but the same fraction of the total IAA taken up was recovered in the receiving block in the presence and absence of the inhibitor.This research was supported by Grant Number 83/6 to Professor M. B. Wilkins from the U. K. Agricultural Research Council.     

Basipetally polarised transport of [3H]gibberellin A1 and [14C]gibberellin A3, and acropetal polarity of [14C]indole-3-acetic acid transport,in stelar tissues of Phaseolus coccineus roots

Summary Movement of both [³H]GA₁ and [¹⁴C]GA₃ through root segments from P. coccineus seedlings was basipetally polarised. The basipetal/acropetal ratio of radioactivity from [³H]GA₁ in agar receiver blocks was 9.2 for apical, elongating segments, and 4.0 for more basal, non-elongating segments. Polarity of gibberellin transport was restricted to the stele, and absent from cortical tissues. Transport of [¹⁴C]IAA through root segments to agar receivers was preferentially acropetal, particularly so in the stele. Despite the existence of basipetal polarity of gibberellin transport in the root, [³H]GA₁ injected into cotyledons moved into and acropetally along the seedling root.     

Auxin-Promoted Transport of Metabolites in Stems of Phaseolus vulgaris L.: EFFECTS REMOTE FROM THE SITE OF HORMONE APPLICATION

Phloem transport in stems of Phaseolus vulgaris was found tobe sensitive to treatment with the auxin transport inhibitor,2,3,5-triidobenzoic acid (TIBA). The response was dependenton the concentration of TIBA applied. A concentration of TIBA(0?5% in lanolin) which did not interfere with normal phloemtransport proved inhibitory to both basipetal transport of IAAand the acropetal component of IAA-promoted metabolite transport.In contrast, both acropetal IAA transport and basipetal IAA-promotedmetabolite transport were unaffected by TIBA treatment. Theinhibitory effect of TIBA on acropetal IAA-promoted transportwas overcome by providing IAA below the point of TIBA application.Both acropetal and basipetal IAA-promoted transport in stemsegments were unaccompanied by any corresponding changes inthe accumulation of [¹⁴C]sucrose by the segments.     

Auxin transport in roots

Summary The net uptake and movement of radioactivity by 12-mm root segments of Zea mays have been studied as a function of time at 5, 15 and 25° C. Segments were supplied with an agar donor block containing 1 M IAA-1-¹⁴C or IAA-2-¹⁴C continuously or for a limited period of time (pulse-labelling). In the latter case the original donor block was replaced either by a plain agar block or by one containing 1 M unlabelled IAA. Receiver blocks were placed at the other end of the segments.The net uptake of radioactivity from the donor block at 15° C was greater at the basal end than at the apical end of the segment. At 5 and 15° C, the net uptake from a basal donor was virtually linear with time but at 25° C the rate of net accumulation decreased after about 10 h. Decarboxylation of IAA undoubtedly occurred at 15 and 25° C when the concentration in the tissue attained a high value.An acropetally polarised movement of radioactivity into the receiver blocks occurred regardless of whether the results were based on the actual amounts of radioactivity in the receiver block, or on the amounts in the receiver block expressed as a percentage of the net total radioactivity accumulated from the donor block. Only one radioactive substance was present in the receiver block and it ran to the same R_f as IAA in the isopropanol: ammonium: water solvent system.The amounts of radioactivity moving into that part of the root segment at least 6 mm distant from the end in contact with either an apical or a basal donor block were assessed. An acropetal polarity in the movement of radioactivity was observed on the basis of the actual amounts of radioactivity in these distal parts of the segments, but no such polarity was evident when the amounts of radioactivity were expressed as a percentage of the net total accumulated from the donor block. At least 3 radioactive substances were present in the tissue in addition to the substance running to the same R_f as IAA. The distribution of radioactivity in the segment cannot therefore be used to assess the distribution of IAA.Acropetal movement of radioactivity into an apical receiver block is not dependent upon the continued uptake of IAA at the basal end of the segment. No distinct pulses of radioactivity were detected moving through the root segments.Only a small part of the radioactivity in the root segment appears to be located in the polar transport system, while the bulk is not. The polarity found in the movement of the bulk radioactivity within the segment seems to be related to the polarity in IAA uptake from the donor blocks.     

Basipetal and Acropetal Auxin Transport in Relation with Temperature

In stem sections of lentil seedlings, there is a typical polar movement of IAA labelled with ¹⁴C. The degree of polarity, expressed as the ratio of basipetal to acropetal transport, was (25°C) 7.6. A decrease (from 25° to 15°C) and an increase (from 25° to 30°C) of temperature cause a reduction of the IAA uptake by the sections and a decrease of both the basipetal and the acropetal translocation of IAA. Results suggest that the basipetal as well as the acropetal movement of auxin, are dependent of a metabolical component which is discussed.     

Decarboxylation and transport of auxin in segments of sunflower and cabbage roots

Summary The movement of ¹⁴C from indole-3-acetic acid (IAA) ¹⁴C has been examined in 5 mm root segments of dark-grown seedlings of Helianthus annuus and Brassica oleracea. Contaminants from distilled water, phosphate buffer and the razor-blade cutter increase the decarboxylation of IAA-¹⁴C, and cutting of root segments results in an activation of IAA-destroying enzymes at the cut surfaces. When these sources of errors were eliminated the following was shown: a) Both in sunflower and cabbage there is a slight acropetal flux of ¹⁴C through the root segments into the agar receiver blocks. The amount of ¹⁴C found in the receiver blocks increases with the lenght of the transport period. b) When the root segments, after the transport period, are cut in two equal parts and these assayed separately, the amounts of ¹⁴C in the two parts indicate a greater acropetal than basipetal transport. c) The total radioactivity of the receiver blocks is in part due to IAA-¹⁴C and in part to ¹⁴CO₂, the latter being a result of enzymatic destruction of auxin. d) Addition of ferulic acid, an inhibitor of IAA oxidases, to the receiver blocks markedly inhibits the decarboxylation of IAA-¹⁴C and thus increases the amount transported. This effect is more pronounced after a 20 hr than after a 6 hr transport period.     

Influence of Gamma Radiation on the Transport of Indolyl-3-Acetic Acid in Bark Tissue from Apple Trees

The influence of gamma radiation on the polarity of IAA translocation was investigated. Pieces of apple tree shoots, taken from 3-year-old trees, were irradiated in a dark irradiation chamber BK-⁶⁰Co source, 10 kCi. Doses from 20 to 150 krad were given. Strips of bark were peeled off both irradiated and control shoots. The middle parts of the bark strips were placed on agar blocks loaded with IAA-1-¹⁴C. The radioactivity of basal and acropetal segments was determined after about 20 h and results expressed as polarity quotient. It has been shown that irradiation with 20 krad (200 Gy) decreases the polarity quotient remarkably (polarity quotient of control equals 7; for 20 krad—2.9; 100 krad—1.6). The decrease in polarity results from inhibition of the basipetal translocation of IAA. Irradiated tissue shows a decrease in IAA uptake. Possible mechanisms of gamma irradiation effects on polar translocation of IAA are discussed.     

MOVEMENT OF IAA IN SECTIONS FROM SPIDER FLOWER (CLEOME HASSLERIANA) STAMEN FILAMENTS

Movement of IAA in spider flower (Cleome hassleriana Chod.) stamen filaments was studied by placing 2-mm sections horizontally between donor agar blocks containing ¹⁴C-labeled IAA and plain agar receiver blocks and measuring radioactivity in the donor and receiver blocks and filament sections by scintillation counting after the desired transport time. Movement was strictly polar and basipetal at all stages of floral development, except in open flowers just before stamen abscission when the amounts moving acropetally and basipetally were equal. The amount of IAA moved depended upon the stage of development. As buds aged more IAA was moved, until the buds opened and the stamen filaments reached maximum elongation; then the amount of IAA moving basipetally dropped drastically. There was an insignificant amount of acropetal IAA movement except just before stamen abscission. This change in IAA movement is not due to a change in filament diameter. In time-course studies the amount of IAA moved basipetally increased with time up to 5 hr and then declined slightly. The amount of radioactivity retained by sections increased until 8 hr. The amount of IAA moved in tip sections was less than that in mid or base sections; however, this can be partially explained by differences in uptake area of these sections. The relationship of these results to the hypothesis that changes in IAA movement are important in the control of stamen filament elongation and abscission is discussed.     

Auxin Transport in Secondary Tissues1

Auxin transport was investigated in excised stem segments ofNicotiana tabacum L. by the agar block technique using [1-¹⁴C]indol-3yl-acetic acid (IAA). The ability of the stems to transportauxin basipetally increased as secondary development proceeded;by contrast the ability of the pith to transport auxin declinedwith age. By separation of the stem tissues it was shown thatthe great majority of auxin transport took place in cells associatedwith the internal phloem and in cells close to the cambium;in both cases similar velocities of transport were found (c.5.0 mm h^–1 at 22°C). The effects of osmotic gradientson auxin transport through the internal phloem were investigated.IAA was found by chromatography to account for practically allthe radioactivity in receiver blocks and other extracts of stemsegments. The significance of these results is discussed.     

Auxin, transport and growth in intact roots of Vicia faba

Transport of IAA applied to the intact root of Vicia seedlingswas investigated in relation to root growth. The root was treatedat 3–4, 4–5 or 7–8 mm from the tip with anarrow ring of lanolin paste containing IAA or IAA-2-¹⁴C ingrowth or transport experiments, respectively. The growth processalong the root axis was examined in every 1-mm part from thetip at 30 min, 1 or 4 hr intervals. The elongation zone of thecontrol root was 1–9 mm from the tip. IAA treatment broughtabout no significant change in the growth of the region apicalto the treated site, whereas distinct inhibition of growth occurredin the region basal to the treated site within 1 hr. The transportof radioactivity was observed in both acropetal and basipetaldirections within 1 hr, but the latter predominated for 8 hror more; the nearer to the tip the treatment site, the longerthe predominance lasted. The velocities of acropetal and basipetaltransport were estimated at about 4 and 8 mm/hr, respectively.Autoradiographs of transverse section of roots showed that basipetaltransport occurred mainly through the outer part of the root,whereas acropetal transport occurred mainly through the innerpart, the central cylinder. It may be concluded that the basipetallytransported IAA which passed through the outer part of the rootinhibited the elongation of the intact root. (Received November 25, 1975; )     

The Polarity of Movement of Endogenously Produced IAA in Relation to a Gravity Perception Mechanism

The polarity of radioactive IAA transport in segments of theleaf sheath base of Avena fatua L. is reversed upon their inversionthrough 180° transport towards the apex being greater thantowards the base. Changes in rates of transport leading to thisreversal can be detected within 10–20 min, correlatingwith the timing of statolith movements from the base to theapex of statocyte cells and conforming to a proposed model forthe gravity control of auxin transport. Transport of H³-trytophan-derived H³-IAA was found to undergosimilar changes in polarity upon re-orientation as did exogenouslyapplied H³ or C¹⁴ IAA. It is concluded that the proposed modelalso relates to the movement of endogenously produced IAA.     

Movement of Indoleacetic Acid in Coleoptiles of Avena sativa L. II. Suspension of Polarity by Total Inhibition of the Basipetal Transport

Acropetal and basipetal movement of indole-3-acetic acid through coleoptiles of Avena sativa L. was studied. Sections 10-mm long were supplied with either apical or basal sources containing C(14) carboxyl-labeled indoleacetic acid (10(-5)m). Anaerobic conditions inhibit metabolically dependent movement (transport) thus reducing basipetal but not acropetal movement. Total inhibition of basipetal transport abolishes the polarity of auxin uptake and movement. The nonpolar movement that remains in anaerobic sections is free diffusion with an average diffusion coefficient of approximately 1 x 10(-4) mm(2) per second. During an 8-hour diffusion, at least the first millimeter of the section comes to equilibrium at approximately the same concentration as the donor.Acropetal movement is probably by diffusion and is accompanied by an aerobic immobilization of indoleacetic acid that increases more than proportionally to concentration. Anaerobic conditions totally prevent this immobilization and reduce acropetal uptake but not the amount of indoleacetic acid moving into the upper parts of the section; there is, therefore, no evidence for acropetal transport.Polarity of auxin movement in aerobic coleoptile sections is achieved by strict basipetal transport of auxin. The basipetal transport may intensify the polarity by recycling auxin that is moving acropetally.     

Influence of 2,3,5-Triiodobenzoic Acid and 1-N-Naphthylphthalamic Acid on Indoleacetic Acid Transport in Carnation Cuttings: Relationship with Rooting

³H-IAA transport in excised sections of carnation cuttings was studied by using two receiver systems for recovery of transported radioactivity: agar blocks (A) and wells containing a buffer solution (B). When receivers were periodically renewed, transport continued for up to 8 h and ceased before 24 h. If receivers were not renewed, IAA transport decreased drastically due to immobilization in the base of the sections. TIBA was as effective as NPA in inhibiting the basipetal transport irrespective of the application site (the basal or the apical side of sections). The polarity of IAA transport was determined by measuring the polar ratio (basipetal/acropetal) and the inhibition caused by TIBA or NPA. The polar ratio varied with receiver, whereas the inhibition by TIBA or NPA was similar. Distribution of immobilized radioactivity along the sections after a transport period of 24 h showed that the application of TIBA to the apical side or NPA to the basal side of sections, increased the radioactivity in zones further from the application site, which agrees with a basipetal and acropetal movement of TIBA and NPA, respectively. The existence of a slow acropetal movement of the inhibitor was confirmed by using ³H-NPA. From the results obtained, a methodological approach is proposed to measure the variations in polar auxin transport. This method was used to investigate whether the variations in rooting observed during the cold storage of cuttings might be related to changes in polar auxin transport. As the storage period increased, a decrease in intensity and polarity of auxin transport occurred, which was accompanied by a delay in the formation and growth of adventitious roots, confirming the involvement of polar auxin transport in supplying the auxin for rooting. Received April 19, 1999; accepted December 2, 1999     

Decarboxylation and Transport of Auxin in Segments of Sunflower and Cabbage Roots. II. A Chromatographic Study Using IAA-1-14C and IAA-5-3H

The polar movement of IAA has been examined in 5-mm root segments of Brassica oleracea and Helianthus annum. The movement was studied partly with IAA-1-¹⁴C and partly with IAA-5-³H. In both plants a slight acropetal flux of ¹⁴C and IAA-³H was found through the segments. The recovered radioactivity in the agar receiver blocks and in the receiver end of the segments increased as a function of time. A large portion of the applied IAA was converted on the cut surfaces and in the tissues of the segments. Chromatographic analysis indicated different destruction products when estimated by scintillation counting and by spraying with in-dole reagent (DMCA). Chromatograms run in isopropanol: ammonia: water, 8:1:1, yielded three different substances, one spot near the starting line and one near the front, neither of which has been identified. Finally there was a spot with R_f 0.4–0.6, probably representing IAA.     

Auxin transport in roots

Summary The reasons underlying the initial increase and subsequent decrease in the amount of radioactivity in the receiver block at the apical end of a Zea root segment supplied with a basal donor block containing labelled IAA have been investigated.The phenomenon was observed in segments supplied with IAA-1-¹⁴C, IAA-2-¹⁴C and IAA-5-³H. An acropetal polarity in the movement of radioactivity into the receiver blocks was observed using donor blocks containing IAA-5-³H at concentrations as low as 10^-10M.The decrease in the amount of radioactivity in the receiver block begins after 6–8 h of transport at 25° C, and is unaffected by renewal of the donor block every 2 h, or the presence of 2% sucrose in the donor and receiver blocks.The net export of radioactivity into the receiver block at the apical end of the segment virtually ceases after 6–8 h of transport at 25° C, and is not prolonged by the presence of 2% sucrose in the donor and receiver blocks. At 10° C, net export of radioactivity continues for at least the first 50 h of transport, and the amount of radioactivity in a continuously applied receiver block continues to increase over this period.Receiver blocks removed from the apical end of segments after 8 h of transport and placed on planchettes show little or no decrease in the amount of radioactivity they contain as a function of time, in marked contrast to those left in contact with the segment.There is a marked, and metabolically dependent, resorption of radioactivity from the receiver block at the apical end of the segment after about 8 h of transport at 25° C; most of the resorbed radioactivity remains in the apical 2–4 mm of the segment.There is a loss of radioactive CO₂ from segments supplied with a basal donor block containing 10^-6M IAA-1-¹⁴C at 25° C, the emission beginning after 6–8 h of transport. Segments similarly supplied with 10^-6M IAA-2-¹⁴C did not begin to lose radioactive CO₂ until after about 10–12 h of transport.The ability of the segments to transport radioactivity in a polar manner declines with time after they are excised from the root, regardless of whether their cut ends are kept in the intervening period in contact with plain agar blocks, or ones containing unlabelled IAA at 10^-6M. By the 6th h after excision at 25° C no transport of radioactivity through the segments and into the receiver blocks could be detected in either the aropetal or basipetal direction.The decrease in radioactivity in the receiver block after transport periods of 6–8 h at 25° C is therefore due to (1) a cessation of net export of radioactivity into the block, and (2) the onset of a metabolically-dependent, net resorption of radioactivity. At this time substantial amounts of radioactive CO₂ begin to be evolved from segments supplied with IAA-1-¹⁴C, whereas with IAA-2-¹⁴C radioactive CO₂ is not evolved for a further 4–6 h.