Auxin Transport in Roots: V. EFFECTS OF TEMPERATURE ON THE MOVEMENT OF IAA IN ZEA ROOTS

The effects of temperature on the polar movement of IAA through6-mm and 12-mm segments of Zea mays roots have been investigatedover the range from 1 to 50°C. At all temperatures an acropetal polar movement of IAA predominated,although at low temperatures and at 50°C the 6-mm segmentsshowed a transient basipetal polarity, before the persistentacropetal polarity developed. At 1°C the differences betweenacropetal and basipetal movement of IAA were less distinct thanat the other temperatures. There is, however, a marked metabolically-dependentacropetal movement of IAA through the tissues at 1°C, becausewhen the segments were deprived of oxygen the acropetal movementwas severely reduced while the basipetal movement was reducedto a smaller extent. At 1°C and at 5°C there was alwaysa persistent basipetal polarity of IAA movement through 6-mmand 12-mm segments under anaerobic conditions. The velocity of acropetal movement (mm h^–1) was the samethrough the 6-mm and the 12-mm segments and was markedly affectedby temperature. It increased from 1°C to a maximum valueof 8 mm h^–1 at 31°C and then decreased again at 40and 50°C. The velocity of basipetal movement could be assessedonly at 1 and 5°C at which temperatures it was greater thanthe velocity of acropetal movement, and virtually independentof segment length. The acropetal flux of IAA (cpm h^–1) was much less through12-mm segments than through 6-mm segments. For both lengthsof segment, however, the flux showed a complex relationshipwith ambient temperature, increasing from 1°C to a maximumat 10–15°C, declining to a minimum value at 31°Cand then rising again at 40 and 50°C. The basipetal fluxof IAA could be astimated only at 1 and 5°C at which itwas very much smaller than the acropetal flux. The amount of IAA in the receiver blocks increased linearlywith time at the lower temperatures. At temperatures withinthe range 15°C to about 31°C, however, the amount ofIAA in the receiver blocks began to decline if the transportperiods exceeded a certain length. The time at which this declinein the IAA in the receiver block began was related to the ambienttemperature. Chromatographic analysis indicated one radioactive substancein receiver blocks at the apical end of segments supplied withIAA-1-¹⁴C at the basal end after transport periods of 6 h at25°C, and 72 h at 5°C. The Rf of this substance wasclosely similar to that of the radioactive IAA supplied in thedonor blocks.     

Evidence for Three Different Systems of Movement of Indoleacetic Acid in Intact Roots of Phaseolus coccineus

Indoleacetic acid (IAA)-5-³H (2 × 10⁻⁹M) was applied to intact roots of Phaseolus coccineus seedlings, at the apex or 2 cm above the apex, at various pHs and in the presence of Cu²⁺ and NaCl. The transport of label in the roots was then examined after 6 h by cutting the roots into 1 mm sections above and below the zone of treatment. Basipetal movement from 2 cm above the apex was unafected by pH, Cu²⁺ or NaCl. Acropetal movement from the same area decreased with increasing pH from 5.4 to 8.0, probably due to an effect of pH on the entry of IAA into the cells. pH had no effect on sucrose transport. Cu²⁺ also inhibited acropetal movement but NaCl had no effect. Basipetal movement of label from the apex was reduced by Cu²⁺ and increasing pH, but not as much as with acropetal movement, and increased by the presence of NaCl. These facts are interpreted as showing 3 different systems of IAA movement in intact roots: basipetal from 2 cm up the root in some extracellular physical system; acropetal from 2 cm up the root, and basipetal from the apex, in a metabolically dependent intracellular system, but in different tissues of the root. It is proposed that endogenous IAA not only moves into the root from the stem but is also synthesized in the root apex, and moves basipetally for a short distance to the root growing zone in a separate system from the IAA descending from the stem.     

THE REGULATION OF ABSCISSION AND IAA BY SENESCENCE FACTOR AND ABSCISIC ACID

The effects of the endogenous “senescence factor” (SF) and synthetic (±)-abscisic acid (ABA) on basipetal transport of indoleacetic acid (IAA) in Coleus blumei Benth. were studied. When used to pretreat explant petioles, both SF and ABA accelerated abscission and decreased basipetal movement of IAA through petiolar sections but had no effect on acropetal movement, which was negligible in all experiments. SF decreased the intensity but not the velocity of the basipetal movement of IAA. Pretreatment with either SF or ABA decreased the free IAA and increased the IAA-aspartate extractable from the sections. The results support the hypothesis that SF and ABA hasten abscission by lowering the amounts of IAA transport in the tissue. Known properties of SF are similar to those of ABA. Efforts are under way to elucidate the chemical identity of SF.     

Transport of indoleacetic acid in intact roots of Phaseolus coccineus

Summary Indoleacetic acid (IAA)-5-³H (2×10^-9) was applied to intact roots of Phaseolus coccineus seedlings at the apex or 2 cm above the apex, and the movement of IAA-³H and its metabolites traced by sectioning and chromatography. Basipetal movement of label occurred for 2 cm or less, declining exponentially, and the amount increased with time. Acropetal transport from above the apex showed quantitatively less movement of radioactivity. After a 6h treatment period a decline of label occurred in the first 0.5cm, below which there was a long distance movement of small amounts of label, mainly in IAA, towards the apex where the label concentrated by a factor of approximately 2. Short-distance basipetal movement consisted of about equal amounts of IAA and metabolites, and only metabolites were found in areas more basipetal than 2cm. Label from solutions of sucrose-¹⁴C and ³H₂O followed the same general pattern of movement as label from IAA-³H, except that acropetal movement of water showed a steady decrease in the amount of label as the distance from the area of application increased. The short distance basipetal transport of label with the breakdown of IAA-³H indicates that the extent of basipetal movement was limited by catabolic processes. The acropetal pattern of IAA-³H movement with the concentration of the transported material close to the apex, is possibly the result of transport in the phloem.     

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     

Effect of Inversion on Growth and Movement of Indole-3-acetic Acid in Coleoptiles

The effect of a 180° displacement from the normal vertical orientation on longitudinal growth and on the acropetal and basipetal movement of ¹⁴C-IAA was investigated in Avena sativa L. and Zea mays L. coleoptile sections. Inversion inhibits growth in intact sections (apex not removed) and in decapitated sections supplied apically with donor blocks containing auxin. Under aerobic conditions, inversion inhibits basipetal auxin movement and promotes acropetal auxin movement, whereas under anaerobic conditions, it does not influence the movement of auxin in either direction. Inversion retards the basipetal movement of the peak of a 30-minute pulse of auxin in corn.

The inversion-induced inhibition of basipetal auxin movement is not explained by an effect of gravity on production, uptake, destruction, exit from sections, retention in tissue, or purely physical movement of auxin. It is concluded that inversion (a) inhibits basipetal transport, the component of auxin movement that is metabolically dependent, and as a result (b) inhibits growth and (c) promotes acropetal auxin movement.

     

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.     

Abscisic Acid Action on Basipetal Auxin Transport

Several experiments have been performed to analyse the ABA effects on the basipetal transport of IAA-2-¹⁴C, using sections of epicotyls prepared from etiolated Lens seedlings. The sections were incubated in an ABA solution or ABA was applied in the donor blocks containing IAA. For each type of assay, the uptake (analyses of the donor blocks) and the movement of IAA-C¹⁴ (analyses of the receiver blocks) were inhibited by ABA. The distribution of continuous decrease of the radioactivity, along the sections' axis, showed a ¹⁴C level from the apical towards the basal segments. ABA caused a decrease in the ¹⁴C concentration for the total sections, but a relative increase for the basal segment. When ABA was applied simultaneously with IAA in the donor blocks, the transport velocity of IAA, through the sections, was not changed significantly, while an ABA pretreatment caused a significant decrease.     

Effect of ethylene treatment on transport and metabolism of indole-3-butyric acid in citrus leaf midribs

Transport and metabolism of radiolabeled indole-3-butyric acid (IBA) were studied in midrib sections of Cleopatra mandarin (Citrus reticulata Blanco) and compared to that of indole-3-acetic acid (IAA). Exogenous IBA was metabolized by the midribs to a polar compound, probably an ester conjugate. Ethylene pretreatment of the midribs reduced their capacity to metabolize IBA by ca. 70% as compared to air pretreatment. IBA transport capacity in the leaf midribs was ca. two times greater in the basipetal direction than the acropetal. The basipetal transport capacity of ³H-IBA was lower than that of ¹⁴C-IAA (ca. 24% and 39% of the uptake, respectively). While ethylene treatment reduced basipetal transport of IAA by ca. 70% it did not affect the transport of IBA. Most of the transported label was found as free IBA, but the reduction of IBA conjugation by ethylene treatment did not affect the transport capacity.     

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.     

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.     

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; )     

Polar Movement of Indole-3-acetic Acid-C in Roots of Lens and Phaseolus

A critical review of the few papers on IAA-¹⁴C movement in roots revealed apparent contradictions, as well as flaws in experimental design that would be apt to cause artifacts. The movement of ¹⁴C from IAA-¹⁴C was studied in sections of Lens and Phaseolus roots, using a system 20 or more times as sensitive as any previously used. To make sure that our results with roots could be compared validly with published work on petioles and stems, we used the same techniques as we had earlier used for shoot structures. The results with Lens were similar in many ways to those for shoots: net movement into receiver blocks was very strongly polar, followed a linear course for several hours, and showed a velocity of the same order of magnitude as in shoots (and, in fact, very close in absolute value to that found in Coleus stem cylinders). Also, as with shoots, all the radioactivity in receiver blocks ran to the R_F of IAA. The time-course of loss of counts from donor blocks was similar to that found in shoots. The 2 most striking differences from shoots were 1) the very low percentage of added ¹⁴C that was moved into the receivers (about one-tenth of the values for bean petioles), and 2) the fact that the polar movement was acropetal in roots, rather than basipetal as in shoots. Results with Phaseolus roots were similar to those for Lens, although an additional complication with Phaseolus roots was the indication of a transitory stage of weak basipetal polarity in the first few hours after excising the section. This stage was followed in a few hours by a stronger acropetal polarity.     

Auxin transport and the regulation of petiole abscission in cotton (Gossypium hirsutum L.): A comparison of indol-3yl-acetic acid and phenylacetic acid

Distal applications of indol-3yl-acetic acid (IAA) to debladed cotyledonary petioles of cotton (Gossypium hirsutum L.) seedlings greatly delayed petiole abscission, but similar applications of phenylacetic acid (PAA) slightly accelerated abscission compared with untreated controls. Both compounds prevented abscission for at least 91 h when applied directly to the abscission zone at the base of the petiole. The contrasting effects of distal IAA and PAA on abscission were correlated with their polar transport behaviour-[1-¹⁴C]IAA underwent typical polar (basipetal) transport through isolated 30 mm petiole segments, but only a weak diffusive movement of [1-¹⁴C]PAA occurred.Removal of the shoot tip substantially delayed abscission of subtending debladed cotyledonary petioles. The promotive effect of the shoot tip on petiole abscission could be replaced in decapitated shoots by applications of either IAA or PAA to the cut surface of the stem. Following the application of [1-¹⁴C]IAA or [1-¹⁴C]PAA to the cut surface of decapitated shoots, only IAA was transported basipetally through the stem. Proximal applications of either compound stimulated the acropetal transport of [¹⁴C]sucrose applied to a subtending intact cotyledonary leaf and caused label to accumulate at the shoot tip. However, PAA was considerably less active than IAA in this response.It is concluded that whilst the inhibition of petiole abscission by distal auxin is mediated by effects of auxin in cells of the abscission zone itself, the promotion of abscission by the shoot tip (or by proximal exogenous auxin) is a remote effect which does not require basipetal auxin transport to the abscission zone. Possible mechanisms to explain this indirect effect of proximal auxin on abscission are 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.     

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.     

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.     

Effects of Flavonoids on the Polar Transport of Auxins

The effect of some flavonoids on the polar transport of auxins was investigated in hypocotyl sections of dark grown seedlings of cucumber, Cucumis sativus L., by means of ¹⁴C labelled auxins. In experiments of 4–6 h duration quercitrin, morin, dihydroquercetin, naringin, sulfuretin and ferulic acid increased the polarity of the transport of indol-3yl-acetic acid (stimulation of basipetal, inhibition of acropetal transport). Naringenin, genistein and pinobanksin, on the other hand, decreased the polarity of this transport. For NAA no increase in the polarity of the transport could be observed, but all the substances tested inhibited the basipetal transport. There was no simple correlation between the effects on the polar transport and the effects on IAA oxidase.     

Polarity of Indoleacetic Acid in young Coleus Stems

Young internodes of Coleus blumei Benth. have long been known for their sizable amount of acropetal indoleacetic acid movement. However, plants of the same clone, under improved growing conditions, now show almost absolute basipetal polarity of ¹⁴C-indoleacetic acid, as measured by liquid scintillation counting of ¹⁴C in the receiver cylinders of agar. The ratio of basipetal to acropetal movement is now as much as 85:1, instead of the 3:1 ratio found years ago under conditions providing slower growth.     

The transport of radiolabeled indoleacetic acid and its conjugates in nodal stem segments of Phaseolus vulgaris L.

The transport of radiolabeled indoleacetic acid (IAA), and some of its conjugates, was investigated in nodal stem segments of Phaseolus vulgaris L. Donor agar blocks containing either [2-acetyl-¹⁴C]-IAA; [2-acetyl-¹⁴C]-indole-3-acetyl-L-aspartate (IAAsp); [2-acetyl-¹⁴C]-indole-3-acetyl-L-glycine (IAGly); or [2-acetyl-¹⁴C]-indole-3-acetyl-L-alanine (IAAla) were placed on either the apical or basal cut surface of stem segments each bearing an axillary bud at the midline. In some experiments, a receiver block was placed on the end opposite to the donor. After transport was terminated, the segments were divided into five equal sections plus the bud, and the radioactivity of donors, receivers and each part of the stem segment was counted.For all four substances tested, the amount of ¹⁴C transported to the axillary bud from the base was the same or greater than that from the apical end. After basipetal transport, the distribution of ¹⁴C in the segment declined sharply from apex to base. The inverse was true for acropetal transport. Transport for the three IAA conjugates did not differ substantially from each other.The IAA transport inhibitor, N-1-naphthylphthalamic acid (NPA), inhibited basipetal ¹⁴C-IAA transport to the base of the stem segment but did not alter substantially the amount of ¹⁴C-IAA recovered from the bud. Transport of ¹⁴C-IAA from the apical end to all parts of the stem segment declined when the base of the section was treated with nonradioactive IAA. Taken together with data presented in the accompanying article [Tamas et al. (1989) Plant Growth Regul 8: 165–183], these results suggest that the transport of IAA plays a role in axillary bud growth regulation, but its effect does not depend on the accumulation of IAA in the axillary bud itself.