ALBERT

Notes: Indole-3-butyric acid (IBA) was recently identified by GC/MS analysis as an endogenous constituent of various plants. Plant tissues contained 9 ng g−1 fresh weight of free IBA and 37 ng g−1 fresh weight of total IBA, compared to 26 ng g−1 and 52 ng g−1 fresh weight of free and total indole-3-acetic acid (IAA), respectively. IBA level was found to increase during plant development, but never reached the level of IAA. It is generally assumed that the greater ability of IBA as compared with IAA to promote rooting is due to its relatively higher stability. Indeed, the concentrations of IAA and IBA in autoclaved medium were reduced by 40% and 20%, respectively, compared with filter sterilized controls. In liquid medium, IAA was more sensitive than IBA to non-biological degradation. However, in all plant tissues tested, both auxins were found to be metabolized rapidly and conjugated at the same rate with amino acids or sugar.Studies of auxin transport showed that IAA was transported faster than IBA. The velocities of some of the auxins tested were 7. 5 mm h−1 for IAA, 6. 7 mm h−1 for naphthaleneacetic acid (NAA) and only 3. 2 mm h−1 for IBA. Like IAA, IBA was transported predominantly in a basipetal direction (polar transport). After application of 3H-IBA to cuttings of various plants, most of the label remained in the bases of the cuttings. Easy-to-root cultivars were found to absorb more of the auxin and transport more of it to the leaves.It has been postulated that easy-to-root, as opposed to the difficult-to-root cultivars, have the ability to hydrolyze auxin conjugates at the appropriate time to release free auxin which may promote root initiation. This theory is supported by reports on increased levels of free auxin in the bases of cuttings prior to rooting. The auxin conjugate probably acts as a ‘slow-release’ hormone in the tissues. Easy-to-root cultivars were also able to convert IBA to IAA which accumulated in the cutting bases prior to rooting. IAA conjugates, but not IBA conjugates, were subject to oxidation, and thus deactivation. The efficiency of the two auxins in root induction therefore seems to depend on the stability of their conjugates. The higher rooting promotion of IBA was also ascribed to the fact that its level remained elevated longer than that of IAA, even though IBA was metabolized in the tissue.IAA was converted to IBA by seedlings of corn and Arabidopsis. The Km value for IBA formation was low (approximately 20 μM), indicating high affinity for the substrate. That means that small amounts of IAA (only a fraction of the total IAA in the plant tissues) can be converted to IBA. It was suggested that IBA is formed by the acetylation of IAA with acetyl-CoA in the carboxyl position via a biosynthetic pathway analogous to the primary steps of fatty acid biosynthesis, where acetyl moieties are transferred to an acceptor molecule. Incubation of the soluble enzyme fraction from Arabidopsis with 3H-IBA, IBA and UDP-glucose resulted in a product that was identified tentatively as IBA glucose (IBGIc). IBGIc was detected only during the first 30 min of incubation, showing that it might be converted rapidly to another conjugate.

Notes: Indole-3-butyric acid (IBA) was much more effective than indole-3-acetic acid (IAA) in inducing adventitious root formation in mung bean (Vigna radiata L.) cuttings. Prolonging the duration of treatment with both auxins from 24 to 96 h significantly increased the number of roots formed. Labelled IAA and IBA applied to the basal cut surface of the cuttings were transported acropetally. With both auxins, most radioactivity was detected in the hypocotyl, where roots were formed, but relatively more IBA was found in the upper sections of the cuttings. The rate of metabolism of IAA and IBA in these cuttings was similar. Both auxins were metabolized very rapidly and 24 h after application only a small fraction of the radioactivity corresponded to the free auxins. Hydrolysis with 7 M NaOH indicates that conjugation is the major pathway of IAA and IBA metabolism in mung bean tissues. The major conjugate of IAA was identified tentatively as indole-3-acetylaspartic acid, whereas IBA formed at least two major conjugates. The data indicate that the higher root-promoting activity of IBA was not due to a different transport pattern and/or a different rate of conjugation. It is suggested that the IBA conjugates may be a better source of free auxin than those of IAA and this may explain the higher activity of IBA.

Notes: Auxin conjugates play a role in the regulation of free indole-3-acetic acid (IAA) content in plants. Not much is known about the enzymes involved in either conjugate synthesis or hydrolysis. In this study we have isolated and characterized an auxin conjugate hydrolase from Chinese cabbage seedlings and investigated it during the development of both the Chinese cabbage plants and the clubroot disease. The hydrolase isolated from light- and dark-grown seedlings accepted the amide conjugates indole-3-acetic acid-alanine (IAAla), IAA-phenylalanine (IAPhe), but not IAA-aspartate (IAAsp) as substrates. We also found a substantial amount of hydrolysis of an ester conjugate (IAA-glucose, IAGlu) in our enzyme preparation. The tentative reaction product IAA was identified by HPLC and subsequent GC-MS analysis. The pH optima for the different substrates were not identical, suggesting several hydrolase isoforms. After gel filtration chromatography we found at least two peaks containing different hydrolase isoforms. The isoform, which converted IAGlu to IAA, exhibited a molecular mass of ca 63 kDa, and an isoform of ca 21 kDa converted IAAla and IAPhe. The increased free IAA content in clubroot-diseased roots of Brassicaceae can be due to either de novo synthesis or release of IAA from conjugates. To answer this question free, ester- and amide-bound IAA was measured in 24- and 30-day-old leaves and roots of healthy and Plasmodiophora brassicae-infected Chinese cabbage, and the hydrolase activity with different substrates measured in the same tissues. The amide conjugates were dramatically enhanced in infected roots, whereas free IAA was only slightly enhanced compared to the control tissue. Hydrolase activity was also enhanced in clubbed roots, but the substrate specificity differed from that found in the seedlings. Especially, IAAsp hydrolysis was induced after inoculation with P. brassicae. We conclude that different auxin conjugates can be hydrolyzed at different developmental stages or under stress.

Notes: The translocation and metabolism of ethephon at pH 7.0 and its effect on abscission of olive fruit, were studied in attached and detached fruits. In detached olives, the lowest fruit removal force values were achieved when the fruits were treated at their proximal cavity and kept under humid conditions. Following application of 14C-ethephon to the proximal cavity 63% of the label was absorbed within 4 h; evolution of 14C-ethylene proceeded at a slow rate, mainly from 14C-ethephon remaining on the olive surface, totaling 37% of the applied ethephon in 20 days. 14C-ethephon disappeared rapidly after its application to olive on the tree. More 14C-ethephon could be extracted from olives on the trees after distal application than after proximal application.Since the response of detached olives to treatments with ethephon was similar to that of attached olives in regard to fruit removal force reduction, the former can be used for the study of many aspects of ethephon and ethylene metabolism in olives.In our study no ethephon metabolites other than 14C-ethylene could be detected after 14C-ethephon application to attached or detached olive fruits.

Notes: Changes in levels of free and conjugated indole-acetic acid (IAA) resulting from ethylene treatment were examined in leaves and callus of olive (Olea europea var. Manzanillo) by electron capture gas chromotography and by isotope dilution analysis. In both olive leaves and olive tissue cultures, the level of free IAA did not change much after the exposure to ethylene, but the levels of bound IAA increased almost three times in comparison with untreated controls. This is the first report of changes in endogenous, bound auxin accompanying ethylene treatment.

Notes: Abstract The role of auxins in induction of roots byAgrobacterium rhizogenes was studied in carrot root disks. Transformed roots were produced on root disks by inoculation withA. rhizogenes, A4. Measurement of indole-3-acetic acid (IAA) by gas chromatography-mass spectrometry (GC-MS) indicated that there was a significant increase in the concentration of IAA in transformed callus and induced roots compared with initial IAA concentrations in carrot disks. Indole-3-butyric acid (IBA) was found to occur naturally in carrot roots. The presence of IBA, a potent root inducer, must be taken into account when assessing the role of auxin during transformation and induction of roots byA. rhizogenes.