Publications Publications

J. Normanly "Auxin Metabolism" Physiologia Plantarum (1997) Vol 100:431-442

Auxin metabolism encompasses transport, conjugation, deconjugation, conversion, and catabolism. The balance between auxin metabolism and biosynthesis determines the actual level of the hormone in a given cell and consequently plays an important role in many developmental processes from seed germination to fruit ripening. Mass spectrometry used in conjunction with stable isotope labeling studies has enabled comprehensive examination of auxin biosynthesis and turnover along witht the identification of many auxin conjugates. It appears that the conjugate moiety may signal the metabolic fate (e.g. storage and eventual hydrolysis to free hormone, or catabolism). Recently identified auxin-metabolizing enzymes are encoded by gene families which vary in specificity for auxin metabolites. The expression patterns of these genes will reveal a great deal about the mechanics of auxin metabolism.

N. Ilic, J. Normanly, and J. D. Cohen "Quantification of free plus conjugated indole-3-acetic acid in Arabidopsis requires correction for the non-enzymatic conversion of indolic nitriles" Plant Phys. (1996) Vol 111:781-788

The genetic advantages to the use of Arabidopsis mutants for the study of auxin metabolism have previously been partially offset by the complexity of indolic metabolism in this plant and the lack of proper methods. To address some of these problems, we developed isotopic labeling methods to determine amounts and examine the metabolism of indolic compounds in Arabidopsis. Isolation and identification of endogenous indoleacetonitrile (IAN) [a possible precursor of the auxin indoleacetic acid (IAA)] was carried out under mild conditions, thus proving its natural occurrence. We describe here the synthesis of [13C]labeled IAN and its utility in the GC-MS quantification of endogenous IAN levels. We also quantified the non enzymatic conversion of IAN to IAA under conditions used to hydrolyze IAA conjugates. [13C]Labeled IAN was employed to asses the contribution of IAN to measured IAA following hydrolysis of IAA conjugates. We studied the stability and breakdown of the indolic glucosinolate glucobrassicin, which is known to be present in Arabidopsis. This is potentially an important concern when using Arabidopsis for studies of indolic chemistry, since levels of indolic auxins and auxin precursors are well below the levels of the indolic glucosinolates. We found that under conditions of extraction and base hydrolysis, formation of IAA from glucobrassicin was negligible.

J. Normanly, J. P. Slovin, and J. D. Cohen "Update on hormones: Rethinking auxin biosynthesis and metabolism" Plant Phys.,107:323-329 (1995)

Review article
This review focuses on the new and more complex picture of IAA biosynthesis that has emerged as a result of recent experiments.

J. Normanly, J. D. Cohen, and G. R. Fink "Arabidopsis thaliana auxotrophs reveal a tryptophan-independent biosynthetic pathway for indole-3-acetic acid" Proc. Natl. Acad. Sci., 90:10355-10359 (1993)

We used tryptophan auxotrophs of the dicot Arabidopsis thaliana (wall cress) to determine whether tryptophan has the capacity to serve as a precursor to the auxin, indole-3-acetic acid (IAA). Quantitative gas chromatography-selected ion monitoring-mass spectrometry (GC-SIM-MS) revealed that the trp2-1 mutant, which is defective in the conversion of indole to tryptophan, accumulated amide- and ester-linked IAA at levels 38-fold and 19-fold, respectively, above those of wild type. Tryptophan and free IAA were isolated from the trp2-1 mutant grown in the presence of [15N]anthranilate and [2H5]tryptophan, and the relative 15N and 2H5 enrichments of tryptophan and IAA were determined via GC-SIM-MS. The 15N enrichment of tryptophan, 13%+/- 4%, was less than the 15N enrichment of the IAA pool, 39% +/- 4%; therefore, IAA biosynthesis occurs via a tryptophan-independent pathway. The amount of 2H5 incorporated by the plant into IAA from tryptophan (95+/- 4%) was low and only slightly above the level of spontaneous, nonenzymatic conversion of [2H5]tryptophan to [2H5]IAA. These results show that the dicot Arabidopsis is similar to the monocot Zea mays in that the major route of IAA biosynthesis does not occur through tryptophan.

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