Metabolism and Synthesis of Indole-3-Acetic Acid (IAA) in Zea mays (Levels of IAA during Kernel Development and the Use of in Vitro Endosperm Systems for Studying IAA Biosynthesis)

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Abstract
Kernels of Zea mays on an intact plant accumulate indole-3-acetic acid (IAA) at the rate of 190 ng g-1 fresh weight h-1. Of the IAA synthesized, 97% is in the esterified form and less than 3% remains as the free acid. The site of biosynthesis of the IAA, whether synthesized in the leaf and transported to the kernel, or in the kernel and remaining in the kernel, has not been established. In an attempt to determine the locus of synthesis, we grew isolated kernels on agar media not containing tryptophan or other possible aromatic precursors of IAA and observed IAA synthesis of 99 ng g-1 fresh weight h-1, approximately 52% of the in situ rate. Thus, the kernel contains all of the enzymes required for de novo aromatic biosynthesis of IAA and its ester conjugates. Furthermore, endosperm cells in suspension culture, grown on hormone-free media and in the absence of aromatic precursors, are able to synthesize IAA at a rate of 9.2 ng g-1 fresh weight h-1, or 4.8% of the in situ rate. This finding establishes that all of the enzymes of IAA biosynthesis occur in the endosperm and that the endosperm is a site of IAA biosynthesis. Isolated endosperm, prepared from developing kernels, synthesized IAA from labeled anthranilate at a rate of 8.6 ng g-1 fresh weight h-1, or 4.5% of the in situ rate. Frozen endosperm preparations maintained the ability to synthesize labeled IAA from labeled anthranilate. The identity of the synthesized IAA was established by mass spectral analysis. We suggest that endosperm preparations of Z. mays are suitable for study of the mechanism(s) of IAA biosynthesis because they (a) have high rates of synthesis; (b) show stability to freezing, enabling enzyme storage; (c) provide a system with a known rate of in situ synthesis; and (d) are available in large amounts for use as an enzyme source.