Metabolism of Gibberellin A12-7-Aldehyde by Soybean Cotyledons and Its Use in Identifying Gibberellin A7 as an Endogenous Gibberellin

Abstract

The level of gibberellin(GA)-like material in cotyledons of soybean (Glycine max L.) was highest at mid-pod fill-about 10 nanograms GA(3) equivalents per gram fresh weight of tissue, assayed in the immersion dwarf rice bioassay. This amount is about 1000-fold less than levels in Pisum and Phaseolus seed, other legume species whose spectrum of endogenous gibberellins (GAs) is well known. The metabolism of [(14)C]-GA(12)-7-aldehyde (GA(12)ald)-the universal GA precursor-by intact, mid-pod-fill, soybean cotyledons and their cell-free extracts was investigated. In 4 hours, extracts converted GA(12)ald to two products-[(14)C]GA(12) (42% yield) and [(14)C]GA(15) (7%). Within 5 minutes, intact embryos converted GA(12)ald to [(14)C]GA(12) and [(14)C]GA(15) in 15% yield; 4 hour incubations afforded at least 22 products (96% total yield). The putative [(14)C]GA(12) was identified as a product of [(14)C]GA(12)ald metabolism on the basis of co-chromatography with authentic GA(12) on a series of reversed and normal phase high pressure liquid chromatography (HPLC) and thin-layer chromatography (TLC) systems, and by a dual feed of the putative [(14)C]GA(12) and authentic [(14)C]GA(12) to cotyledons of both peas and soybeans. The [(14)C]GA(15) was identified as a metabolite of [(14)C]GA(12)ald by capillary gas chromatography (GC)-mass-spectrometry-selected ion monitoring, GC-radiocounting, HPLC, and TLC. By adding the [(14)C] metabolites of [(14)C]GA(12)ald to a different and larger extract (about 0.2 kg fresh weight of soybean reproductive tissue) and purifying endogenous substances co-chromatographing with these metabolites, at least two GA-like substances were obtained and one identified as GA(7) by GC-mass spectrometry. Since [(14)C]GA(9) was not found as a [(14)C]metabolite of [(14)C]GA(12)ald, soybean embryos might have a pathway for biosynthesis of active, C-19 gibberellins like that of the cucurbits; GA(12)ald --> GA(12) --> GA(15) --> GA(24) --> GA(36) --> GA(4) --> GA(7).