Gibberellin biosynthesis from gibberellin A(12)-aldehyde in endosperm and embryos of Marah macrocarpus

Soluble enzyme preparations from embryos and endosperm of Marah macrocarpus (previously Echinocystis macrocarpa) were incubated with [C-14(4)]gibberellin(GA)(12)-aldehyde, [C-14(4)]GA(12), [C-14(1)] GA(9), 2,3-didehydro[C-14(1)]GA(9), [C-14(1)]GA(20), and [17-C-13,H-3]GA(5). Embryo preparations converted GA(12)-aldehyde, GA(12), and GA(9) to GA(4) and GA(7); 2,3-didehydroGA(9) to GA(7); GA(5) to GA(3); and GA(20) (incompletely) to GA(1) and GA(60), but not to GA(3). Endosperm preparations converted GA(12)-aldehyde and GA(12) to GA(15), GA(24), GA(25), and GA(9), but, unlike embryo preparations, not to GA(4) or GA(7). However, GA(4) and GA(7) were formed from GA(9) and GA(7) was formed from 2,3-didehydroGA(9). Metabolism of GA(5) to GA(3) and GA(20) to GA(1) was low. 2,3-DidehydroGA(9) accumulated when GA(9) was incubated with a desalted endosperm preparation. A cDNA clone (M3-8), selected from an embryo-derived cDNA library using a DNA fragment generated by reverse transcriptase polymerase chain reaction, was expressed in Escherichia coli. The fusion protein converted GA(12) to GA(9) (major) and GA(25) (minor); GA(53) was metabolized less effectively and only to GA(44). Thus, the M3-8 protein is functionally similar to GA 20-oxidases from Arabidopsis thaliana, Spinacia oleracea, and Pisum sativum, but different from that from Cucurbita maxima seeds, to which its amino acid sequence is most closely related. mRNA hybridizing to M3-8 accumulated in embryos and endosperm of M. macrocarpus, but was absent in vegetative tissues.