Gibberellin biosynthesis from gibberellin A12-aldehyde in endosperm and embryos of Marah macrocarpus.
Soluble enzyme preparations from embryos and endosperm of Marah macrocarpus (previously Echinocystis macrocarpa) were incubated with [14C4]gibberellin(GA)12-aldehyde, [14C4]GA12, [14C1] GA9, 2,3-didehydro[14C1]GA9, [14C1]GA20, and [17-13C, 3H]GA5. Embryo preparations converted GA12-aldehyde, GA12, and GA9 to GA4 and GA7; 2,3-didehydroGA9 to GA7; GA5 to GA3; and GA20 (incompletely) to GA1 and GA60, but not to GA3. Endosperm preparations converted GA12-aldehyde and GA12 to GA15, GA24, and GA9, but, unlike embryo preparations, not to GA4 or GA7. However, GA4 and GA7 were formed from GA9 and GA7 was formed from 2,3-didehydroGA9. Metabolism of GA5 to GA3 and GA20 to GA1 was low. 2,3-DidehydroGA9 accumulated when GA9 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 GA12 to GA9 (major) and GA25 (minor); GA53 was metabolized less effectively and only to GA44. 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.
MacMillan
J
J
Department of Agricultural Sciences, University of Bristol, Long Ashton, Bristol, United Kingdom.
Ward
D A
DA
Phillips
A L
AL
Sánchez-Beltrán
M J
MJ
Gaskin
P
P
Lange
T
T
Hedden
P
P
D000595:Amino Acid Sequence; D003001:Cloning, Molecular; D018076:DNA, Complementary; D004926:Escherichia coli; D015723:Gene Library; D005875:Gibberellins / Q000378:metabolism*; D006899:Mixed Function Oxygenases / Q000096:biosynthesis / Q000737:chemistry / Q000378:metabolism*; D008969:Molecular Sequence Data; D010944:Plants / Q000378:metabolism*; D016133:Polymerase Chain Reaction; D012333:RNA, Messenger / Q000378:metabolism; D011993:Recombinant Fusion Proteins / Q000096:biosynthesis / Q000737:chemistry / Q000378:metabolism; D012639:Seeds / Q000378:metabolism; D017386:Sequence Homology, Amino Acid; D013379:Substrate Specificity
D018076:DNA, Complementary; D005875:Gibberellins; D012333:RNA, Messenger; D011993:Recombinant Fusion Proteins; C048463:gibberellin A12 aldehyde; D006899:Mixed Function Oxygenases; C089210:gibberellin, 2-oxoglutarate-oxygen oxidoreductase (20-hydroxylating, oxidizing)