Gibberellin structure and function: biological activity and competitive inhibition of gibberellin 2‐ and 3‐oxidases

Abstract

Some gibberellin (GA) analogues, especially with C-16,17 modifications of GA(5), can inhibit growth of plants apparently by acting as competitors with the endogenous substrate of GA biosynthetic enzymes. Here, we directly confirm the competitive action of GA derivatives but also show that some analogues may retain significant bioactivity. A recombinant 3-oxidase from pea, which converts GA(20) to bioactive GA(1), was inhibited by GA(5), and 16,17-dihydro-GA(5) derivatives, especially if the C-17 alkyl chain length was increased by up to three carbons or if the C-13 hydroxyl was acetylated. Genetic confirmation that GA(5) analogues target 3-oxidases in vivo was provided by comparing the growth response of a WT (LE) pea with a 3-oxidase mutant (le-1). Two pea 2-oxidases that inactivate bioactive GAs, were inhibited by GA(1) and GA(3) but were generally insensitive to GA(5) analogues. alpha-Amylase production by barley half-seeds in response to GA analogues provided a method to study their action when effects on GA biosynthesis were excluded. This bioactivity assay showed that 16,17-dihydro GA(5) analogues have some inherent activity but mostly less than for GA(5) (5-50-fold), which in turn was 100-fold less active than GA(1) and GA(3). However, although C-17 alkyl derivatives with one or two added carbons showed little bioactivity and were purely 3-oxidase inhibitors, adding a third carbon (the 17-n-propyl-16,17-dihydro GA(5) analogue) restored bioactivity to that of GA(5). Furthermore, this analogue has lost its capacity to inhibit stem elongation of Lolium temulentum (Mander et al., Phytochemistry 49:1509-1515, 1998a), although it strongly inhibits the 3-oxidase. Thus, the effectiveness of a GA derivative as a growth retardant will reflect the balance between its bioactivity and its capacity to inhibit the terminal enzyme of GA biosynthesis. The weaker growth inhibition in dicots including pea (approximately 10%) than in monocots such as L. temulentum (>35%) is suggestive of taxonomic differences in the bioactivity of GAs and/or their effects on GA biosynthesis.

16,17-Dihydro Gibberellin A5 Competitively Inhibits a Recombinant Arabidopsis GA 3β-Hydroxylase Encoded by the GA4 Gene

“…The K m values of AtGA3ox1 fusion protein (in a pGEX-2T vector) for the conversions of GA 9 to GA 4 and GA 20 to GA 1 were approximately the same as those obtained by Williams et al (1998) with lysate preparations and also roughly comparable to those seen for other GA3ox1 enzymes expressed in different vectors (Martin et al, 1997;King et al, 2004). However, purified AtGA3ox1 fusion protein was much more , 5 mM ascorbate, 5 mM 2-oxoglutarate, 2 mM NADPH, and 2 mg BSA mL 21 .…”

“…16, (Fig. 1) and other ring D-modified GA 5 derivatives (Mander et al, 1995(Mander et al, , 1998a(Mander et al, , 1998b are structurally very similar to GA 20 .When these ring D-modified GAs are applied to many higher plant species, and especially to grasses, they can effectively inhibit shoot growth (Evans et al, 1994a(Evans et al, , 1994bTakagi et al, 1994;Foster et al, 1997;King et al, 1997King et al, , 2004. Associated with the reduction in shoot elongation is a concomitant reduction in levels of endogenous 3b-hydroxylated GAs and an increase in one or more of the 3-deoxy precursors (Foster et al, 1997;Junttila et al, 1997;Zhou, 2000).…”

“…inhibit conversion of GA 20 to GA 1 by AtGA3ox1). A parallel approach has also been taken by King et al (2004) using cell lysates containing recombinant 3b-and 2b-hydroxylases from Pisum sativum, where their range of ring D-modified GA 5 derivatives included the 17-ethyl, n-propyl, and n-butyl derivatives of dihydro GA 5 . et al (1998) showed that AtGA3ox1, which is encoded by the GA4 gene, was a GA 3b-hydroxylase.…”

Regulation of Flowering in the Long-Day Grass Lolium temulentum by Gibberellins and the FLOWERING LOCUS T Gene

“…Preliminary studies of metabolism of GA 5 in L. temulentum showed its conversion to GA 6 but not to GA 3 (King et al, 2004). Although applied GA 6 is as florally effective as GA 5 and it increases in LD apex along with GA 5 (King et al, 2001, its detection was unreliable in the leaf so no results could be presented.…”

Chemical Regulators of Gibberellin Status and Their Application in Plant Production

“…In order to explain the high biological activity it can be assumed that exo-16,17-dihydro-GA 5 -13-acetate and related structures compete very effectively in grasses with the natural GA substrates, e.g. GA 20 , for the respective enzymatic sites (Takagi et al, 1994;King et al, 2004). In contrast to graminaceous plants, exo-16,17-dihydro-GA 5 -13-acetate and related structures are virtually inactive in reducing shoot growth in any other plant species tested (Rademacher et al, 1999).…”