Gibberellins Inhibit Flavonoid Biosynthesis and Promote Nitrogen Metabolism in Medicago truncatula.

Hao Sun,Mingna Li,Jiaju Zhang,Ruicai Long,Zhen Wang,Fengyan Yi,Junmei Kang,Qingchuan Yang,Huiting Cui

doi:10.3390/ijms22179291

Abstract

Bioactive gibberellic acids (GAs) are diterpenoid plant hormones that are biosynthesized through complex pathways and control various aspects of growth and development. Although GA biosynthesis has been intensively studied, the downstream metabolic pathways regulated by GAs have remained largely unexplored. We investigated Tnt1 retrotransposon insertion mutant lines of Medicago truncatula with a dwarf phenotype by forward and reverse genetics screening and phylogenetic, molecular, biochemical, proteomic and metabolomic analyses. Three Tnt1 retrotransposon insertion mutant lines of the gibberellin 3-beta-dioxygenase 1 gene (GA3ox1) with a dwarf phenotype were identified, in which the synthesis of GAs (GA3 and GA4) was inhibited. Phenotypic analysis revealed that plant height, root and petiole length of ga3ox1 mutants were shorter than those of the wild type (Medicago truncatula ecotype R108). Leaf size was also much smaller in ga3ox1 mutants than that in wild-type R108, which is probably due to cell-size diminution instead of a decrease in cell number. Proteomic and metabolomic analyses of ga3ox1/R108 leaves revealed that in the ga3ox1 mutant, flavonoid isoflavonoid biosynthesis was significantly up-regulated, while nitrogen metabolism was down-regulated. Additionally, we further demonstrated that flavonoid and isoflavonoid biosynthesis was induced by prohexadione calcium, an inhibitor of GA3ox enzyme, and inhibited by exogenous GA3. In contrast, nitrogen metabolism was promoted by exogenous GA3 but inhibited by prohexadione calcium. The results of this study further demonstrated that GAs play critical roles in positively regulating nitrogen metabolism and transport and negatively regulating flavonoid biosynthesis through GA-mediated signaling pathways in leaves.

Highlights

Plant hormones, through endogenous biosynthesis and signal transduction in plants, perform complex physiological functions at very low concentrations
Mtga3ox mutant has been recently characterized in other mutant lines of M. truncatula [26], the mutant lines used in our experiment were identified by forward genetics combined with reverse genetics
In a phylogenetic analysis of MtGA3ox1, based on the entire protein sequences, MtGA3ox1 was clustered with a group of proteins that have been identified as the GA3ox family of proteins (Supplementary Materials Figure S1B), which are responsible for catalyzing the reactions from GA9 to GA4, as well as from GA20 to GA1 (Supplementary Materials Figure S1C) [5]

Summary

Introduction

Through endogenous biosynthesis and signal transduction in plants, perform complex physiological functions at very low concentrations. Plant hormones play important roles in plant developmental and physiological processes, such as morphogenesis, growth, metabolism, etc. Gibberellic acids (GAs), a diterpenoid plant hormone, play an important role in various processes in the whole life cycle of plants, including seed germination, hypocotyl and stem elongation, leaf expansion, trichome development, flowering time, flower development and fruit development [2,3]. Ent-kaurene oxidase (KO) catalyzes the formation of ent-kaurenoic acid, which occurs mainly in the plastid membrane. In the third and last stage, which occurs in the nucleus and cytoplasm [8], bioactive GAs (including GA1, GA3, GA4 and GA7) are synthesized from the precursor GA12 and catalyzed by the oxidases GA20ox and GA3ox [3]. As a key enzyme in GA synthesis, GA3ox is responsible for catalyzing GA9/GA20 to generate GA4/GA1

Methods

Results

Conclusion