title: Inhibition of aconitase in citrus fruit callus results in a metabolic shift towards amino acid biosynthesis.
abstract: Citrate, a major determinant of citrus fruit quality, accumulates early in fruit development and declines towards maturation. The isomerization of citrate to isocitrate, catalyzed by aconitase is a key step in acid metabolism. Inhibition of mitochondrial aconitase activity early in fruit development contributes to acid accumulation, whereas increased cytosolic activity of aconitase causes citrate decline. It was previously hypothesized that the block in mitochondrial aconitase activity, inducing acid accumulation, is caused by citramalate. Here, we investigated the effect of citramalate and of another aconitase inhibitor, oxalomalate, on aconitase activity and regulation in callus originated from juice sacs. These compounds significantly increased citrate content and reduced the enzyme's activity, while slightly inducing its protein level. Citramalate inhibited the mitochondrial, but not cytosolic form of the enzyme. Its external application to mandarin fruits resulted in inhibition of aconitase activity, with a transient increase in fruit acidity detected a few weeks later. The endogenous level of citramalate was analyzed in five citrus varieties: its pattern of accumulation challenged the notion of its action as an endogenous inhibitor of mitochondrial aconitase. Metabolite profiling of oxalomalate-treated cells showed significant increases in a few amino acids and organic acids. The activities of alanine transaminase, aspartate transaminase and aspartate kinase, as well as these of two γ-aminobutyrate (GABA)-shunt enzymes, succinic semialdehyde reductase (SSAR) and succinic semialdehyde dehydrogenase (SSAD) were significantly induced in oxalomalate-treated cells. It is suggested that the increase in citrate, caused by aconitase inhibition, induces amino acid synthesis and the GABA shunt, in accordance with the suggested fate of citrate during the acid decline stage in citrus fruit.
authors.affiliation: Department of Fruit Tree Sciences, ARO, The Volcani Center, P.O. Box 6, Bet Dagan 50250, Israel.
authors.forename: Asfaw
authors.initials: A
authors.lastname: Degu
authors.forename: Bayissa
authors.initials: B
authors.lastname: Hatew
authors.forename: Adriano
authors.initials: A
authors.lastname: Nunes-Nesi
authors.forename: Ludmila
authors.initials: L
authors.lastname: Shlizerman
authors.forename: Naftali
authors.initials: N
authors.lastname: Zur
authors.forename: Ehud
authors.initials: E
authors.lastname: Katz
authors.forename: Alisdair R
authors.initials: AR
authors.lastname: Fernie
authors.forename: Eduardo
authors.initials: E
authors.lastname: Blumwald
authors.forename: Avi
authors.initials: A
authors.lastname: Sadka
mesh_terms: D000154:Aconitate Hydratase / Q000037:antagonists & inhibitors* / Q000378:metabolism; D000595:Amino Acid Sequence; D000596:Amino Acids / Q000096:biosynthesis* / Q000378:metabolism; D019343:Citric Acid / Q000378:metabolism; D002957:Citrus / Q000187:drug effects / Q000201:enzymology / Q000235:genetics / Q000378:metabolism*; D004791:Enzyme Inhibitors / Q000378:metabolism; D018506:Gene Expression Regulation, Plant; D007557:Israel; D008293:Malates / Q000378:metabolism / Q000494:pharmacology; D008969:Molecular Sequence Data; D010070:Oxalates / Q000378:metabolism / Q000494:pharmacology
chemical_list: D000596:Amino Acids; D004791:Enzyme Inhibitors; D008293:Malates; D010070:Oxalates; D019343:Citric Acid; C015407:oxalomalic acid; D000154:Aconitate Hydratase; C011729:citramalate