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Inhibition of aconitase in citrus fruit callus results in a metabolic shift towards amino acid biosynthesis

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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.

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Abbreviations

GABA:

γ-Aminobutyrate

ICDH:

Isocitrate dehydrogenase

2OG:

2-Oxoglutarate

SSAR:

Succinic semialdehyde reductase

SSAD:

Succinic semialdehyde dehydrogenase

TA:

Total titratable acid

TCA:

Tricarboxylic acid

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Acknowledgments

This work was supported by the Binational Agricultural Research and Development Fund (Grant number US-4010-07) and by Grant number 203-552 of the Chief Scientist of the Ministry of Agriculture and Rural Development. Contribution from the Agricultural Research Organization, The Institute of Plant Sciences, Bet Dagan, Israel, No. 106/2011.

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Authors and Affiliations

  1. Department of Fruit Tree Sciences, ARO, The Volcani Center, P.O. Box 6, Bet Dagan, 50250, Israel

    Asfaw Degu, Bayissa Hatew, Ludmila Shlizerman, Naftali Zur & Avi Sadka

  2. Max Planck Institute of Molecular Plant Physiology, Wissenschaftspark Golm, Am Mühlenberg 1, 14476, Potsdam-Golm, Germany

    Adriano Nunes-Nesi & Alisdair R. Fernie

  3. Department of Plant Sciences, Mail Stop 5, University of California, 1 Shields Ave, Davis, CA, 95616, USA

    Ehud Katz & Eduardo Blumwald

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  1. Asfaw Degu
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  2. Bayissa Hatew
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  5. Naftali Zur
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Correspondence to Avi Sadka.

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Degu, A., Hatew, B., Nunes-Nesi, A. et al. Inhibition of aconitase in citrus fruit callus results in a metabolic shift towards amino acid biosynthesis. Planta 234, 501–513 (2011). https://doi.org/10.1007/s00425-011-1411-2

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