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Modulation of higher-plant NAD(H)-dependent glutamate dehydrogenase activity in transgenic tobacco via alteration of beta subunit levels

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An Erratum to this article was published on 08 February 2007

Abstract

Glutamate dehydrogenase (GDH; EC 1.4.1.2–1.4.1.4) catalyses in vitro the reversible amination of 2-oxoglutarate to glutamate. In vascular plants the in vivo direction(s) of the GDH reaction and hence the physiological role(s) of this enzyme remain obscure. A phylogenetic analysis identified two clearly separated groups of higher-plant GDH genes encoding either the α- or β-subunit of the GDH holoenzyme. To help clarify the physiological role(s) of GDH, tobacco (Nicotiana tabacum L.) was transformed with either an antisense or sense copy of a β-subunit gene, and transgenic plants recovered with between 0.5- and 34-times normal leaf GDH activity. This large modulation of GDH activity (shown to be via alteration of β-subunit levels) had little effect on leaf ammonium or the leaf free amino acid pool, except that a large increase in GDH activity was associated with a significant decrease in leaf Asp (~51%, P=0.0045). Similarly, plant growth and development were not affected, suggesting that a large modulation of GDH β-subunit titre does not affect plant viability under the ideal growing conditions employed. Reduction of GDH activity and protein levels in an antisense line was associated with a large increase in transcripts of a β-subunit gene, suggesting that the reduction in β-subunit levels might have been due to translational inhibition. In another experiment designed to detect post-translational up-regulation of GDH activity, GDH over-expressing plants were subjected to prolonged dark-stress. GDH activity increased, but this was found to be due more likely to resistance of the GDH protein to stress-induced proteolysis, rather than to post-translational up-regulation.

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Abbreviations

NH +4 :

Ammonium

GDH:

Glutamate dehydrogenase

GOGAT:

Glutamate synthase

GS:

Glutamine synthetase

2-OG:

2-oxoglutarate

PVP:

Polyvinylpyrrolidone

PVPP:

Polyvinylpolypyrrolidone

UPGMA:

Unweighted Pair Group Mean Average

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Acknowledgements

We thank Andrew Fletcher for help with amino acid analysis, Jo Stringer for assistance with statistical analyses, and Scott Hermann and Agnelo Furtado for critical reading of the manuscript.

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Author notes

  1. Matthew P. Purnell

    Present address: BSES Ltd., 50 Meiers Road, Indooroopilly, QLD, 4068, Australia

Authors and Affiliations

  1. Department of Botany, University of Queensland, Queensland, QLD, 4072, Australia

    Matthew P. Purnell & José R. Botella

  2. Department of Biology, University of Crete, P.O. Box 2208, 71409, Heraklion, Greece

    Damianos S. Skopelitis & Kalliopi A. Roubelakis-Angelakis

Authors
  1. Matthew P. Purnell
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  2. Damianos S. Skopelitis
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  3. Kalliopi A. Roubelakis-Angelakis
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  4. José R. Botella
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Correspondence to José R. Botella.

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An erratum to this article is available at http://dx.doi.org/10.1007/s00425-007-0481-7.

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Purnell, M.P., Skopelitis, D.S., Roubelakis-Angelakis, K.A. et al. Modulation of higher-plant NAD(H)-dependent glutamate dehydrogenase activity in transgenic tobacco via alteration of beta subunit levels. Planta 222, 167–180 (2005). https://doi.org/10.1007/s00425-005-1510-z

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