Plant Physiology 132:2205-2217 (2003)
© 2003 American Society of Plant Biologists
ENVIRONMENTAL STRESS AND ADAPTATION
Modulation of Citrate Metabolism Alters Aluminum Tolerance in Yeast and Transgenic Canola Overexpressing a Mitochondrial Citrate Synthase1
Valar M. Anoop*,
Urmila Basu,
Mark T. McCammon,
Lee McAlister-Henn and
Gregory J. Taylor
Department of Biological Sciences, University of Alberta, Edmonton,
Alberta, Canada T6G 2E9 (V.M.A., U.B., G.J.T.); and Department of
Biochemistry, University of Texas Health Science Center at San Antonio, San
Antonio, Texas, 78229–3900 (M.T.M., L.M.-H.)
Aluminum (Al) toxicity is a major constraint for crop production in acid
soils, although crop cultivars vary in their tolerance to Al. We have
investigated the potential role of citrate in mediating Al tolerance in
Al-sensitive yeast (Saccharomyces cerevisiae; MMYO11) and canola
(Brassica napus cv Westar). Yeast disruption mutants defective in
genes encoding tricarboxylic acid cycle enzymes, both upstream (citrate
synthase [CS]) and downstream (aconitase [ACO] and isocitrate
dehydrogenase [IDH]) of citrate, showed altered levels of Al
tolerance. A triple mutant of CS ( cit123) showed lower levels
of citrate accumulation and reduced Al tolerance, whereas
aco1- and idh12-deficient mutants showed
higher accumulation of citrate and increased levels of Al tolerance.
Overexpression of a mitochondrial CS (CIT1) in MMYO11 resulted in a
2- to 3-fold increase in citrate levels, and the transformants showed enhanced
Al tolerance. A gene for Arabidopsis mitochondrial CS was overexpressed in
canola using an Agrobacterium tumefaciens-mediated system. Increased
levels of CS gene expression and enhanced CS activity were observed in
transgenic lines compared with the wild type. Root growth experiments revealed
that transgenic lines have enhanced levels of Al tolerance. The transgenic
lines showed enhanced levels of cellular shoot citrate and a 2-fold increase
in citrate exudation when exposed to 150 µM Al. Our work with
yeast and transgenic canola clearly suggest that modulation of different
enzymes involved in citrate synthesis and turnover (malate dehydrogenase,
CS, ACO, and IDH) could be considered as potential targets
of gene manipulation to understand the role of citrate metabolism in mediating
Al tolerance.
1 This work was supported by the Natural Sciences and Engineering Research
Council of Canada, by the Department of Biological Sciences, University of
Alberta (Edmonton, Alberta, Canada), and by the National Institutes of Health
(grant no. AG17477 to L.M.-H.).
*
Corresponding author; e-mail
mv{at}ualberta.ca;
fax 780–492–9234.
Received March 19, 2003;
returned for revision April 21, 2003;
accepted May 12, 2003.
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