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JXB Advance Access originally published online on August 13, 2004
Journal of Experimental Botany 2004 55(405):2005-2014; doi:10.1093/jxb/erh224
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Journal of Experimental Botany, Vol. 55, No. 405, © Society for Experimental Biology 2004; all rights reserved

RESEARCH PAPER

Quantitative trait loci analysis of nitrate storage in Arabidopsis leading to an investigation of the contribution of the anion channel gene, AtCLC-c, to variation in nitrate levels

Hisatomi Harada1,*, Takashi Kuromori2, Takashi Hirayama2, Kazuo Shinozaki2 and Roger A. Leigh3

1National Institute of Livestock and Grassland Science, 768, Senbonmatsu, Nasu, Tochigi, 329-2793, Japan
2RIKEN Genomic Sciences Center, 1-7-22 Suehiro-cho, Tsurumi-Ku, Yokohama 230-0045, Japan
3Department of Plant Sciences, University of Cambridge, Downing Street, Cambridge CB2 3EA, UK

* Present address and to whom correspondence should be sent: Ogata Experimental Farm, Akita Agricultural Experiment Station, 1-1, Higashi, Ogata, Minami-Akita, Akita, 010-0442, Japan. Fax: +81 185 22 4008. E-mail: hisatomi{at}affrc.go.jp

Storage of excess nitrate in the vacuole and its subsequent remobilization is an important aspect of a plant's nitrogen economy, but the genes controlling the underlying processes have not all been identified and characterized. Cape Verdi Island (Cvi)/Landsberg erecta (Ler) and Columbia (Col)/Landsberg erecta recombinant inbred line (RIL) populations of Arabidopsis thaliana were used to identify quantitative trait loci (QTL) controlling natural variation in nitrate concentrations. One major and two minor QTLs were found for the Cvi/Ler population and one minor QTL for the Col/Ler RIL. These were designated NA1 to NA4. The major Cvi/Ler QTL (NA3) was located at the bottom of chromosome 5. No interaction among the QTLs was found by two-way ANOVA. By comparing in silico the locations of the QTLs with a physical map of the Arabidopsis genome, candidate genes for each QTL were identified. Several of these were anion channels of the AtCLC family. One of these, AtCLC-c, coincided with NA3 and its role was investigated using a mutant with a transposon insertion in AtCLC-c. Mutant plants homozygous for the insertion (designated clcc-1) had less than 5% of AtCLC-c mRNA compared with wild-type (WT) shoots. They also had significantly lower nitrate concentrations when grown at a range of external nitrate concentrations. The concentrations of chloride, malate, and citrate were also affected in the mutant. In wild-type plants, expression of AtCLC-c was down-regulated in the presence of nitrate, but ammonium had a much smaller effect while chloride and sulphate did not affect expression. These and published results suggest that multiple genes affect nitrate concentrations in plants and that AtCLC-c and other members of the AtCLC gene family play some role in this.

Key words: Anion channel, Arabidospsis, AtCLC-c, choride, nitrate accumulation, QTL


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