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JXB Advance Access published online on September 4, 2007
Journal of Experimental Botany, doi:10.1093/jxb/erm179
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© 2007 The Author(s).
This is an Open Access article distributed under the terms of the Creative Commons Attribution Non-Commercial License (http://creativecommons.org/licenses/by-nc/2.0/uk/) which permits unrestricted non-commercial use, distribution, and reproduction in any medium, provided the original work is properly cited.This paper is available online free of all access charges (see http://jxb.oxfordjournals.org/open_access.html for further details)

RESEARCH PAPER

Integrated metabolomic and transcriptomic analyses of high-tryptophan rice expressing a mutant anthranilate synthase alpha subunit

Joseph G. Dubouzet1, Atsushi Ishihara1,2, Fumio Matsuda1 *, Hisashi Miyagawa1,2, Hiroyoshi Iwata3 and Kyo Wakasa1,4,5,{dagger}

1CREST, Japan Science and Technology Agency, Tokyo 103-0027, Japan
2Division of Applied Life Sciences, Department of Agriculture, Kyoto University, Kyoto 606-8502, Japan
3Department of Information Science and Technology, National Agricultural Research Center, 3-1-1 Kannondai, Tsukuba, Ibaraki 305-8666, Japan
4Department of Rice Breeding, National Institute of Crop Science, 2-1-18 Kannondai, Tsukuba, Ibaraki 305-8518, Japan
5Tokyo University of Agriculture, Faculty of Agriculture, 1737 Funako, Atsugi, Kanagawa 243-0034, Japan

{dagger} To whom correspondence should be addressed at Tokyo University of Agriculture, 1737 Funako, Atsugi, Kanagawa 243-0034, Japan. E-mail: k3wakasa{at}nodai.ac.jp

Transgenic rice plants overexpressing a mutant rice gene for anthranilate synthase alpha subunit (OASA1D) accumulate large amounts of free tryptophan (Trp) with few adverse effects on the phenotype, except for poor germination and weak seedling growth. Metabolic profiling of 8-d-old seedlings of Nipponbare and two high-Trp lines, HW1 and HW5, by high performance liquid chromatography-photo diode array (HPLC-PDA) confirmed that, relative to Nipponbare, only the peak attributed to Trp was significantly changed in the profiles of the OASA1D lines. More detailed and targeted analysis using HPLC coupled with tandem mass spectrometry revealed that the OASA1D lines had higher levels of anthranilate, tryptamine, and serotonin than Nipponbare, but these metabolites were at much lower levels than free Trp. The levels of phenylalanine (Phe) and tyrosine (Tyr) were not affected by the overproduction of Trp. Transcriptomic analysis by microarray validated by quantitative Real-Time PCR (qRT-PCR) revealed that at least 12 out of 21 500 genes showed significant differential expression among genotypes. Except for the OASA1D transgene and a putative IAA ß-glucosyltransferase, these were not related to Trp metabolism. Most importantly, the overexpression of the OASA1D and the consequent accumulation of Trp in these lines had little effect on the overall transcriptome, consistent with the minimal effects on growth and the metabolome. Integrated analysis of the metabolome and transcriptome of these OASA1D transgenic lines indicates that the over-accumulation of free Trp may be partly due to the low activity of Trp decarboxylase or other metabolic genes that directly utilize Trp as a substrate.

Key words: Metabolic profiling, microarray analysis, OASA1D, Oryza sativa, tryptophan

* Present address: Metabolome Research Group, RIKEN Plant Science Center, Yokohama, Kanagawa 230-0045, Japan.

Received 9 March 2007; Revised 18 June 2007 Accepted 26 June 2007


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