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J. Biol. Chem., Vol. 283, Issue 19, 13407-13417, May 9, 2008
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111
2
From the
Laboratory of Plant Biotechnology and Laboratory of Plant Molecular Physiology, Graduate School of Agricultural and Life Sciences, the University of Tokyo, 1-1 Yayoi, Bunkyo-ku, Tokyo 113-8657 and ¶Plant Genetic Engineering Research Unit, Division of Plant Sciences, National Institute of Agrobiological Sciences, 2-1-2, Kannondai, Tsukuba, Ibaraki 305-8602, Japan
Iron is essential for most living organisms, and thus iron deficiency poses a major abiotic stress in crop production. Plants induce iron utilization systems under conditions of low iron availability, but the molecular mechanisms of gene regulation under iron deficiency remain largely unknown. We identified a novel transcription factor of rice and barley, IDEF2, which specifically binds to the iron deficiency-responsive cis-acting element 2 (IDE2) by yeast one-hybrid screening. IDEF2 belongs to an uncharacterized branch of the NAC transcription factor family and exhibits novel properties of sequence recognition. An electrophoretic mobility shift assay and cyclic amplification and selection of targets experiment revealed that IDEF2 predominantly recognized CA(A/C)G(T/C)(T/C/A)(T/C/A) within IDE2 as the core-binding site. IDEF2 transcripts are constitutively present in rice roots and leaves. Repression of the function of IDEF2 by the RNA interference (RNAi) technique and chimeric repressor gene-silencing technology (CRES-T) caused aberrant iron homeostasis in rice. Several genes up-regulated by iron deficiency, including the Fe(II)-nicotianamine transporter gene OsYSL2, were less induced by iron deficiency in the RNAi rice of IDEF2, suggesting that IDEF2 is involved in the regulation of these genes. Many genes with repressed expression in IDEF2 RNAi rice possessed the IDEF2-binding core sites in their promoters, and the flanking sequences were also highly homologous to IDE2. IDEF2 bound to OsYSL2 promoter region containing the binding core site, suggesting direct regulation of OsYSL2 expression. These results reveal novel cis-element/trans-factor interactions functionally associated with iron homeostasis.
Received for publication, October 22, 2007 , and in revised form, February 4, 2008.
The nucleotide sequence(s) reported in this paper has been submitted to the DDBJ/GenBankTM/EBI Data Bank with accession number(s) AB362160 and AB362161.
* This work was supported by a grant-in-aid for scientific research on priority areas from the Ministry of Education, Culture, Sports, Science and Technology of Japan and a grant from the Green Technology Project from the Ministry of Agriculture, Forestry and Fisheries of Japan. The costs of publication of this article were defrayed in part by the payment of page charges. This article must therefore be hereby marked "advertisement" in accordance with 18 U.S.C. Section 1734 solely to indicate this fact.
The on-line version of this article (available at http://www.jbc.org) contains supplemental Tables S1-S3.
1 These authors contributed equally to this work.
2 To whom correspondence should be addressed. Tel./Fax: 81-3-5841-7514; E-mail: annaoko{at}mail.ecc.u-tokyo.ac.jp.
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