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Originally published In Press as doi:10.1074/jbc.M007405200 on December 22, 2000

J. Biol. Chem., Vol. 276, Issue 17, 14139-14152, April 27, 2001
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A Rice Functional Transcriptional Activator, RISBZ1, Responsible for Endosperm-specific Expression of Storage Protein Genes through GCN4 Motif*

Yasuyuki OnoderaDagger , Akihiro SuzukiDagger §, Chuan-Yin WuDagger , Haruhiko WashidaDagger ||, and Fumio TakaiwaDagger **

From the Dagger  Department of Biotechnology, National Institute of Agrobiological Resources, Kannondai 2-1-2, Tsukuba, Ibaraki 305-8602 and the || Faculty of Horticulture, Chiba University, Matsudo, Chiba 271-8510, Japan

The GCN4 motif, a cis-element that is highly conserved in the promoters of cereal seed storage protein genes, plays a central role in controlling endosperm-specific expression. This motif is the recognition site for a basic leucine zipper transcriptional factor that belongs to the group of maize Opaque-2 (O2)-like proteins. Five different basic leucine zipper cDNA clones, designated RISBZ1-5, have been isolated from a rice seed cDNA library. The predicted gene products can be divided into two groups based on their amino acid sequences. Although all the RISBZ proteins are able to interact with the GCN4 motif, only RISBZ1 is capable of activating (more than 100-fold expression) the expression of a reporter gene under a minimal promoter fused to a pentamer of the GCN4 motif. Loss-of-function and gain-of-function experiments using the yeast GAL4 DNA binding domain revealed that the proline-rich N-terminal domain (27 amino acids in length) is responsible for transactivation. The RISBZ1 protein is capable of forming homodimers as well as heterodimers with other RISBZ subunit proteins. RISBZ1 gene expression is restricted to the seed, where it precedes the expression of storage protein genes. When the RISBZ1 promoter was transcriptionally fused to the beta -glucuronidase reporter gene and the chimeric gene was introduced into rice, the beta -glucuronidase gene is specifically expressed in aleurone and subaleurone layer of the developing endosperm. These findings suggest that the specific expression of transcriptional activator RISBZ1 gene may determine the endosperm specificity of the storage protein genes.

* This work was supported by research grants from the Bio-oriented Technology Research Advancement Institute (PROBRAIN), Science and Technology Agency (Enhancement System of Center of Excellence), and the Ministry of Agriculture, Forestry and Fishery of Japan (to F. T.).The costs of publication of this article were defrayed in part by the payment of page charges. The article must therefore be hereby marked "advertisement" in accordance with 18 U.S.C. Section 1734 solely to indicate this fact.

§ Present address: Faculty of Science, Kagoshima University, Kagoshima 890-0065, Japan.

Present address: Ceres, Inc., Malibu, CA 90265.

** To whom correspondence should be addressed. Tel.: 81-298-38-8373; Fax: 81-298-38-8397; E-mail: takaiwa@abr.affrc.go.jp.

Copyright © 2001 by The American Society for Biochemistry and Molecular Biology, Inc.
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