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Published on June 23, 2006; 10.1104/pp.106.082826

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Received May 1, 2006
Returned for revision May 10, 2006
Accepted June 14, 2006

Synergism between RPBF Dof and RISBZ1 bZIP activators in the regulation of rice seed expression genes

Masayuki P. Yamamoto , Yasuyuki Onodera , Satoru M. Touno , and Fumio Takaiwa *

Transgenic Crop Research and Development Center, National Institute of Agrobiological Sciences, Kannondai 2-1-2, Tsukuba Ibaraki 305-8602, Japan
Transgenic Crop Research and Development Center, National Institute of Agrobiological Sciences, Kannondai 2-1-2, Tsukuba Ibaraki 305-8602, Japan; Graduate School of Science and Technology, Chiba University, Matsudo, Chiba 271-8510, Japan

* Corresponding author; email: takaiwa{at}nias.affrc.go.jp.

The Dof zinc finger protein called RPBF (rice prolamin box binding factor), which is involved in gene regulation of rice seed storage proteins, has been isolated from rice cDNA EST clones containing the conserved Dof. RPBF is found as a single gene per haploid genome. Comparison of RPBF genomic and cDNA sequences revealed that the genomic copy is interrupted by one long intron of 1892 bp in the 5' noncoding region. We demonstrated by transient expression in rice callus protoplasts that the isolated RPBF trans-activated several storage protein genes via an AAAG target sequence located within their promoters, and with methylation interference experiments that additional AAAG-like sequences in promoters of genes expressed in maturing seeds were recognized by the RPBF protein. Binding was sequence-specific, since mutation of the AAAG motif or its derivatives decreased both binding and trans-activation by RPBF. Synergism between RPBF and RISBZ1 recognizing the GCN4 motif (TGA(G/C)TCA) was observed in the expression of many storage protein genes. Over-expression of both transcription factors gave rise to much higher levels of expression than the sum of individual activities elicited by either RPBF or RISBZ1 alone. Furthermore, mutation of recognition sites suppressed reciprocal trans-activation ability, indicating that there are mutual interactions between RISBZ1 and RPBF. The RPBF gene is predominantly expressed in maturing endosperm and coordinately expressed with seed storage protein genes, and is involved in the quantitative regulation of genes expressed in the endosperm in co-operation with RISBZ1.




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