Wu C Y, Suzuki A, Washida H, Takaiwa F
Department of Biotechnology, National Institute of Agrobiological Resources, Ibaraki, Japan.
Plant J. 1998 Jun;14(6):673-83. doi: 10.1046/j.1365-313x.1998.00167.x.
The GCN4 motif is conserved in a number of seed storage protein genes, and promoter fragments containing this motif have been shown to be involved in controlling seed-specific expression of the genes studied. All genes encoding the rice seed storage protein glutelin contain the GCN4 motif at similar sites in their 5' flanking regions. Using a stable homologous transgenic system, we have analysed the promoter of the rice glutelin gene GluB-1 and demonstrated that the GCN4 motif functions as an essential cis-element for endosperm-specific gene expression. Moreover, a 21 bp GluB-1 promoter fragment spanning the GCN4 motif, as a multimer, directed GUS gene expression in endosperm of transgenic rice plants, when fused directly to the core promoter (-46) of CaMV 35S. In transiently transfected rice protoplasts, over a hundred-fold transactivation was observed from the 21 bp sequence by the bZIP type transcriptional activator Opaque-2 (O2) co-expressed under a CaMV 35S promoter. The transactivation was also evident in transgenic plants containing both O2 and the 21 bp sequence/GUS fusion. The O2-mediated activation requires binding of O2 to an intact GCN4 motif. Our results suggest that a bZIP protein functionally similar to O2 may exist in rice and participate in controlling the endosperm-specific expression of GluB-1 through the GCN4 motif.
GCN4基序在许多种子贮藏蛋白基因中是保守的，并且已表明含有该基序的启动子片段参与了所研究基因的种子特异性表达的调控。所有编码水稻种子贮藏蛋白谷蛋白的基因在其5'侧翼区域的相似位点都含有GCN4基序。利用稳定的同源转基因系统，我们分析了水稻谷蛋白基因GluB-1的启动子，并证明GCN4基序作为胚乳特异性基因表达的必需顺式元件发挥作用。此外，跨越GCN4基序的21 bp GluB-1启动子片段作为多聚体，当直接与CaMV 35S的核心启动子（-46）融合时，可指导转基因水稻植株胚乳中的GUS基因表达。在瞬时转染的水稻原生质体中，在CaMV 35S启动子控制下共表达的bZIP型转录激活因子不透明-2（O2）从21 bp序列观察到了超过百倍的反式激活。在同时含有O2和21 bp序列/GUS融合体的转基因植物中，反式激活也很明显。O2介导的激活需要O2与完整的GCN4基序结合。我们的结果表明，水稻中可能存在一种功能上与O2相似的bZIP蛋白，并通过GCN4基序参与控制GluB-1的胚乳特异性表达。
