Molecular Mechnisms of Meristematic Tissue-Specific Expression of Plnat Histone Genes Introduced by a Ti-Plasmid Vector, Especially concerning Cis-Acting Control Elements for the Initiation and Termination of

在过去的两年里，为了研究细胞周期中S期特异表达的基因转录调控的分子机制，我们利用克隆的小麦组蛋白H3和H4基因对组蛋白基因的顺式和反式调控元件进行了研究。为了检测组蛋白基因的转录活性，利用钛载体介导的基因转移系统，将小麦组蛋白基因通过钛质粒导入向日葵下胚轴细胞，并通过S1检测小麦基因在转化细胞中的转录活性。对H3和H4基因5‘端缺失突变体的转录活性分析表明，一些促进转录高效的顺式元件位于-668和191位之间，六聚体序列位于目前所检测的所有植物组蛋白基因中存在的八聚体序列的上游，似乎与转录调控有关。通过凝胶滞留实验、甲基化干扰实验和DNA酶I足迹实验证明，小麦细胞中存在与六聚体序列特异结合的核蛋白(S)。核蛋白，暂定为HBP-1，在萌发中比休眠的萌发中含有更丰富的蛋白质。此外，在用杀虫双处理部分同步细胞周期的小麦幼苗中，从S相丰富的细胞群中提取的细胞核提取液中，HBP-1的含量最高。这些数据表明，六聚体序列和HBP-1分别是参与小麦组蛋白基因细胞周期调控转录的顺式和反式调控元件。

During the last two years, in order to study the molecular mechanisms of the transcriptional regulation of the genes which are specifically experssed at the S phase during cell cycle, we have examined the cis- and trans-acting Control elements of the histone genes using the cloned wheat histone H3 and H4 genes. To examine the transcriptional activity of the histone gene, the Ti-plasmid mediated gene transfer system was used; the wheat histone genes were introduced into sunflower hypocotyl cells via Ti-plasmid and the transcripts of the wheat genes in transformed cells were measured by the S1 assay.The analysis of the transcriptional activities of the 5' deletion mutants of the H3 and H4 genes revealed that some cis-elements promoting a high efficiency of the transcription are located between the positions -668 and 191 and that the hexameric sequence just upstream from the octameric sequence which exists in all plant histone genes examined so far seemd to have some connection with the transcriptional regulation. On the other hand, the octameric sequence was not shown to be a motif to regulate the transcription of the histone genes.Several experiments using the gel retardation assay, methylation interference assay and DNase I footprint method have demonstrated that there exist nuclear protein(s) specifically binding to the hexameric sequence in wheat cells. The nuclear protein Which was tentatively termed HBP-1 are more abundantly contained in germinating than dormant germs. Furthermore, in wheat seedling in which the cell cycle was partially synchronized by aphidicoline treatment, HBP-1 was persent in the highest concentration in the nuclear extract from an S phase-abundant cell population. These data have suggested that the hexameric sequence and HBP-1 are cis- and trans-acting control elements, respectively, involved in cell cycle-dependent transcription of the wheat histone genes.