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Moleculaabiological analysis of phytochrome siggnal transduction

光敏色素信号转导的分子生物学分析

基本信息

批准号：
06454018
负责人：
NAGATANI Akira
金额：
$ 4.61万
依托单位：
UNIVERSITY of TOKYO (1995)The Institute of Physical and Chemical Research (1994)
依托单位国家：
日本
项目类别：
Grant-in-Aid for General Scientific Research (B)
财政年份：
1994
资助国家：
日本
起止时间：
1994 至 1995
项目状态：
已结题

项目摘要

Phytochrome is a chromoprotein which is common in green plants.Phytochrome is know to be act as a major photoreceptor mediating divergent plant responses to light.Although intensive work has been done on phytochrome, molecular mechanism in which phytochrome transduce the light signal to signal transduction factors.We addressed this problem with molecular as well as biochemical techniques in the present study.To examine biological activity of the C-terminal half, which has been speculated to be involved in the signal transduction process, we prepared transgenic Arabidopsis expressing a C-terminal fragment of phytochrome B(phyB).The analysis of those plants indicated that the C-terminal region alone could not interact with the endogenous signal transduction mechanism.Next, we investigated intracellular localization of phyB with the aid of a reporter protein.C-terminal fragments of phyB was fused to GUS reporter protein and expressed in transgenic Arabidopsis.Histochemical staining of GUS activity in those plants revealed that the fusion protein localized to the nucleus.Nuclear localization of endogenous phyB was confirmed by immunoblotting analysis of nuclei isolated from light-grown Arabidopsis.Furthermore, the level of phyB in the isolated nuclei was substantially decreased if plants was irradiated with far-red light and then kept in darkness.Taken together, is possible that nuclear localization of phyB is an important step of phytochrome signaling process.We have also examined biolochemical and biological activity of fern phytochrome expressed in higher plants.The results indicated that the fern phytochrome is photochemically active in higher plants.Nevertheless, it did show only very limited biological activity.So, we concluded that fern phytochrome is structurally too diverse from higher plant phytochrome to transduce the signal in higher plants.

光敏色素是一种存在于绿色植物中的色素蛋白。光敏色素是一种重要的光感受器，参与调节植物对光的各种反应。光敏色素的分子机制是光敏色素将光信号传递给信号转导因子。本研究利用分子生物学技术对光敏色素的生物学活性进行了研究。其被推测参与信号转导过程，我们制备了表达光敏色素B（phyB）C-末端片段的转基因拟南芥。对这些植物的分析表明，单独的C-末端区域不能与内源信号转导机制相互作用。接下来，我们借助于报道蛋白研究了phyB的细胞内定位。将phyB基因的末端片段与GUS报告蛋白融合，在转基因拟南芥中表达，GUS组织化学染色结果表明，融合蛋白定位于细胞核，免疫印迹分析证实了内源phyB基因的细胞核定位此外，如果植物用远红光照射然后保持在黑暗中，则分离的细胞核中的phyB水平显著降低。PhyB的核定位可能是光敏色素信号转导过程中的一个重要步骤.我们还研究了在高等植物中表达的蕨类光敏色素的生物化学和生物活性.结果表明，蕨类光敏色素是一种重要的光敏色素，因此，我们认为蕨类植物光敏色素在结构上与高等植物光敏色素的差异太大，不可能抑制高等植物光敏色素的光化学活性。