ECD8蛋白调控叶绿体基因转录的分子机制

叶绿体是植物特有的细胞器，是光合作用的场所。叶绿体基因正确的表达与调控，决定着叶绿体的发育进程以及光合功能的实现。叶绿体细菌类型的多亚基RNA聚合酶PEP是叶绿体基因主要的转录机器，其功能的正常行使需要核基因编码的蛋白因子的精细调控。我们通过荧光成像高通量筛选的方法，获得了一个叶绿体发育受阻突变体ecd8。该突变体为幼苗白化致死的表型，其光系统Ⅱ的功能完全丧失。通过实验，我们确定ECD8影响PEP的转录活性。ECD8是核基因编码的功能未知叶绿体蛋白，含有两个DNA结合结构域GAT1。我们推断，ECD8是通过结合叶绿体DNA来实现对叶绿体基因的转录调控。本项目将综合运用遗传学、分子生物学、生物化学等多种技术手段，深入研究ECD8的生物学功能。再结合结构生物学，解析ECD8-DNA复合物的晶体结构，揭示ECD8与DNA的结合模式，阐明ECD8调控叶绿体基因转录的分子机制。

The chloroplast is unique to higher plants, in which photosynthesis takes place. The correct expression and regulation of chloroplast genes is essential for chloroplast development and photosynthesis. Plastid-encoded polymerase (PEP) is the major transcriptional machine and is composed by both prokaryotic core subunits and essential subunits encoded by nucleus genes. Thousands of nuclear-encoded protein factors are required to regulate the transcriptional activity of PEP. We have screened a T-DNA insertion mutant ecd8 with the lethal phenotype of albino through high throughput screening of high chlorophyll fluorescence imaging. The function of PSⅡ in ecd8 is completely abolished. Through our experiments, we have found that ECD8 plays an important role in chloroplast gene transcription. ECD8 is a nucleus-encoded chloroplast protein. It contains two GAT1 domains with Rossmann fold which can bind DNA. So, we deduce that ECD8 regulates chloroplast gene transcription by binding with DNA. But what is the binding model and how ECD8 regulates the transcription? By multiple technical means of genetics, molecular biology, biochemistry, we will investigate the biological function of ECD8. Through the structural research of ECD8-DNA complex, we attempt to shed light on the molecular mechanism of ECD8 in regulating the transcription of chloroplast genes.