Molecular analysis of cell death through plant plastid

质体是植物细胞的主要细胞器之一，它有自己的基因组。叶绿体是质体的一种形式，它捕获光能以ATP的形式保存自由能，并将NADP还原为NADPH产生糖。本研究旨在阐明质体调控植物细胞死亡的机制。拟南芥Cdf1最初是作为酵母细胞死亡诱导剂分离出来的。在拟南芥基因组中，有3个与cdf相关的基因（Cdf1-3）。T-DNA插入植株表明，杂合子Cdf1突变体在最佳生长条件下表现出正常的营养表型，但Cdf1纯合子胚胎在球形阶段停滞。这种表型由Cdf1的过表达补充。为了研究Cdf1基因在胚胎发生过程中的表达，我们进行了启动子- gus实验。包含Cdf1启动子区和编码区的5kb基因组片段与一个报告基因GUS相连。将合成的质粒导入农杆菌，转化为拟南芥。GUS实验表明Cdf1B在球状和心脏阶段之间的一个阶段表达。这些数据表明，质体包膜蛋白Cdf1对胚胎发生至关重要。此外，在Cdf1-3中插入T-DNA的植物在黑暗诱导的叶片衰老中表现出延迟表型。进一步的生化分析将阐明Cdf基因在植物中的分子功能。

The plastid, one of the major organelles in the plant cell, has its own genome. Chloroplast, which is a one form of plastid, capture light energy to conserve free energy in the form of ATP and reduce NADP to NADPH to produce sugars. In this research, we aimed to clarify the mechanisms of plant cell death regulated by plastid. Arabidopsis Cdf1 was originally isolated as a cell death inducer in yeast. In Arabidopsis genome, there are three Cdf-related genes (Cdf1-3). T-DNA inserted plants demonstrated that the heterozygous Cdf1 mutant showed normal vegetative phenotypes under optimal growth conditions, but embryos homozygous for Cdf1 arrested at the globular stage. Such phenotype was complemented by the overexpression of Cdf1. To investigate the gene expression of Cdf1 during embryogenesis, promoter-GUS assay was performed. The 5 kb genomic fragment containing Cdf1 promoter region and coding region was connected with a reporter gene, GUS. Resultant plasmid was introduced into Agrobacterium, and transformed to Arabidopsis. GUS assay demonstrated that Cdf1B expressed during a stage between globular and heart stages. These data indicate that plastid envelope protein Cdf1 is essential for embryogenesis. Furthermore, T-DNA inserted plants in Cdf1-3 demonstrated delayed phenotype in dark-induced leaf senescence. Further biochemical analysis will clarify the molecular function of Cdf genes in plant.