Molecular basis of induction and execution of programmed cell death in plants

1.微阵列分析油菜素内酯（BRs）诱导的综合基因表达：为了解BRs诱导的气管元件（TE）细胞死亡机制，我采用微阵列技术对BRs诱导的基因表达进行了综合分析。因此，我揭示了大多数参与TE细胞死亡的基因都被BRs快速诱导，这表明BRs是细胞死亡相关基因转录激活的发起者。我还发现了转录因子在br中表现出短暂和快速的变化，这些转录因子是细胞死亡相关基因调控的候选因子。细胞死亡相关基因表达调控机制利用细胞死亡相关基因ZCP4作为标记物，鉴定出负责te特异性表达的顺式元件，命名为气管元件调节顺式元件（TERE）。该元件不仅在细胞死亡相关基因的启动子中保守，而且在细胞壁相关基因的启动子中也保守。利用酵母双杂交系统，分离出两个结合TERE序列的因子，均为bzip型转录因子。核酸酶分析esi对拟南芥S1核酸酶基因所有成员（BFN1-BFN5）进行启动子分析。BFN1仅是一个S1核酸酶基因，在发展中的TEs中表达。在走向细胞死亡的不同类型细胞中，S1核酸酶基因表达不同，表明每个S1核酸酶基因参与不同的细胞死亡过程。我还发现了两个钙依赖性核酸酶的候选基因，它们的活性在TE细胞死亡前短暂增加。对一个核定位脂肪酶的功能分析表明，百日草脂肪酶基因的mRNA以te特异性的方式增加。本文分离了拟南芥同源植物，分析了拟南芥脂肪酶基因的表达和功能。该基因在TEs的发生过程中优先表达，过量表达导致血管形成异常，提示其参与TEs的细胞死亡过程。

1.Microarray analysis of comprehensive gene expression induced by brassinosteroids (BRs) :To understand tracheary element (TE) cell death mechanism induced by BRs, I analyzed gene expression induced by BRs comprehensively using microarray. As a result, I revealed that most genes involved in TE cell death are induced by BRs rapidly, suggesting that BRs are an initiator of transcriptional activation of cell death-related genes. I also identified transcription factors showing transient and rapid changes by BRs, which are candidates of regulators of cell death related genes.2.Regulation mechanism of the expression of cell death-related genesUsing a cell-death related gene, ZCP4 as a marker, I identified a cis-element responsible for TE-specific expression, which is designated Tracheary Element Regulating cis Element (TERE). This element is conserved in promoters not only of cell death-related genes but also of cell wall-related genes. Using yeast two hybrid system, I isolated two factors that bind the TERE sequence, both of which were bZip-type transcription factors.3.Analysis of nucleasesI performed promoter analysis of all the S1 nuclease gene members in Arabidopsis (BFN1-BFN5). BFN1 was only a S1 nuclease gene that was expressed in developing TEs. In various types of cells going to cell death, different S1 nuclease genes were expressed, suggesting that each S1 nuclease gene is involved in distinctive cell death process. I also identified two candidate genes of calcium-dependent nucleases whose activity increases transiently before TE cell death.4.Function analysis of a nucleus-locating lipaseI have indicated a Zinnia lipase gene whose mRNA increases in a TE-specific manner. Here I isolated its Arabidopsis ortholog and analyzed the expression and function of the Arabidopsis lipase gene. This gene is expressed preferentially in developing TEs and overproduction of the gene caused abnormal vascular formation, suggesting its involvement in cell death process of TEs.