DECIPHERING THE RELATIONSHIP BETWEEN GLUTATHIONE AND PROGRAMMED CELL DEATH IN TR

这个子项目是许多利用资源的研究子项目之一 由NIH/NCRR资助的中心拨款提供。子项目的主要支持 而子项目的主要调查员可能是由其他来源提供的， 包括其它NIH来源。 列出的子项目总成本可能 代表子项目使用的中心基础设施的估计数量， 而不是由NCRR赠款提供给子项目或子项目工作人员的直接资金。 程序性细胞死亡（PCD）是一种基本的细胞过程，其特征在于细胞收缩、核浓缩和碎裂，导致细胞的选择性消除，并且已被报道在囊性纤维化、肿瘤发生和其他疾病发生中起主要作用。有趣的是，在植物中似乎存在一些进化上保守的类似植物程序性细胞死亡（AL-PCD）的信号通路。我们建议使用植物模型，烟草（烟草），谷胱甘肽在细胞凋亡中的作用的特点。谷胱甘肽（GSH）被认为是最重要的水溶性抗氧化剂，存在于植物和动物中。谷氨酰胺通过GSH还原酶降低细胞中的氧化应激和水平，并且GSH水平的降低已显示直接激活促凋亡途径。将使用具有葡萄糖氧化酶（GOX）基因的诱导型表达的转基因烟草植物。当表达时，GOX蛋白催化植物细胞中葡萄糖的分解;该过程的副产物是产生H2 O2和随后的氧化应激。我们的具体目的是（1）使用蛋白质印迹和LCMS分析来测量GSH表达的变化，这是转基因GOX烟草植物中H2 O2产生的结果，（2）使用DNA梯状分析来测量凋亡，以确定GSH水平与植物凋亡的关系。我们的假设是GSH水平将由于H2 O2的产生而增加，并导致AL-PCD。或者，（3）我们计划通过测量光合参数的变化来测量GOX表达对叶片生理的影响。如果成功，我们的项目将建立使用植物作为模型系统来研究动物PCD的可行性。使用植物作为模型系统是本科实验室的理想选择。植物比动物系统更容易维护，成本更低，需要的技术技能和经验也更少。

This subproject is one of many research subprojects utilizing the resources provided by a Center grant funded by NIH/NCRR. Primary support for the subproject and the subproject's principal investigator may have been provided by other sources, including other NIH sources. The Total Cost listed for the subproject likely represents the estimated amount of Center infrastructure utilized by the subproject, not direct funding provided by the NCRR grant to the subproject or subproject staff. Programmed cell death (PCD) is an essential cellular process characterized by cell shrinkage, nuclear condensation and fragmentation that leads to the selective elimination of cells and has been reported to play a major role in cystic fibrosis, oncogenesis and other disease occurrences. Interestingly, there appears to be some evolutionarily conserved signaling pathways for an apoptotic-like programmed cell death (AL-PCD) in plants. We propose to use a plant model, tobacco (Nicotiana tobacum), to characterize the role of glutathione in apoptosis. Glutathione (GSH) is considered to be the most important water-soluble antioxidant and is found in both plants and animals. Glutathione functions to reduced oxidative stress and levels in the cell via GSH reductase and decreases in GSH levels have been shown to directly activate pro-apoptotic pathways. Transgenic tobacco plants with the inducible expression of a glucose oxidase (GOX) gene will be employed. When expressed the GOX protein catalyzes the breakdown of glucose in the plant cell; a byproduct of this process is the generation of H2O2 and subsequently oxidative stress. Our Specific Aims are (1) to measure changes in GSH expression, using western blot and LCMS analysis, as result of H2O2 production in transgenic GOX tobacco plants, (2) measure apoptosis using DNA laddering assays to determine the relationship of GSH levels and apoptosis in plants. It is our hypothesis that GSH levels will increase as a result of H2O2 production and result in AL-PCD. Alternately, (3) we plan to measure the effects of GOX expression on leaf physiology by measuring changes in photosynthetic parameters. If successful, our project will establish the viability of using plants as a model system to study PCD in animals. The use of plants as a model system is ideal for the undergraduate lab. Plants are easier and cheaper to maintain than animals systems and require less technical skill and experience.