Mitochondrial Retrograde Regulation in Plants: Identification of Regulated Genes and Signaling Pathway Components and Involvement in Stress Responses

真核细胞含有被称为细胞器(如细胞核、叶绿体、线粒体和过氧化物体)的膜结合亚室。细胞器之间的通讯对细胞的功能至关重要。特别令人感兴趣的是，叶绿体和线粒体等细胞器向细胞核传递其状态的方式，从而导致基因表达的变化。每种代谢活动的状态，如线粒体中的呼吸作用和三羧酸(TCA)循环，叶绿体中的光合作用，以及细胞器中发生的其他关键代谢途径，对细胞的功能至关重要。如果细胞器的代谢活动没有正常发挥作用，那么，一般而言，细胞器似乎能够将这一点传递给细胞核，从而使基因表达发生变化，这将有助于在分子水平上缓解这种情况。人们对这一过程的实现方式知之甚少，尤其是在高等真核生物中。环境变化是可能改变细胞器功能的一种重要的刺激类型。这包括光照水平和环境压力的变化，如冷应激、热应激、营养缺乏和病原体攻击。植物是固着的，因此，必须通过试图维持或恢复细胞内稳态来适应环境条件的变化。很明显，植物通过改变基因表达来响应压力和新陈代谢的变化。环境变化可能依赖于细胞器和细胞核之间的通讯(这一过程被称为基因表达的逆行调节)，以改变对特定胁迫的成功反应至关重要的关键基因的表达。本项目的主要研究重点是检验这一假说，即植物中从线粒体到细胞核有多种通讯方式，称为信号通路：一种途径与细胞核沟通，说明呼吸受到了扰动，这可能是由某些环境胁迫引起的；另一种途径是传达代谢过程受到扰动，如TCA循环，这也可能是由一些环境胁迫引起的。如果线粒体到细胞核的信号是植物对胁迫的整体反应的关键部分，那么确定线粒体到细胞核的信号在个体胁迫反应中的确切作用将是非常有趣的。将使用几种方法来解决这一问题。负责响应线粒体逆行信号的AtAOX1a基因的启动子(基因表达控制)区域将在特定位置发生突变，以确定对该反应重要的区域。还将确定由于线粒体呼吸或TCA周期的扰动而导致的基因表达的基因组变化的完整视图。遗传筛选将被用来分离缺乏线粒体逆行信号通路的突变体拟南芥植株。将对这些突变植物进行分析，以确定突变对每一株植物的影响。本科生，特别是那些在生命科学中代表性不足的群体的学生，将被招募参加这一研究计划。研究生培训和博士后培训也将是该项目的主要部分。由于我的实验室使用的方法多种多样，而且有对细胞器间交流感兴趣的亲密同事的存在，因此为学生和博士后研究员提供的培训环境应该非常令人兴奋。

Eukaryotic cells contain membrane-bound sub-compartments called organelles (e.g. nuclei, chloroplasts, mitochondria, and peroxisomes). Communication between organelles is of central importance to the functioning of cells. Of particular interest is the way in which organelles such as chloroplasts and mitochondria communicate their status to the nucleus to cause changes in gene expression. The status of each of the metabolic activities, such as respiration and the tricarboxylic acid (TCA) cycle in the mitochondria, photosynthesis in the chloroplasts, and other key metabolic pathways that take place in organelles, is vital to the functioning of cells. If the metabolic activities of the organelles are not functioning properly, then, in general, the organelles seem to be able to communicate this to the nucleus so that changes in gene expression can occur that will help to alleviate the situation at the molecular level. Relatively little is known about the way in which this is accomplished, especially in higher eukaryotes. One important category of the type of stimuli that may alter organelle function is environmental changes. This includes changes in light level and environmental stresses such as cold stress, heat stress, nutrient deprivation and pathogen attack. Plants are sessile and, therefore, must adjust to changes in environmental conditions by attempting to maintain or regain cellular homeostasis. It is clear that plants respond to stress and changes in metabolism by altering gene expression. Environmental changes may rely on communication between organelles and the nucleus (a process referred to as retrograde regulation of gene expression) to alter the expression of key genes vital to a successful response to the particular stress. The main focus of the research in this project it to test the hypothesis that there are multiple modes of communication, referred to as signaling pathways, from mitochondria to the nucleus in plants: one pathway to communicate to the nucleus that there has been a perturbation of respiration, which may be caused by some environmental stresses, and another pathway to communicate that there has been a perturbation of metabolic processes, such as the TCA cycle, which also may be caused by some environmental stresses. If mitochondria-to-nucleus signaling is a key part of the overall response of plants to stresses, then it will be very interesting to determine the exact roles of mitochondria-to-nucleus signaling in individual stress responses. Several approaches will be used to address this problem. The promoter (gene expression controlling) region of the AtAOX1a gene, which responds to mitochondrial retrograde signaling, will be mutated at specific sites to determine regions that are important for the response. A full view of the genomic changes in gene expression that result from perturbations of mitochondrial respiration or perturbation of the TCA cycle will also be determined. A genetic screen will be utilized to isolate mutant Arabidopsis thaliana plants that lack mitochondrial retrograde signaling pathways. These mutant plants will be analyzed to determine the effects of the mutation in each. Undergraduate students, especially those of groups that are under-represented in the life sciences, will be recruited for participation in this research program. Graduate student training and post-doctoral training are also intended to be a major part of this project. Because of the diverse approaches used in my laboratory and the presence of close colleagues interested in inter-organellar communication, the training environment for students and post-doctoral fellows should be very stimulating.