OXYGEN SENSING AND REGULATION OF YEAST GENES

细胞、组织和生物体对氧张力变化的适应取决于大量不同蛋白质组表达水平的变化。 目前，尚不清楚大多数生物体是如何感受到氧气的，尽管人们普遍认为血红素在这一过程中起着至关重要的作用。 同样不清楚的是，“氧传感器”如何将其信号传递到信号转导途径，以激活或抑制氧反应基因。 在酿酒酵母中，响应氧可用性变化的基因可以分为两大类：好氧基因，其在常氧条件下最佳表达，和低氧基因，其在低氧或缺氧条件下最佳表达。 目前酵母中的氧传感模型假设血红素浓度受氧浓度的影响，血红素的功能是调节氧响应基因的转录因子的活性，并且血红素是这些转录因子的氧化还原不敏感的辅因子。最近的研究结果对这个简单的模型提出了质疑，并表明多种途径/机制参与了氧的感知。这项建议的研究目标是基于新的实验发现，即有氧基因的表达水平取决于氧的浓度，而不仅仅是它的存在或不存在。 这种现象被称为氧浓度依赖性（OCD）基因表达，意味着细胞实际上检测并响应氧浓度本身。 这种类型的基因表达，连同独特的发酵罐系统，允许其发现，打开了一种方法来检查上游事件，参与在酵母中的氧传感，通过问以下问题：目的1：可以在好氧基因的表达水平的变化被解释的血红素水平的变化？ 目的2：细胞色素c氧化酶、YHb黄素血红蛋白或其他氧化还原敏感的氧结合蛋白是否参与强迫症基因的表达？ 目的3：氧气是否影响转录本周转率？目的4：COX 6、COX 5a和CYC 1的顺式启动子位点介导OCD的表达？

Adaptation to changes in oxygen tension in cells, tissues, and organisms depends on changes in the level of expression of a large and diverse group of proteins. Currently, it is not clear how oxygen is sensed in most organisms although it is generally agreed that heme plays a vital role in this process. Also unclear is how an 'oxygen sensor' transmits its signal to a signal transduction pathway for the activation or repression of oxygen-responsive genes. In the yeast Saccharomyces cerevisiae genes that respond to changes in oxygen availability can be placed into two broad categories: aerobic genes, which are optimally expressed under normoxic conditions, and hypoxic genes, which are optimally expressed under low oxygen or anoxic conditions. The current model for oxygen sensing in yeast assumes that heme concentration is affected by oxygen concentration, that heme functions to regulate the activity of transcription factors for oxygen-responsive genes, and that heme is a redox-insensitive cofactor for these transcription factors. Recent findings have cast doubt on this simple model and suggest that multiple pathways/mechanisms are involved in sensing oxygen. The research objectives of this proposal are based on the novel experimental finding that the level of expression of aerobic genes is determined by the concentration of oxygen and not merely its presence or absence. This phenomenon, called oxygen concentration-dependent (OCD) gene expression, implies that cells actually detect and respond to oxygen concentration per se. This type of gene expression, together with the unique fermentor system that permitted its discovery, opens a way to examine upstream events that are involved in oxygen sensing in yeast, by asking the following questions: Aim 1: Can changes in level of expression of aerobic genes be explained by changes in heme levels? Aim 2: Is cytochrome c oxidase, YHb flavohemoglobin, or some other redox-sensitive oxygen binding protein involved in OCD gene expression? Aim 3: Does oxygen affect transcript turnover rate? Aim 4: What cis promoter sites mediate the OCD expression of COX6, COX5a, and CYC1?