OXYGEN SENSING AND REGULATION OF YEAST GENES

细胞、组织和生物体对氧分压变化的适应取决于一大群不同蛋白质表达水平的变化。目前，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，但目前还不清楚大多数生物体是如何感受到氧气的。同样不清楚的是，“氧传感器”如何将其信号传输到激活或抑制氧反应基因的信号转导途径。在酵母中，对氧气供应变化作出反应的基因可以分为两大类：在常氧条件下最佳表达的需氧基因和在低氧或缺氧条件下最佳表达的低氧基因。目前酵母氧感应模型假定，血红素浓度受氧浓度的影响，血红素调节氧反应基因转录因子的活性，并且血红素是这些转录因子的氧化还原不敏感辅因子。最近的发现对这一简单的模型提出了质疑，并表明氧气感知涉及多个途径/机制。这项建议的研究目标是基于一项新的实验发现，即有氧基因的表达水平取决于氧气的浓度，而不仅仅是氧气的存在或不存在。这种现象被称为氧浓度依赖(OCD)基因表达，意味着细胞本身实际上检测到氧浓度并对其做出反应。这种类型的基因表达，加上允许它发现的独特的发酵系统，通过提出以下问题，打开了一种检查酵母氧气感应上游事件的方法：目标1：有氧基因表达水平的变化能否通过血红素水平的变化来解释？目的2：细胞色素c氧化酶、YHb黄化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？目标3：氧气是否会影响成绩单的周转率？目的4：COX6、COX5a和Cyc1的强迫症表达由哪些顺式启动子调控？

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?