OXYGEN SENSING AND CELL SIGNALING IN HYPOXIA

描述（改编自申请人的摘要）：虽然细胞 呼吸是由常氧期间的代谢需要决定的，有证据表明， 细胞可以通过减少能量需求来适应缺氧，从而降低 ATP的使用和对O2的需求。 在复氧时，正常的代谢 恢复了细胞活动。 减少的能力 低氧时的能量需求，同时保持高能量磷酸盐水平 （缺氧适应）可能是保护在严重缺氧， ATP用于基本过程。 低氧适应需要细胞内的氧气 传感器能够检测PO2。 虽然可能存在多个传感器， 数据表明，细胞色素氧化酶作为传感器在缺氧 适应 具体目标1将测试细胞色素氧化酶是否起作用 作为氧传感器在缺氧期间通过降低其表观Vmax。 这将 使用抑制剂进行测试，该抑制剂降低氧化酶的Vmax， 常氧，确定这些是否激活缺氧适应 反应 缺氧期间O2传感器的激活必须与 随后激活细胞内信号级联， 最终抑制ATP的利用。 具体目标2将测试 活性氧（ROS）作为第二信使发挥作用假说 in this signaling信号pathway途径. 完整细胞中PO2依赖的ROS产生 将被研究并与氧化酶的功能相关联。 其他 研究将证实线粒体是否是ROS的来源， 将这些信号与低氧适应反应的激活联系起来。 对分离的线粒体的研究将确定线粒体的位点和机制。 缺氧时PO2依赖的ROS生成。 总的来说，这些研究 将阐明缺氧时线粒体ROS产生的机制 并将这些信号与细胞色素氧化酶的功能和缺氧 适应性反应 假设是下游的信号元件 ROS的释放导致ATP依赖性酶系统的抑制。 具体目标3 将开始检验蛋白激酶作为 低氧反应中的下游信号元件。 的参与 基于先前的研究，将对该途径中的蛋白激酶C进行检测 表明其被ROS或缺氧激活。 的长期目标 该项目旨在确定O2传感机制和下游 参与低氧适应的信号序列。 这些研究将 确定一种新的细胞O2检测途径，并可能有助于阐明 了解细胞如何适应低氧环境。

DESCRIPTION (Adapted from the applicant's abstract): Although cellular respiration is set by metabolic need during normoxia, evidence suggests that cells can adapt to hypoxia by reducing their energy demand, thereby lowering the use of ATP and the need for O2. At reoxygenation, normal metabolic processes are restored and cellular activity recovers. An ability to reduce energy demands in hypoxia while preserving high energy phosphate levels (hypoxic adaptation) may be protective during severe hypoxia by conserving ATP for essential processes. Hypoxic adaptation requires a cellular O2 sensor capable of detecting PO2. Although multiple sensors likely exist, data suggest that cytochrome oxidase acts as the sensor during hypoxic adaptation. Specific Aim 1 will test whether cytochrome oxidase functions as the O2 sensor during hypoxia by decreasing its apparent Vmax. This will be tested using inhibitors that reduce the Vmax of the oxidase during normoxia, determining whether these activate the hypoxic adaptation response. Activation of the O2 sensor during hypoxia must be coupled to subsequent activation of an intracellular signaling cascade, which ultimately inhibits ATP utilization. Specific Aim 2 will test the hypothesis that reactive oxygen species (ROS) function as a second messenger in this signaling pathway. The PO2-dependent ROS generation in intact cells will be studied and correlated with the function of the oxidase. Other studies will confirm whether mitochondria are the source of the ROS and will link these signals to the activation of the hypoxic adaptation response. Studies with isolated mitochondria will identify the sites and mechanisms of PO2-dependent ROS generation during hypoxia. Collectively, these studies will clarify the mechanisms of mitochondrial ROS generation during hypoxia and link these signals to the function of cytochrome oxidase and the hypoxic adaptation response. The hypothesis is that signaling elements downstream of ROS lead to inhibition of ATP-dependent enzyme systems. Specific Aim 3 will begin to test the hypothesis that protein kinases function as downstream signaling elements in the hypoxic response. The involvement of protein kinase C in this pathway will be tested, based on previous studies demonstrating its activation by ROS or by hypoxia. The long term goal of this project is to identify O2 sensing mechanisms and the downstream signaling sequence involved in hypoxic adaptation. These studies will identify a novel pathway of cellular O2 detection, and may help clarify understanding of how cells adapt to lowered O2 conditions.