Mechanisms of Oxygen Sensing

描述（由申请人提供）：氧稳态对生命和健康至关重要，包括肺和心血管健康。慢性缺氧与多种病理有关，包括肺部相关疾病，如COPD、肺癌、肺动脉高压、纤维化和炎症。对氧感应途径的研究已经揭示了氧稳态的重要介质，然而，氧感应的分子机制的详细知识仍未被揭示。更全面地了解细胞氧感应途径可能导致肺部和心血管疾病治疗的新治疗靶点的鉴定。在细胞水平上，缺氧激活缺氧诱导转录因子（hif）。hif与大量靶基因的启动子/增强区域中的缺氧反应元件结合，导致基因程序的激活，包括糖酵解、血管生成和红细胞生成的上调。在常氧条件下，HIF¿蛋白亚基被脯氨酸羟化酶结构域蛋白2 （PHD2）羟基化，被von Hippel-Lindau （VHL）泛素连接酶靶向泛素化，并被蛋白酶体降解。在缺氧条件下，PHD2的活性被抑制，从而允许HIF¿蛋白的积累和随后与HIF¿亚基的结合来诱导转录反应。氧水平降低PHD2活性的潜在机制尚不完全清楚。我们之前已经证明，缺氧会增加线粒体复合体III中ROS的产生，从而抑制HIF¿蛋白羟基化和随后的HIF¿蛋白稳定。在这项提议中，我们将利用蛋白质组学方法来测试缺氧诱导的线粒体ROS是否通过结合未知蛋白质、翻译后修饰PHD2和/或氧化PHD2内半胱氨酸残基来抑制PHD2活性。目前临床上还没有成功的靶向抑制hif的药物。这项探索性资助将提供详细的了解

DESCRIPTION (provided by applicant): Oxygen homeostasis is essential for life and health, including pulmonary and cardiovascular health. Chronic hypoxia is associated with a myriad of pathologies including pulmonary related diseases such as COPD, lung cancer, pulmonary hypertension, fibrosis and inflammation. Research on oxygen sensing pathways has revealed important mediators of oxygen homeostasis, however, detailed knowledge of the molecular mechanisms of oxygen sensing remain uncovered. Fuller understanding of the cellular oxygen sensing pathways may lead to identification of novel therapeutic targets for treatment of pulmonary and cardiovascular disease. At the cellular level, hypoxia activates the Hypoxia Inducible transcription Factors (HIFs). HIFs bind to hypoxia-response elements in the promoter/enhance regions of a large number of target genes resulting in activation of a genetic program that includes upregulation of glycolysis, angiogenesis, and erythropoiesis. Under normoxic conditions, the HIF¿ protein subunit is hydroxylated by Prolyl Hydroxylase Domain protein 2 (PHD2), targeting it for ubiquitination by the von Hippel-Lindau (VHL) ubiquitin ligase, and proteasomal degradation. The activity of PHD2 is inhibited under hypoxic conditions, allowing the accumulation of HIF¿ protein and the subsequent binding to HIF¿ subunit to induce the transcriptional response. The underlying mechanism by which oxygen levels diminish PHD2 activity is not fully understood. We have previously demonstrated that hypoxia increases the generation of ROS from mitochondrial complex III resulting in inhibition of HIF¿ protein hydroxylation and subsequent HIF¿ protein stabilization. In this proposal we will utilize a proteomic approach to test whether hypoxia-induced mitochondrial ROS lead to inhibition of PHD2 activity by binding of unidentified proteins, post-translational modification of PHD2, and/or oxidizing of cysteine residues within PHD2. Currently, there have been no successful drugs in the clinic targeting inhibition of HIFs. This exploratory grant will provide detailed understanding of how PHD2 is regulated during hypoxia leading to rationale molecular basis for therapeutic targeting of HIFs.