Prolyl Hydroxylation and Neuronal Cell Death

描述（由申请人提供）：细胞死亡在神经系统发育过程中广泛存在，它有助于确保神经元及其靶点之间形成适当数量和类型的连接。在发育过程中，当神经元不能从营养因子如神经生长因子（NGF）接收足够的存活信号时，细胞死亡发生。越来越多的证据表明，营养因子剥夺诱导的死亡也有助于人类神经元疾病和退行性疾病。我们以前确定脯氨酰羟化酶EGLN 3作为介质的神经生长因子剥夺诱导的细胞死亡。EGLN 3是脯氨酰羟化酶的小家族之一，其作为转录因子缺氧诱导因子（HIF）的氧依赖性调节剂起关键作用。除了调节HIF外，最近的报道表明EGLN具有HIF非依赖性功能。我们已经获得了EGLN 3和促凋亡Bcl-2家族蛋白BIM（EL）之间新的相互作用的初步证据。此外，我们发现EGLN 3和BIM（EL）各自与von Hippel-Lindau蛋白（pVHL）相互作用，pVHL是E3泛素连接酶的一个亚基，靶向HIF降解。在这里，我们将测试的假设，EGLN 3，pVHL，BIM（EL）协调功能，以调节营养因子剥夺诱导的细胞死亡。在目标1中，我们将确定EGLN 3和BIM（EL）之间的功能关系，在神经生长因子剥夺诱导的细胞死亡，使用过表达和基因敲除和敲低的方法相结合。目的2阐述EGLN 3/BIM（EL）相互作用的生物化学意义。实验将表征EGLN 3和BIM（EL）之间的相互作用如何影响BIM（EL）的促凋亡功能和/或EGLN 3的脯氨酰羟化酶活性。在目标3中，我们将确定pVHL如何调节EGLN 3和BIM（EL）的表达和功能。我们还将测试我们的假设，pVHL在营养因子剥夺诱导的细胞死亡中起促凋亡作用。这些研究将进一步加深我们对发育过程中导致神经元死亡的机制的理解，以及在神经系统疾病如中风、脊髓损伤和其他神经退行性疾病中，营养因子剥夺有助于神经元的损失。神经营养因子剥夺有助于人脑疾病和障碍如中风、脊髓损伤、神经元损伤和其他神经退行性疾病中神经元的损失和功能障碍。周围神经病和神经变性疾病。这个项目将描述新的机制，调节神经营养因子剥夺引起的细胞死亡。从该项目中获得的信息可能有助于确定旨在预防神经元损失的治疗的新靶点。神经营养因子剥夺导致人脑疾病中神经元的丧失和功能障碍， 例如中风、脊髓损伤、周围神经病变和神经变性疾病。这 该项目将描述由神经营养因子剥夺引起的调节细胞死亡的新机制。 从该项目中获得的信息可能有助于确定旨在预防神经元损伤的治疗的新靶点。 损失

DESCRIPTION (provided by applicant): Cell death is widespread during the development of the nervous system, where it helps to ensure that the proper number and types of connections are formed between neurons and their targets. During development, cell death occurs when neurons fail to receive adequate survival signals from trophic factors such as nerve growth factor (NGF). Accumulating evidence suggests that trophic factor deprivation-induced death also contributes to human neuronal disorders and degenerative diseases. We previously identified the prolyl hydroxylase EGLN3 as a mediator of NGF deprivation-induced cell death. EGLN3 is one of a small family of prolyl hydroxylases that function key as oxygen-dependent regulators of the transcription factor hypoxia- inducible factor (HIF). Besides regulating HIF, recent reports suggest that EGLNs have HIF-independent functions. We have obtained preliminary evidence for a novel interaction between EGLN3 and the pro- apoptotic Bcl-2 family protein BIM(EL). Moreover, we have found that EGLN3 and BIM(EL) each interact with the von Hippel-Lindau protein (pVHL), a subunit of the E3 ubiquitin ligase that targets HIF for degradation. Here we will test the hypothesis that EGLN3, pVHL, and BIM(EL) function coordinately to regulate trophic factor deprivation-induced cell death. In Aim 1 we will determine the functional relationship between EGLN3 and BIM(EL) during NGF deprivation-induced cell death using a combination of over-expression and gene knock-out and knock-down approaches. Aim 2 addresses the biochemical significance of the EGLN3/BIM(EL) interaction. Experiments will characterize how the interaction between EGLN3 and BIM(EL) influences the pro- apoptotic function of BIM(EL) and/or the prolyl hydroxylase activity of EGLN3. In Aim 3, we will determine how pVHL regulates EGLN3 and BIM(EL) expression and function. We will also test our hypothesis that pVHL plays a pro-apoptotic role in trophic factor deprivation-induced cell death. These studies should further our understanding of the mechanisms that lead to neuronal death during development, and in nervous systems disorders such as stroke, spinal cord injury, and other neurodegenerative conditions in which trophic factor deprivation contributes to the loss of neurons.Neurotrophic factor deprivation contributes to the loss and dysfunction of neurons in human brain diseases and disorders such as stroke, spinal cord injury, peripheral neuropathy, and neurodegenerative disease. This project will characterize new mechanisms that regulate cell death caused by neurotrophic factor deprivation. Information gained from this project may help identify new targets for therapies aimed at preventing neuronal loss. Neurotrophic factor deprivation contributes to the loss and dysfunction of neurons in human brain diseases and disorders such as stroke, spinal cord injury, peripheral neuropathy, and neurodegenerative disease. This project will characterize new mechanisms that regulate cell death caused by neurotrophic factor deprivation. Information gained from this project may help identify new targets for therapies aimed at preventing neuronal loss.