Kinase/Phosphatase-mediated Mitochondrial Restructuring in Neuroprotection

激酶/磷酸酶介导的线粒体重构在神经保护中的作用

基本信息

批准号：
7389640
负责人：
STEFAN STRACK
金额：
$ 32.27万
依托单位：
UNIVERSITY OF IOWA
依托单位国家：
美国
项目类别：
财政年份：
2007
资助国家：
美国
起止时间：
2007-04-01 至 2012-03-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/7389640
关键词：
A kinase anchoring protein Abbreviations Adult Alternative Splicing Antibodies Apoptosis Apoptotic Attention Behavioral Biological Assay Brain Brain Injuries CAG repeat Calcium Cell Death Cells Chromosome Pairing Class Collaborations Complex Condition Confocal Microscopy Corpus striatum structure Cyclic AMP-Dependent Protein Kinases Cytochromes Data Development Dominant-Negative Mutation Event Figs - dietary Free Radicals Functional RNA Galactosidase Genes Genotype Glucose Glutamates Guanosine Triphosphate Guanosine Triphosphate Phosphohydrolases Hippocampus (Brain)Holoenzymes Homeostasis Hydrolysis Hypoglycemia Hypoglycemic Agents Hypoxia IGFBP2 gene In Vitro Injection of therapeutic agent Injury Intervention Ischemia Ischemic Stroke Knock-out Knockout Mice Length Localized Mediating Metabolic Middle Cerebral Artery Occlusion Mitochondria Modeling Molecular Morphology Mus Mutagenesis Nerve Degeneration Neurodegenerative Disorders Neuronal Injury Neurons Neuroprotective Agents Outer Mitochondrial Membrane Oxygen PC12 Cells Personal Satisfaction Phosphoric Monoester Hydrolases Phosphorylation Phosphorylation Site Phosphotransferases Plasmids Predisposition Process Protein Overexpression Protein phosphatase Proteins RNA Interference RNA Splicing Rattus Recombinant Proteins Recombinants Regulation Reporter Research Research Infrastructure Research Personnel Resistance Role Rotenone Shapes Signal Transduction Slice Spinocerebellar Ataxias Staging Stream Stroke Subfamily lentivirinae Synapses Testing Therapeutic Toxic effect Transient Cerebral Ischemia Variant Virus brain cell concept density deprivation follow-up in vivo inhibitor/antagonist killings loss of function neglect neuron apoptosis neuronal survival neuroprotection prevent pro-apoptotic protein programs response scaffold stroke therapy

项目摘要

DESCRIPTION (provided by PI): Ischemic stroke kills brain cells by causing massive depolarization and glutamate release, which in turn disrupts calcium homeostasis and generates free radicals. Mitochondria are central to these events, as well as to the wave of delayed cell death that follows the ischemic injury. Attention has focused on neuroprotective drugs that prevent the initial calcium overload, while interventions into later stages of neuronal damage have been largely neglected despite the promise of a wider treatment window. Protein phosphatase 2A (PP2A) is an essential and ubiquitous Ser/Thr phosphatase that predominately exists as a heterotrimer of two core subunits (a catalytic and a scaffolding subunit) complexed to a third, regulatory subunit Bp2 is an alternative splice variant of a PP2A regulatory subunit gene that is expressed only in the adult brain. A non-coding CAG repeat expansion in the B(3 gene causes the neurodegenerative disorder spinocerebellar ataxia type 12. The alternative N terminus of B(32 promotes translocation of PP2A to the outer mitochondrial membrane (OMM) to induce apoptosis in PC 12 cells (JBC 278:24976; JBC 280:27375), whereas interfering with endogenous mitochondrial PP2A protects neurons against ischemic insults in vitro. The proposal is centered on the observation that mitochondrial shape transitions underlie the pro- apoptotic activity of PP2A/Bp2, with overexpression of Bp2 fragmenting mitochondria and Bp2 silencing causing mitochondrial elongation in hippocampal neurons. Opposing the fission activity of the phosphatase is protein kinase A (PKA) anchored to the OMM via A-kinase anchoring protein D-AKAP1. Aim 1 test the hypothesis that OMM-localized PP2A and PKA control neuronal survival by reciprocally modulating mitochondrial fission and fusion, which in turn alters free radical and calcium homeostasis. Aim 2 explores the fission GTPase Drp1 as an effector substrate for outer-mitochondrial PP2A and PKA. In Aim 3, in vivo delivery of viruses expressing three kinds of PP2A/Bp2 antagonists is intended to provide proof-of-concept evidence that this neuron-specific pro-apoptotic protein is a promising target for stroke therapy. Finally, Aim 4 characterizes Bp2 null mice for mitochondrial changes and resistance to ischemic injury.

描述（由 PI 提供）：缺血性中风通过引起大量去极化和谷氨酸释放来杀死脑细胞，进而破坏钙稳态并产生自由基。线粒体是这些事件以及缺血性损伤后延迟细胞死亡波的核心。人们的注意力集中在防止最初钙超载的神经保护药物上，而尽管有望提供更宽的治疗窗口，但对神经元损伤后期的干预却在很大程度上被忽视了。蛋白磷酸酶 2A (PP2A) 是一种必需且普遍存在的 Ser/Thr 磷酸酶，主要以与第三个调节亚基复合的两个核心亚基（催化亚基和支架亚基）的异三聚体形式存在。Bp2 是 PP2A 调节亚基的替代剪接变体仅在成人大脑中表达的亚基基因。 B(3 基因中的非编码 CAG 重复扩增会导致 12 型脊髓小脑共济失调神经退行性疾病。B(32 的替代 N 末端促进 PP2A 易位至线粒体外膜 (OMM)，从而诱导 PC 12 细胞凋亡。 JBC 278:24976；JBC 280:27375)，而干扰内源性线粒体 PP2A 可以保护神经元该提案的重点是观察到线粒体形状转变是 PP2A/Bp2 促凋亡活性的基础，Bp2 的过度表达会导致线粒体断裂，而 Bp2 沉默会导致海马神经元的线粒体伸长。磷酸酶是通过 A 激酶锚定蛋白 D-AKAP1 锚定到 OMM 的蛋白激酶 A (PKA)。测试 OMM 定位的 PP2A 和 PKA 通过相互调节线粒体裂变和融合来控制神经元存活，进而改变自由基和钙稳态的假设。目标 2 探索裂变 GTPase Drp1 作为外线粒体 PP2A 和 PKA 的效应底物。在目标 3 中，体内递送表达三种 PP2A/Bp2 拮抗剂的病毒旨在提供概念验证证据，证明这种神经元特异性促凋亡蛋白是中风治疗的有希望的靶标。最后，Aim 4 描述了 Bp2 缺失小鼠的线粒体变化和对缺血性损伤的抵抗力。