SOD1/Bc1-2 Complex: A Role in Regulating Motor Neuron Cell Death

SOD1/Bc1-2 复合物：调节运动神经元细胞死亡的作用

基本信息

批准号：
7271129
负责人：
PIERA PASINELLI
金额：
$ 41.75万
依托单位：
THOMAS JEFFERSON UNIVERSITY
依托单位国家：
美国
项目类别：
财政年份：
2006
资助国家：
美国
起止时间：
2006-08-02 至 2010-01-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/7271129
关键词：
Affect Affinity Amyotrophic Lateral Sclerosis Antibodies Antioxidants Apoptosis Apoptotic Appearance B-Lymphocytes BH3 Domain Binding Biological Assay Brain Stem Cell Death Cell Fractionation Cell Survival Cells Cerebellum Cessation of life Co-Immunoprecipitations Complex Confocal Microscopy Correlation Studies Cultured Cells Cyclosporine Cytochromes Data Doctor of Philosophy Down-Regulation Drosophila genus Epitopes Exhibits Flow Cytometry Goals Hippocampus (Brain)Human Immunofluorescence Immunologic Immunoprecipitation In Vitro Injection of therapeutic agent Knockout Mice Link Liver Localized Longevity Mediating Mitochondria Modeling Modification Molecular Molecular Weight Motor Cortex Motor Neurons Mus Nature Nerve Degeneration Nerve Growth Factor 1 Nerve Growth Factor Pathway Neuroblastoma Neurons Oxidative Stress Pathogenesis Patients Peptides Physiological Production Property Proteins Rattus Regulation Reporting Research Personnel Resistance Role Series Specificity Spinal Spinal Cord Staurosporine Structure Superoxide Dismutase Swelling Testing Time Tissues Toxic effect Transgenic Organisms Translating Withdrawal copper zinc superoxide dismutase design expression vector gain of function in vitro Model in vivo mutant neuron apoptosis neuron loss novel research study

项目摘要

DESCRIPTION (provided by applicant): In this project we will define a role for copper-zinc superoxide dismutase (SOD1) in the regulation of cell survival and death. While the wild-type (WT) SOD1 is a pro-survival protein, amyotrophic lateral sclerosis (ALS)-linked SOD1 mutants are toxic both in vitro and in vivo. We recently found that both WT and mutant SOD1 interact with the anti-apoptotic protein Bcl-2. However, the nature of the mutant SOD1 binding with Bcl-2 differs from WT SOD1. Contrary to WT SOD1, mutant SOD1 specifically localizes to spinal cord mitochondria where it forms SDS-resistant high molecular weight aggregates that bind and entrap Bcl-2. (Pasinelli et al, 2004, Neuron 43: 19-30). These studies suggest a potentially novel function for WT SOD1 in regulating cell survival and death, and a novel, toxic gain-of-function for mutant SOD1. Thus, while WT SOD1 may protect against cell death through its interaction with Bcl-2, mutant SOD1 may become toxic by aberrantly binding to Bcl-2 and converting Bcl-2 into a toxic or non-functional protein. In support of this hypothesis, we now have preliminary data indicating that both WT and mutant SOD1 might require Bcl-2 to exert their anti-and-pro apoptotic function respectively. With the present proposal we intend to characterize the anti and pro-death function of WT and mutant SOD1 and their respective interactions with Bcl-2. The ultimate goal is to understand the mechanism(s) of mutant SOD1-mediated toxicity and to define a potential role for the mitochondrial mutant SOD1/Bcl-2 complex in ALS pathogenesis. The specific aims are: 1) (A) To determine whether WT SOD1 pro-survival activity depends on its binding to Bcl-2, and (B) to determine whether mutant SOD 1-mediated toxicity depends on the aberrant interaction with Bcl-2. 2) (A) To identify the region(s) in SOD1 essential for the binding with Bcl-2, and (B) to study the difference in binding strength between WT SOD1 and Bcl-2 and mutant SOD1 and Bcl-2. 3) (A) To determine whether Bcl-2 undergoes conformational modifications upon binding with mutant SOD1 and.(B) to test the potential benefit of Bcl-2 and SOD1 like-peptides that abolish binding between Bcl-2 and mutant SOD1 on our cell culture model of mutant SOD1-linked ALS. 4) To determine whether Bcl-2 mediates mutant SOD1 mitochondrial translocation. 5) To study the correlation between mutant SOD1/Bcl-2-containing aggregates and ALS using transgenic ALS mice and patients.

描述（由申请人提供）：在此项目中，我们将定义铜锌超氧化物歧化酶（SOD1）在细胞存活和死亡调节中的作用。虽然野生型SOD1是一种蛋白质蛋白，但肌萎缩性侧索硬化症（ALS）与SOD1突变体在体外和体内都有毒性。我们最近发现WT和突变体SOD1都与抗凋亡蛋白Bcl-2相互作用。但是，与Bcl-2结合的突变体SOD1的性质与WT SOD1不同。与WT SOD1相反，突变体SOD1专门定位于脊髓线粒体，在该线粒体中形成了耐SDS的高分子量聚集体，该高分子量聚集体结合并置入BCl-2。（Pasinelli等，2004，Neuron 43：19-30）。这些研究表明，WT SOD1在调节细胞存活和死亡方面具有潜在的新功能，以及一种新型的突变体SOD1功能获得的功能。因此，尽管WT SOD1可以通过与Bcl-2相互作用来预防细胞死亡，但突变体SOD1可能会因与Bcl-2的异常结合并将Bcl-2转化为有毒或非功能性蛋白质而变得有毒。为了支持这一假设，我们现在拥有初步数据，表明WT和突变体SOD1可能需要Bcl-2分别发挥其抗凋亡功能。通过本建议，我们打算表征WT和突变体SOD1的抗死功能以及它们与Bcl-2的相互作用。最终目标是了解突变体SOD1介导的毒性的机制，并定义ALS发病机理中线粒体突变体SOD1/BCL-2复合物的潜在作用。具体目的是：1）（a）确定WT SOD1促生物活性是否取决于其与Bcl-2的结合，并且（b）确定突变SOD 1介导的毒性是否取决于与Bcl-2的异常相互作用。 2）（a）确定与Bcl-2结合所必需的SOD1中的区域，以及（b）研究WT SOD1和Bcl-2和突变体SOD1和BCL-2之间结合强度的差异。 3）（a）确定Bcl-2在与突变体SOD1和。（b）结合后是否经历构象修饰，以测试Bcl-2和SOD1类似肽的潜在益处，这些肽在我们的细胞培养模型中取消了Bcl-2和突变体Sod1之间的结合突变SOD1-SOD1链接的ALS。 4）确定Bcl-2是否介导了突变体SOD1线粒体易位。 5）研究使用转基因ALS小鼠和患者的含有突变体SOD1/Bcl-2的聚集体和ALS之间的相关性。