INVESTIGATION OF THE ROLE OF SMN IN NEURONAL APOPTOSIS

SMN 在神经元凋亡中的作用研究

• 批准号: 6024691
• 负责人: DOUGLAS A KERR
• 金额: $ 10.08万
• 依托单位: JOHNS HOPKINS UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 1999
• 资助国家: 美国
• 起止时间: 1999-09-30 至 2002-08-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6024691
• 关键词: RNA splicing; Sindbis virus; apoptosis; cysteine endopeptidases; disease /disorder model; enzyme activity; genetically modified animals; laboratory mouse; laboratory rat; motor neurons; neural growth associated protein; neuroregulation; neurotropic virus; precursor mRNA; protein structure function; site directed mutagenesis; superoxide dismutase; transfection /expression vector

The goal of this proposal is to investigate the role of the Survival of Motor Neuron (SMN) protein in neuronal apoptosis. We have generated a Sindbis virus model system to investigate the role of Survival of Motor Neuron (SMN) protein in neuronal apoptosis in vivo. Sindbis virus is a neuronotropic virus that causes a severe infection in young mice characterized by neuronal apoptosis. In this model system, the virus serves both as a cell death stimulus and as a vector expressing SMN specifically in CNS neurons. Preliminary studies reveal the first evidence that SMN modulates the susceptibility of neurons to programmed cell death in vivo and we propose that this function underlies the neuronal death seen in patients with Spinal Muscular Atrophy (SMA). Not only does SMN protein protect neurons from apoptotic death, but naturally occurring mutant SMN proteins found in SMA patients accelerate neuronal death, causing increased mortality in mice. Further data suggest a model in which SMN is cleaved by Caspase-6 in neurons undergoing apoptosis, creating a truncated protein that resembles a truncated SMN protein found in a patient with SMA, and that this truncated protein may have pro-death function. We now propose to further this analysis to determine the mechanisms that underlie the regulation of apoptosis by SMN. We will employ the Sindbis virus vector model system for in vivo gene delivery and induction of neuronal apoptosis. Domains of SMN critical for both pro- and anti-apoptotic functions will be investigated, and the identity of the SMN cleavage product created during apoptosis will be determined. The role of Caspase-6 in cleavage of SMN in vivo and whether suppression of this cleavage modulates the ability of SMN to protect neurons from apoptosis will be examined. Investigations will examine potential mechanisms that underlie the ability of SMN to modulate neuronal apoptosis, including whether SMN derivatives modulate differential splicing of selected pre-mRNA transcripts during apoptosis. Finally, a transgenic mouse approach will be employed to further the mechanistic exploration of SMN's ability to alter neuronal survival, and to determine the role of SMN in neural development and in mediating cellular responses to apoptotic stimuli.

该建议的目的是研究运动神经元（SMN）蛋白在神经元凋亡中的生存作用。 我们已经生成了一个信德氏病毒模型系统，以研究运动神经元（SMN）蛋白在体内神经元凋亡中的生存作用。 信德氏病毒是一种神经型病毒，在以神经元凋亡为特征的年轻小鼠中引起严重的感染。 在此模型系统中，该病毒既用作细胞死亡刺激，又是在CNS神经元中专门表达SMN的载体。 初步研究表明，第一个证据表明，SMN调节了神经元对体内编程细胞死亡的敏感性，我们建议该功能是脊柱肌肉萎缩（SMA）患者中神经元死亡的基础。 SMN蛋白不仅可以保护神经元免受凋亡死亡的影响，而且在SMA患者中发现的自然存在的突变体SMN蛋白会加速神经元死亡，从而导致小鼠死亡率增加。进一步的数据表明，在经历凋亡的神经元中，caspase-6裂解SMN的模型，形成截断的蛋白质，类似于在SMA患者中发现的截断的SMN蛋白，并且该截断的蛋白质可能具有促侵袭功能。现在，我们建议进一步分析，以确定SMN调节凋亡的基础的机制。 我们将利用sindbis病毒载体模型系统进行体内基因递送和神经元凋亡的诱导。 将研究对促凋亡功能至关重要的SMN领域，并将确定在凋亡期间产生的SMN裂解产物的身份。 CASPASE-6在体内SMN裂解中的作用以及该裂解的抑制是否会调节SMN保护神经元免受凋亡的能力。 研究将研究SMN调节神经元凋亡能力的潜在机制，包括SMN衍生物是否调节凋亡过程中选定的MRNA转录物的差异剪接。 最后，将采用转基因小鼠方法来进一步探索SMN改变神经元存活能力，并确定SMN在神经发育中的作用以及介导细胞对凋亡刺激的反应。