Analysis of Cytoplasmic Prion Protein Toxicity

细胞质朊病毒蛋白毒性分析

• 批准号: 7113660
• 负责人: WALKER S JACKSON
• 金额: $ 5.04万
• 依托单位: WHITEHEAD INSTITUTE FOR BIOMEDICAL RES
• 依托单位国家: 美国
• 财政年份: 2004
• 资助国家: 美国
• 起止时间: 2004-09-01 至 2007-08-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7113660
• 关键词: cell component structure /function; cell line; cytoplasm; cytotoxicity; embryonic stem cell; endoplasmic reticulum; family genetics; fibroblasts; gene expression; gene targeting; glia; intracellular transport; laboratory mouse; molecular pathology; mutant; neurons; postdoctoral investigator; prions; proteasome; protein biosynthesis; protein degradation; protein localization; protein quantitation /detection; protein structure function; spongiform encephalopathy

DESCRIPTION (provided by applicant): Evidence produced in the mid 1990's suggests that bovine spongiform encephalopathy (commonly known as mad cow disease) was probably transmitted to humans. The agents widely believed to be responsible for these diseases are abnormally shapened prion proteins. Along with an infectious route, humans can also develop prion diseases either sporadically or by expressing a mutant prion protein (penetrance approximately 100%). Recent work points to the cytosol as being the site of toxicity of prion proteins. Mutant prion proteins may be highly toxic because they have an increased probability of misfolding and therefore mislocalizing to the cytosol. To study the mechanism of increased toxicity of the mutant prion proteins and how it relates to cytosolic prion toxicity, ES cells will be genetically altered to express mutant prion proteins directly into the cytosol. These cell lines will be differentiated into neurons, gila, or fibroblasts and used to study the effects these mutations have on subcellular traffciking, degradation, and toxicity of PrP. The gene-targeting approach will allow for a high probability of equivalent expression of these constructs at the mRNA level. The selection of ES cells as a model system will permit efficient transition of cell culture experiments into mice.

描述（由申请人提供）：20世纪90年代中期产生的证据表明牛海绵状脑病（俗称疯牛病）可能传播给人类。人们普遍认为导致这些疾病的因素是形状异常的朊病毒蛋白。除了感染途径外，人类还可能偶尔或通过表达突变朊病毒蛋白（外显率约为 100%）而患上朊病毒疾病。最近的研究表明细胞质是朊病毒蛋白的毒性部位。突变的朊病毒蛋白可能具有剧毒，因为它们错误折叠的可能性增加，因此错误定位到细胞质中。为了研究突变型朊病毒蛋白毒性增加的机制及其与胞质朊病毒毒性的关系，将对胚胎干细胞进行基因改造，将突变型朊病毒蛋白直接表达到胞浆中。这些细胞系将分化为神经元、吉拉细胞或成纤维细胞，并用于研究这些突变对 PrP 的亚细胞运输、降解和毒性的影响。基因靶向方法将允许这些构建体在 mRNA 水平上有很高的等效表达可能性。选择 ES 细胞作为模型系统将允许细胞培养实验有效地转移到小鼠身上。