DBH as a Modifying Gene in Neurodegenerative Diseases

描述（由申请人提供）：申请人Cyrus Zabetian博士在耶鲁大学/ VACHS担任博士后研究员已有三年。明年，他将加入华盛顿大学的神经病学系，他未来的导师托马斯伯德博士和杰拉德谢伦贝格博士在那里建立了一个极好的神经遗传学研究项目。他的培训将包括参加实验室会议、研讨会、结构化课程和年度科学会议。他将成为具有临床和分子神经遗传学，儿茶酚胺生物化学和生物统计学专业知识的研究人员丰富的合作网络的一部分。Zabetian博士的长期计划是在五年内成为一名独立的实验室研究员，并继续积极参与患者护理和神经病学服务的住院医师培训。 在神经退行性疾病研究中，确定存活神经元代偿性变化的遗传机制有望改善诊断和治疗策略。本申请中提出的项目旨在确定新发现的启动子多态性（C-1021 T）是否影响DBH基因的调节，并在帕金森病（PD）中具有潜在的临床后果，并分为三个部分。第一部分的目的是评估C-1021 T等位基因的纯合性（与血浆DBH酶水平低相关）是否可预测PD患者交感神经功能衰竭的早期发作和更严重的症状。从400名临床PD患者中选择一组40名C或T等位基因纯合子受试者，并使用交感神经功能指数进行纵向评估。第二部分旨在确定C-1021 T是否与去甲肾上腺素能组织中DBH的表达密切相关。将分别使用蛋白质印迹和定量真实的时间RT-PCR，比较C或T等位基因纯合的死后人肾上腺髓质标本中DBH蛋白和mRNA的水平。第三部分将评估C-1021 T是否直接起作用。如果初步结果是有利的，则将产生对于T或C等位基因纯合的两个转基因小鼠系，其中内源小鼠DBH启动子的近端2kb被同源人序列替换。比较两个品系中DBH蛋白和儿茶酚胺的血浆和组织水平将检测每个等位基因对DBH表达的影响。

DESCRIPTION (provided by applicant): The applicant, Dr. Cyrus Zabetian, has spent the past three years as a postdoctoral fellow at Yale University/ VACHS. He will join the neurology faculty at the University of Washington next year where his future mentors, Drs. Thomas Bird and Gerard Schellenberg, have established a superb research program in neurogenetics. His training will include participation in laboratory meetings, seminars, structured courses, and annual scientific meetings. He will become part of a rich collaborative network of researchers with expertise in clinical and molecular neurogenetics, catecholamine biochemistry, and biostatistics. Dr. Zabetian's long-term plans are to become established as an independent laboratory investigator within five years, and remain actively involved in patient care and resident training on the neurology service. In neurodegenerative disease research, identifying genetic mechanisms underlying compensatory changes in surviving neurons promises to lead to improved strategies of diagnosis and treatment. The project proposed in this application seeks to determine if a newly discovered promoter polymorphism (C-1021T) influences regulation of the DBH gene with potential clinical consequences in Parkinson's disease (PD), and is divided into three parts. The goal of part I is to evaluate whether homozygosity for the T allele of C-1021 T, which is associated with low levels of plasma DBH enzyme, is predictive of an earlier onset and more severe symptoms of sympathetic failure in patients with PD. A group of forty subjects homozygous for either the C or T allele will be selected from a population of 400 clinic patients with PD and assessed longitudinally using indices of sympathetic function. Part II seeks to determine whether C-1021T strongly associates with DBH expression in noradrenergic tissues. Levels of DBH protein and mRNA will be compared in postmortem human adrenal medulla specimens, homozygous for either the C or T allele, using western blots and quantitative real time RT-PCR, respectively. Part III will assess whether C-1021T is directly functional. If preliminary results are favorable, two transgenic mouse lines homozygous for either the T or C allele will be created in which the proximal 2 kb of the endogenous mouse DBH promoter is replaced by homologous human sequence. Comparing plasma and tissue levels of DBH protein and catecholamines in the two lines will detect the effect of each allele on DBH expression.