Proteomic Dissection of M-channel Phosphorylation

M通道磷酸化的蛋白质组学解析

基本信息

批准号：
6658082
负责人：
EDWARD C COOPER
金额：
$ 19.81万
依托单位：
UNIVERSITY OF PENNSYLVANIA
依托单位国家：
美国
项目类别：
财政年份：
2001
资助国家：
美国
起止时间：
2001-09-15 至 2005-08-31
项目状态：
已结题

项目摘要

DESCRIPTION (provided by applicant): Careful study of genes responsible for rare Mendelian forms of human neurological disorders is an powerful approach for gaining insight into the causes, treatment, and potential cure for common, related diseases. The KCNQ genes were recently discovered as the result of the search for mutant genes causing Benign Neonatal Familial Convulsions, an autosomal dominant epileptic syndrome associated with seizures in infancy and throughout life. KCNQ genes encode voltage-dependent potassium channels called M-channels. M-channels regulate neuronal excitability by serving as effectors for neurotransmitter receptors and intracellular signaling pathways. Determining how these receptors and pathways modulate M-channels will enhance our ability to exploit M-channels as therapeutic targets in conditions involving excessive excitability or alterations and imbalances in modulatory neurotransmission, such as epilepsy, pain syndromes, and Alzheimer's disease. Because M-channels are large, oligomeric proteins and appear to be regulated by several distinct intracellular pathways, a novel, comprehensive approach to understanding their regulatory mechanisms is warranted. This proposed research approach exploits new instrumentation and analytical methods in mass spectrometry to identify sites of in vivo phosphorylation on KCNQ subunits. A combination of electrophysiological and cellular and molecular biological methods will then be used to determine the consequences of phosphorylation at the identified sites. The proposal involves a new collaboration between individuals with expertise in the clinical and genetic aspects of epilepsy, ion channel physiology, and proteomics. In addition to addressing specific questions concerning M-channel regulation, the proposed research will contribute to the development of mass spectrometric methods and the adaptation of these methods to the study of signaling proteins that are large, complex, and expressed either transiently, locally, or at low abundance in the brain. Because such signaling proteins play essential roles in brain development and in the maintenance of normal brain structure and activity, the proposed research may result in enhanced methodologies of broad usefulness for research into anomalous neurodevelopment, degeneration, and injury.

描述（由申请人提供）：仔细研究负责的基因罕见的孟德尔形式的人类神经系统疾病是一种有效的方法深入了解常见问题的原因、治疗方法和潜在治愈方法，相关疾病。 KCNQ 基因是最近被发现的寻找引起良性新生儿家族性惊厥的突变基因与婴儿期癫痫发作相关的常染色体显性癫痫综合征一生。 KCNQ 基因编码电压依赖性钾通道，称为 M通道。 M 通道通过充当效应器来调节神经元兴奋性用于神经递质受体和细胞内信号传导途径。确定这些受体和通路如何调节 M 通道将增强我们在某些情况下利用 M 通道作为治疗靶点的能力涉及过度兴奋或调节的改变和不平衡神经传递，例如癫痫、疼痛综合征和阿尔茨海默病。因为 M 通道是大的寡聚蛋白，并且似乎受几种不同的细胞内途径，一种新颖的综合方法了解其监管机制是有必要的。这项拟议的研究方法大规模利用新的仪器和分析方法光谱法鉴定 KCNQ 亚基的体内磷酸化位点。一个电生理学与细胞分子生物学的结合然后将使用方法来确定磷酸化的后果已确定的地点。该提案涉及双方之间的新合作具有癫痫临床和遗传方面专业知识的个人，离子通道生理学和蛋白质组学。除了解决具体问题有关 M 通道监管的问题，拟议的研究将有助于质谱方法的发展和适应这些方法可用于研究大型、复杂的信号蛋白，并在大脑中短暂地、局部地或以低丰度表达。因为此类信号蛋白在大脑发育和在维持正常的大脑结构和活动方面，建议研究可能会产生对研究具有广泛用途的增强方法导致神经发育异常、退化和损伤。