Functional analysis of variant BDNF (Val66Met)

描述（由申请人提供）：脑源性神经营养因子（BDNF）在脊椎动物神经系统发育和功能中起关键作用。最近，BDNF基因中的单核苷酸多态性（Val 66 Met）导致在位置66处的前结构域从缬氨酸（瓦尔）替换为甲硫氨酸（Met），已经显示在人类中导致海马依赖性记忆障碍和对神经精神障碍的易感性。这种BDNF多态性代表了第一个与临床病理学相关的神经营养因子的改变。关于BDNF（BDNF Met）功能改变的分子机制知之甚少。当在海马神经元中过表达时，BDNFMet具有减少的活性依赖性分泌，表明BDNF前结构域中存在特异性信号，该信号是有效的BDNF运输到受调节的分泌途径所需的。初步研究表明，BDNF Met异常参与高度专业化的生化机制，调节BDNF运输到分泌途径，这是BDNF的生物反应的关键决定因素的假设。拟议的研究旨在确定调节异常BDNFMet运输的特定蛋白质，并检查海马结构和功能的体内后果。本提案中的实验将主要利用表达表位标记形式的变体BDNF（BDNF 9 e）的新型转基因敲入小鼠，以能够评估内源性表达条件下的BDNF Met运输事件并分析对海马结构和功能的体内后果。拟议研究的具体目的是：1）确定内源性表达BDNFMet的神经元中的运输缺陷和功能后果，2）鉴定参与异常BDNFMet运输的蛋白质，3）确定BDNFMet对海马功能的体内后果。这些研究将有助于对神经元中异常BDNFMet运输的机制的基本分子理解，并直接解决这种变体BDNF对海马功能的生理相关性。

DESCRIPTION (provided by applicant): Brain derived neurotrophic factor (BDNF) plays critical roles in vertebrate nervous system development and function. Recently, a single nucleotide polymorphism (Val66Met) in the BDNF gene leading to a prodomain substitution at position 66 from a valine (Val) to methionme (Met) has been shown to lead in humans to hippocampal dependent memory impairments and susceptibility to neuropsychiatric disorders. This BDNF polymorphism represents the first alteration in a neurotrophin that has been linked to clinical pathology. Less is known about the molecular mechanisms underlying altered variant BDNF (BDNFMet) functioning. When overexpressed in hippocampal neurons, BDNFMet has reduced activity dependent secretion, suggesting the presence of a specific signal in the BDNF prodomain that is required for efficient BDNF trafficking to the regulated secretory pathway. Preliminary studies suggest the hypothesis that BDNFMet aberrantly engages the highly specialized biochemical mechanisms that regulate BDNF trafficking to secretory pathways, which are critical determinants of BDNF's biological responses. The proposed studies are designed to identify specific proteins that regulate aberrant BDNFMet trafficking, and to examine the in vivo consequences on hippocampal structure and function. Experiments in this proposal will primarily utilize a novel transgenic knock-in mouse expressing an epitope tagged version of variant BDNF (BDNF 9 e) to enable assessment of BDNFMet trafficking events under endogenously expressed conditions and analysis of in vivo consequences on hippocampal structure and function. The Specific Aims of the proposed studies are to 1) define the trafficking defect and functional consequences in neurons endogenously expressing BDNFMet, 2) identify proteins that are involved in aberrant BDNFMet trafficking, and 3) determine the in vivo consequences of BDNFMet on hippocampal function. These studies will contribute to a fundamental molecular understanding of the mechanisms that underlie aberrant BDNFMet trafficking in neurons, and directly address the physiological relevance of this variant BDNF on hippocampal function.