THE ROLE OF AGRIN IN THE DEVELOPMENT OF CENTRAL NEURONS

描述(申请人提供)：在过去的二十年里，对哺乳动物中枢神经系统中突触发生的研究引起了极大的关注。许多这些研究都是基于这样一种信念，即通过更好地了解中枢神经元突触形成的基本机制，可以诊断和预防与几种神经退行性疾病相关的突触丢失。尽管这些机制尚不完全清楚，但人们很容易推测，在神经肌肉连接形成中发挥关键作用的蛋白质，如集聚蛋白，可能也在神经元之间突触接触的形成中发挥重要作用。最近，我们发现，集聚蛋白对海马神经元中轴突和树突的伸长以及突触的形成具有不同的调节作用。然而，这些神经元中的集聚蛋白受体(S)尚未被确定。本文提出的实验旨在验证以下假设：ROR蛋白是两种被认为是“孤儿受体”的酪氨酸激酶受体，可以介导集聚蛋白对中枢神经元早期发育的影响。我们将结合西方印迹分析、免疫细胞化学、RT-PCR、零突变的产生、结合分析和免疫沉淀等技术来分析：1)orr1和orr2在中枢神经元中的表达和定位；2)这些酪氨酸激酶受体在轴突延长和突触形成中的作用；以及3)orr1和orr2在集聚蛋白信号通路中的作用。

DESCRIPTION (provided by applicant): In the past two decades, a great deal of attention has been directed to the study of synaptogenesis in the mammalian central nervous system. Many of these studies are driven by the belief that the diagnosis and the prevention of synapse loss associated with several neurodegenerative diseases can be achieved through a better understanding of the basic mechanisms of synapse formation in central neurons. Although these mechanisms are not completely understood, it is tempting to speculate that proteins, such as agrin, that play a key role in the formation of the neuromuscular junction might also play an important role in the formation of synaptic contacts between neurons. Recently, we have shown that agrin differentially regulates the rates of axonal and dendritic elongation as well as synapse formation in hippocampal neurons. However, the agrin receptor(s) has yet to be identified in these neurons. The experiments proposed here are intended to test the following hypothesis: ror proteins, two tyrosine kinase receptors considered "orphan receptors", could mediate the effects of agrin on early stages of development in central neurons. A combination of techniques including: Western blot analysis, immunocytochemistry, RT-PCR, the generation of null mutations, binding assays, and immunoprecipitation will be used to analyze: 1) the pattern of expression and localization of ror 1 and ror 2 in central neurons; 2) the role of these tyrosine kinase receptors in neurite elongation and synapse formation; and 3) the participation of ror 1 and ror 2 in the agrin-signaling pathway.