DROSOPHILA MODELS FOR STUDYING FUNCTION OF THE NF1 GENE

该建议利用果蝇模型研究肿瘤抑制基因神经纤维瘤病1（NF 1）的机制和功能。 我们特别感兴趣的是它在调节G蛋白依赖的cAMP通路激活中的作用。 NF 1蛋白含有与Ras特异性GAPS的催化结构域同源的片段，并且其突变引起常见的神经遗传性疾病，其通过神经纤维瘤和包括特定学习缺陷的其他症状来鉴定。 我们对果蝇NF 1突变体的研究表明，NF 1除了作为Ras-GAP外，还参与调控芜菁甘蓝基因编码的腺苷酸环化酶（AC）的G蛋白依赖性激活。这种NF 1调节的cAMP通路对于介导神经肌肉接头处的神经肽反应、体型发育以及学习和记忆至关重要。 然而，目前尚不清楚NF 1如何调节AC活性，NF 1调节的AC通路如何有助于学习和记忆，以及这种NF 1调节机制是否也在脊椎动物中起作用。 该提案旨在调查这些问题。 它包括两个具体目标。1)在人类NF 1患者中鉴定的18个点突变将用于定义NF 1的哪些部分对于AC活性的调节是重要的以及Ras是否参与这种调节。 然后将小鼠Nf 1基因敲除突变体用于检测Nf 1是否可以调节脊椎动物AC活性。2)生物化学和电生理学分析将被引导，以确定神经递质激活NF 1调节的AC活动在苍蝇的大脑和神经递质激活NF 1调节的AC通路，以调节膜电流在确定的蘑菇体神经元。 这些神经元被认为参与了被NF 1突变破坏的联想学习过程。 我们希望这项研究能为深入了解NF 1的功能和发病机制提供新的线索。

This proposal utilizes Drosophila models for investigating mechanisms and functions of the tumor suppressor gene neurofibromatosis 1 (NF1). We are particularly interested in its role in regulation of G-protein-dependent activation of the cAMP pathway. NF1 protein contains a fragment homologous to catalytic domain of the Ras-specific GAPS and its mutations cause a common neurogenetic disorder that is identified by neurofibromas and other symptoms including specific learning defects. Our study of Drosophila NF1 mutants has revealed that in addition to as a Ras-GAP, NF1 is involved in regulation of G-protein dependent activation of adenylyl cyclase (AC) encoded by rutabaga gene. This NF1-regulated cAMP pathway is crucial for mediating a neuropeptide response at neuromuscular junction, body size development, and learning and memory. However, it is unclear how NF1 regulates AC activity, how the NF1-regulated AC pathway contributes to learning and memory, and whether such NF1-regulated mechanism also works in vertebrates. The proposal is intended to investigate these problems. It includes two specific aims. 1) Eighteen point mutations identified in human NF1 patients will be used to define which parts of NF1 is important for regulation of AC activity and whether Ras is involved in such regulation. Then mouse Nf1 knockout mutants will be used to assay whether NF1 can regulate AC activity in vertebrates. 2) Biochemical and electrophysiological analyses will be directed to determine neurotransmitters that activate NF1-regulated AC activity in the fly brain and neurotransmitters that activate the NF1-regulated AC pathway to modulate membrane currents in identified mushroom body neurons. These neurons have been suggested to be involved in the associative learning process that are disrupted by NF1 mutations. We expect that the proposed research will lead to new insights into NF1 function and provide new clues for understanding pathogenesis of NF1.