Understanding Mechanisms Underlying Drosophila Neurofibromatosis-1 Defects

描述(申请人提供)：神经纤维瘤病1型(NF1)是一种高度可变和不可预测的疾病，其严重程度似乎由未知的修饰基因决定。NF1基因编码神经纤维蛋白，其唯一公认的功能是作为RAS的负调节因子。除了良性肿瘤和恶性肿瘤，NF1患者还患有几种鲜为人知的非肿瘤症状，包括全面的生长缺陷和学习问题。果蝇NF1模型的表型与这些非肿瘤症状相似。值得注意的是，大多数果蝇的NF1表型不容易通过减弱RAS信号而改变，但可以通过cAMP依赖的蛋白激酶A(PKA)途径增加信号来恢复。这使得确定NF1如何影响cAMP信号成为NF1研究的中心问题。我们的结果表明，果蝇NF1突变体的总体生长缺陷反映了对特定幼虫神经元中RasGAP活性的非细胞自主需求。我们最初认为，这种原发的RAS信号缺陷反过来会导致继发性激素缺陷，这可以通过增加cAMP来缓解。新的证据现在提出了另一种双功能假说，即NF1不仅在神经内分泌细胞中需要调节RAS和一种或多种激素的产生，而且在激素反应细胞中也需要有效地将激素受体偶联到腺苷环化酶。我们提出了一种遗传和生化方法的组合来检验这一假说的优点，这可能解释了为什么我们和其他人对NF1在表型拯救中必不可少的功能得出了相互矛盾的结论。我们的新结果表明，神经内分泌Ret酪氨酸激酶、腺苷环化酶偶联的神经肽受体及其配体是NF1大小缺陷的遗传抑制因子。我们提出了四套相互关联的遗传和生化实验来跟进这些结果，并实现我们的总体目标，即提供NF1缺陷潜在机制的分子描述。与公共卫生相关的神经纤维瘤病1型(NF1)是人类最常见的遗传病之一。它的发病率估计为每3000人中就有1人，这表明仅在美国就可能有10万名NF1患者。在这个项目中，我们将结合使用遗传模式生物--黑腹果蝇的生化、分子生物学和遗传学方法，以确定导致NF1缺陷的机制。更好地了解这些机制是NF1合理治疗道路上的必要步骤。

DESCRIPTION (provided by applicant): Neurofibromatosis type 1 (NF1) is a highly variable and unpredictable disease, the severity of which appears to be determined by as yet unknown modifier genes. The NF1 gene encodes neurofibromin, whose only agreed upon function is to serve as a negative regulator of Ras. Beyond benign and malignant tumors, NF1 patients also suffer from several less well understood non-tumor symptoms, including an overall growth deficiency and learning problems. A Drosophila NF1 model displays phenotypes resembling these non-tumor symptoms. Strikingly, most Drosophila NF1 phenotypes are not readily modified by attenuating Ras signaling, but are restored by increasing signaling through the cAMP-dependent protein kinase A (PKA) pathway. This has made determining how NF1 affects cAMP signaling a central issue in NF1 research. Our results indicate that the overall growth defect of Drosophila NF1 mutants reflects a non-cell-autonomous requirement for RasGAP activity in specific larval neurons. We initially suggested that this primary Ras signaling defect in turn causes a secondary hormonal defect, which is mitigated by increasing cAMP. New evidence now suggests an alternative dual-function hypothesis, which states that NF1 is not just required in neuroendocrine cells to regulate Ras and the production of one or more hormones, but also in hormone responding cells to efficiently couple hormone receptors to adenylyl cyclase. We propose a combination of genetic and biochemical approaches to test the merits of this hypothesis, which may explain why we and others have reached contradictory conclusions about NF1 functions essential for phenotypic rescue. Our new results have implicated the neuroendocrine Ret tyrosine kinase, and an adenylyl cyclase-coupled neuropeptide receptor and its ligand as genetic suppressors of the NF1 size defect. We propose four interconnected sets of genetic and biochemical experiments to follow up on these results, and to achieve our overall goal of providing a molecular description of mechanisms underlying NF1 defects. PUBLIC HEALTH RELEVANCE Neurofibromatosis type 1 (NF1) is among the most common genetic diseases of man. Its estimated incidence of 1 in 3,000 suggests there may be 100,000 NF1 patients in the US alone. In this project we will use a combination of biochemical, molecular biological, and genetic approaches in the genetic model organism, Drosophila melanogaster, to identify mechanisms responsible for NF1 defects A better understanding of these mechanisms is a required step on the road to rational therapy for NF1.