Understanding Mechanisms Underlying Drosophila Neurofibromatosis-1 Defects

描述（由申请人提供）：1型神经纤维瘤病（NF1）是一种高度可变和不可预测的疾病，其严重程度似乎由尚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。除了良性和恶性肿瘤外，NF1患者还会出现一些鲜为人知的非肿瘤症状，包括整体生长缺陷和学习问题。果蝇NF1模型显示出类似这些非肿瘤症状的表型。引人注目的是，大多数果蝇NF1表型不容易通过减弱Ras信号来修饰，而是通过camp依赖性蛋白激酶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。我们的研究结果表明，果蝇NF1突变体的整体生长缺陷反映了特定幼虫神经元对RasGAP活性的非细胞自主需求。我们最初认为，这种原发性Ras信号缺陷反过来会导致继发性激素缺陷，这种缺陷可以通过增加cAMP来减轻。新的证据现在提出了另一种双重功能假说，即NF1不仅需要在神经内分泌细胞中调节Ras和一种或多种激素的产生，而且还需要在激素应答细胞中有效地将激素受体与腺苷酸环化酶偶联。我们提出结合遗传和生化方法来检验这一假设的优点，这可以解释为什么我们和其他人对NF1对表型拯救至关重要的功能得出了相互矛盾的结论。我们的新结果暗示神经内分泌Ret酪氨酸激酶和腺苷酸环化酶偶联神经肽受体及其配体是NF1大小缺陷的遗传抑制因子。我们提出了四组相互关联的遗传和生化实验来跟进这些结果，并实现我们提供NF1缺陷机制的分子描述的总体目标。1型神经纤维瘤病（NF1）是人类最常见的遗传性疾病之一。据估计，其发病率为3000分之一，这表明仅在美国就可能有10万名NF1患者。在这个项目中，我们将结合遗传模式生物——黑胃果蝇的生化、分子生物学和遗传学方法，来确定NF1缺陷的机制。更好地理解这些机制是NF1合理治疗的必要步骤。

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.