Dynamic Regulation of Ras via the NF1 tumor Suppressor

描述(由申请人提供)：I型神经纤维瘤是一种流行的家族性癌症综合征，全世界每3500人中就有一人受到影响。NF1肿瘤抑制基因功能丧失突变是该病的基础。NF1编码的蛋白，神经纤维素，已被证明是一种Ras-GTP酶激活蛋白(GAP)；然而，关于它的活性是如何正常调节的，以及它在控制RAS信号通路中的确切作用，人们知之甚少。我们最近发现了第一个已知的调节神经纤维蛋白的机制。我们发现，在生长因子处理后，它被蛋白酶体迅速降解，随后被重新上调以关闭RAS。在我们未发表的研究中，我们进一步观察到重新合成的蛋白质是磷酸化的。由于神经纤维蛋白的磷酸化恰好发生在RAS失活时，我们推测，磷酸化可能增强了神经纤维蛋白的功能，从而强烈地终止了RAS信号。这一假说得到了各种遗传和生化证据的支持。我们的数据还表明，ERK是调节激酶。因此，该事件可能代表RAS信号通路正确终止所需的正常负反馈环。在第一个目标中，我们将确定神经纤维蛋白磷酸化的功能后果，并确定ERK是否是体内的调节性激酶。在第二个目标中，我们将在小鼠模型系统和表达磷酸化突变蛋白的细胞系中确定神经纤维蛋白磷酸化的生物学意义。最后，我们还鉴定了一种新的神经纤维蛋白相互作用蛋白。因为它是在神经纤维蛋白被磷酸化的条件下分离出来的，所以我们将在Aim 3中确定磷酸化在介导这种相互作用中的作用。无论如何，我们将从分子上剖析这种相互作用，并测试它对神经纤维蛋白功能的参与。重要的是，这一研究途径可能使我们能够确定十多年来第一个新的NF1功能。总而言之，这些目标将大大增加我们对NF1肿瘤抑制因子的调节和功能的了解。它们还将极大地有助于我们理解调节适当RAS衰减的机制以及这种微调调节的生物学重要性。因此，这些发现可能最终会影响未来对NF1和非NF1相关肿瘤的治疗策略。

DESCRIPTION (provided by applicant): Neurofibromatis type I is a prevalent familial cancer syndrome affecting 1 in 3500 individuals worldwide. Loss-of-function mutations in NF1 tumor suppressor gene underlie the disease. The NF1-encoded protein, neurofibromin, has been shown to function as a Ras-GTPase activating protein (GAP); however, little is known about how its activity is normally regulated or its precise role in controlling Ras signaling pathways. We have recently identified the first mechanism known to regulate neurofibromin. We found that it is rapidly degraded by the proteasome following growth factor treatment and is subsequently re-elevated to turn off Ras. In our unpublished studies we have further observed that the re-synthesized protein is phosphorylated. Because neurofibromin phosphorylation occurs precisely as Ras becomes inactivated, we hypothesize that phosphorylation may enhance neurofibromin function to acutely terminate the Ras signal. This hypothesis is supported by a variety of genetic and biochemical evidence. Our data also suggest that ERK is the regulatory kinase. Therefore, this event may represent a normal negative feedback loop that is required for proper termination of Ras signaling pathways. In the first aim we will determine the functional consequences of neurofibromin phosphorylation and firmly establish whether ERK is the regulatory kinase in vivo. In the second aim we will determine the biological significance of neurofibromin phosphorylation in a mouse model system and in cell lines expressing the phospho-mutant protein. Finally, we have also identified a novel neurofibromin interacting protein. Because it was isolated under conditions in which neurofibromin is phosphorylated, we will determine the role of phosphorylation in mediating this interaction in Aim 3. Regardless, we will molecularly dissect this interaction and test its involvement in neurofibromin function. Importantly, this avenue of investigation may enable us to identify the first new function for NF 1 in over a decade. Collectively, these aims will serve to dramatically increase our knowledge about the regulation and function of the NF1 tumor suppressor. They will also greatly contribute to our understanding of the mechanisms governing appropriate Ras attenuation and the biological importance of this fine-tuned regulation. As a result these findings may ultimately impact future therapeutic strategies for NF1 and non-NF1 related tumors.