Molecular mechanisms for lipid sensing by mTORC1

项目总结 细胞感知营养的分子机制在很大程度上仍不清楚，但它们的 阐明是我们理解正常和疾病状态下代谢调节的关键。在中心 是一种古老的蛋白激酶，被称为 雷帕霉素复合体1(MTORC1)。作为对营养等代谢输入的综合作用的反应， 生长因子、能量和氧气，mTORC1从细胞质转移到溶酶体表面， 在那里它的激酶功能被激活。越来越多的证据表明，mTORC1的异常激活 溶酶体可能是从癌症到2型糖尿病到其他疾病的驱动力 神经退行性变。因此，对mTORC1如何被激活以响应 营养素可以为这些疾病的新治疗策略指明方向。 目前的提案调查了mTORC1功能的一个中心方面，该功能到目前为止仍然存在 人们对它的研究不足，也就是对它感知血脂的能力知之甚少。我们将以我们最近的发现为基础 MTORC1在溶酶体上感觉到一种重要的脂质--胆固醇。使用创新的方法在细胞和 在体外，我们将阐述和阐明新发现的信号通路的关键方面。特别是，我们将 确定一)调节mTORC1的胆固醇池的细胞位置II)组成 胆固醇对mTORC1的作用及III)胆固醇诱导mTORC1的分子机制 向溶酶体表面募集。此外，我们将研究mTORC1是如何感知胆固醇的 依赖于Niemann-Pick C1蛋白，该蛋白的丢失会导致致命的代谢和神经退行性变 疾病。 最近，我们将通过创新和高度互补的方法来解决这些研究目标 在我们的实验室中进行了优化，包括对选定细胞器的脂质含量进行测量和有针对性的操作 结合基于重建的mTORC1调控分析。 我们的发现将影响目前对细胞脂质分子机制的理解 动态平衡，并将为在疾病环境中操纵mTORC1信号的新方法指明方向。

PROJECT SUMMARY The molecular mechanisms through which cells sense nutrients remain largely unknown, but their elucidation is key to our understanding of metabolic regulation both in normal and disease states. At the center of nutrient sensing and growth regulation is an ancient protein kinase known as the mechanistic Target of Rapamycin Complex 1 (mTORC1). In response to the combined action of metabolic inputs such as nutrients, growth factors, energy and oxygen, mTORC1 translocates from the cytoplasm to the surface of lysosomes, where its kinase function becomes activated. Accumulating evidence indicates that aberrant mTORC1 activation at the lysosome could be a driving force in diseases ranging from cancer to type-2 diabetes to neurodegeneration. Thus, a deep mechanistic understanding of how mTORC1 is activated in response to nutrients could point the way to novel therapeutic strategies in these diseases. The current proposal investigates a central aspect of mTORC1 function that has so far remained understudied and poorly understood, namely, its ability to sense lipids. We will build on our recent discovery that mTORC1 senses an important lipid, cholesterol, at the lysosome. Using innovative approaches both in cells and in vitro, we will address and elucidate key aspects of newly identified signaling pathway. In particular, we will determine i) the cellular location of the cholesterol pools that regulate mTORC1 ii) the transport circuits that make cholesterol available to mTORC1 and iii) the molecular mechanisms through which cholesterol induces mTORC1 recruitment to the lysosomal surface. Moreover, we will investigate how cholesterol sensing by mTORC1 depends on the Niemann-Pick C1 protein, loss of which causes a fatal metabolic and neurodegenerative disease. We will address these research aims via innovative and highly complementary approaches recently optimized in our lab, including measurement and targeted manipulations of the lipid content of selected organelle populations, combined with reconstitution-based assays of mTORC1 regulation. Our findings will impact the current understanding of the molecular mechanisms of cellular lipid homeostasis, and will point the way to novel approaches to manipulate mTORC1 signaling in disease settings.