Spatio-temporal regulation of mTORC1 signaling in normal and disease states

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， 在那里它会被激活。越来越多的证据表明，mTORC1的异常激活在 溶酶体可能是从癌症到2型糖尿病再到神经退化等疾病的驱动力。 溶酶体转位和激活mTORC1需要异二聚体RAG鸟苷 三磷酸酶(GTP酶)与五聚体刺激器复合体一起形成营养调节的 以物理方式将mTORC1固定在溶酶体表面的支架复合体。将动态成像与 细胞的生化重建和结构方法，我们最近发现的Ragator-RAG 复合体不是静态的，而是被营养物质积极地重塑，导致碎屑的空间循环 溶酶体表面和细胞质之间的GTP酶。反过来，RAG自行车对 当营养水平下降时，mTORC1捕获的效率可能会更高，并可能有助于其失活。重要的是，拉格 影响mTORC1信号的癌症特异性突变改变了细胞周期。基于这些发现，我们 假设mTORC1支架的时空调控是一种新的未被认识的机制 来调节mTORC1信号反应的效力和选择性，并且它的干扰可能会驱动 MTORC1驱动的癌症的异常生长，包括肾细胞癌和淋巴瘤。 我们将通过两个互补性和创新性很强的研究目标来检验这一假设。首先，我们将 利用结构导向突变来剖析控制mTORC1-1组装的机制 支架复合体，以响应不断变化的养分输入。第二，我们将描述这一机制 我们最近发现的阻止溶酶体组装的新一代化合物的作用 MTORC1支架复合体，并测定其抑制mTORC1代谢和生长的能力- 导致癌症。 总的来说，拟议的研究将产生关于时空调节的新知识 MTORC1信号转导，并指出了在正常和 疾病状态。

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 it 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. Lysosomal translocation and activation of mTORC1 requires the heterodimeric Rag guanosine triphosphatases (GTPases), which together with the pentameric Ragulator complex, form a nutrient-regulated scaffolding complex that physically anchors mTORC1 to the lysosomal surface. Combining dynamic imaging in cells with biochemical reconstitution and structural approaches, we recently discovered that the Ragulator-Rag complex is not static but is rather actively remodeled by nutrients, leading to spatial cycling of the Rag GTPases between the lysosomal surface and the cytoplasm. In turn, Rag cycling places a limit on the efficiency of mTORC1 capture and may facilitate its inactivation when nutrient levels fall. Importantly, Rag cycling is altered by cancer-specific mutations that affect mTORC1 signaling. Based on these findings, we hypothesize that spatial-temporal regulation of mTORC1 scaffolding is a novel and unrecognized mechanism to modulate the potency and selectivity of mTORC1 signaling responses, and that its disruption may drive the aberrant growth of mTORC1-driven cancers, including renal cell carcinoma and lymphoma. We will test this hypothesis via two highly complementary and innovative research aims. First, we will employ structure-guided mutagenesis to dissect the mechanisms that govern the assembly of the mTORC1- scaffolding complex in response to changing nutrient inputs. Second, we will characterize the mechanism of action of new-generation compounds we recently discovered, which block the assembly of the lysosomal mTORC1 scaffolding complex, and determine their ability to inhibit the metabolism and growth of mTORC1- driven cancers. Collectively, the proposed studies will generate new knowledge on the spatial-temporal regulation of mTORC1 signaling, and point the way to novel strategies to manipulate mTORC1 signaling in both normal and disease states.