Sorting and trafficking in the endosomal system

描述（由申请人提供）：本研究项目的主要目标是阐明内体中的蛋白质和脂质被包装到运输载体中的机制，该运输载体从内体出芽并将货物运送到高尔基体和其他细胞器。这些运输途径，统称为“内体逆行运输”，对细胞生理学和稳态，后生动物发育至关重要，并且它们对抗导致溶酶体贮积病，癌症，阿尔茨海默病和其他疾病的知之甚少的过程。在之前的资助周期中，我们发现逆行途径用于调节质膜的组成以响应环境和营养线索，并且我们确定了称为“retromer”的可溶性蛋白分选复合物和称为分选连接蛋白（SNX）蛋白的磷脂酰肌醇3-激酶（PI 3 K）的三种效应物的作用。这些组分在核内体膜上组装，在那里它们捕获并将货物包装到出芽的运输载体中，但潜在的机制尚不清楚。随着retromer的细胞功能原理已经确立，该领域的主要挑战是阐明它发挥作用的机制。将使用生物化学方法来剖析逆转录聚合物与内体膜上的货物组装的机制。这些实验将与逆行运输的活细胞成像相补充，这将使组装反应处于生理环境中。在认识到，retromer分选连接蛋白是一个更大的SNX蛋白家族的成员，所有的功能，但少数已被描述，无偏的，蛋白质组规模的实验将被用来阐明每个分选连接蛋白的功能，并评估其对细胞生理学的具体贡献。

DESCRIPTION (provided by applicant): The broad goal of this research project is to elucidate the mechanisms by which proteins and lipids in the endosome are packaged into transport carriers that bud from the endosome and ferry cargo to the Golgi apparatus and other organelles. These trafficking pathways, collectively termed 'endosomal retrograde transport,' are vital for cell physiology and homeostasis, metazoan development and they counter poorly understood processes that contribute to lysosomal storage diseases, cancer, Alzheimer's disease, and other diseases. In previous funding cycles we discovered that retrograde pathways are used to modulate the composition of the plasma membrane in response to environmental and nutritional cues, and we established roles for a soluble protein sorting complex called 'retromer' and three effectors of Phosphatidylinositol 3-kinase (PI3K) called sorting nexin (SNX) proteins. These components assemble on the endosome membrane where they capture and package cargo into budding transport carriers but the underlying mechanisms are unknown. With the principle cellular function of retromer now well established, the major challenge for the field is to elucidate the mechanisms by which it functions. Biochemical approaches will be used to dissect the mechanism of retromer assembly with cargo on the endosome membrane. These experiments will complemented with live cell imaging of retrograde transport that will place the assembly reactions in a physiological context. In recognition that the retromer sorting nexins are members of a much larger SNX protein family for which the functions of all but a few have been described, unbiased, proteome scale experiments will be used to elucidate the functions of each sorting nexin and to assess their specific contributions to cell physiology.