Rab11-FIPs interact to define distinct functional nodes in the recycling system

描述（由申请人提供）：蛋白质运输和转运是细胞功能的关键方面，因为细胞通过内化和从细胞表面释放蛋白质与环境相互作用。细胞表面蛋白质的异常加工或转运有助于疾病，如尿崩症、囊性纤维化和阿尔茨海默病。虽然特定的货物，如通道和受体是独特的不同的细胞类型，质膜再循环的过程是高度保守的哺乳动物细胞和一组共同的运输蛋白，包括Rab小GTP酶的调节。Rab11是一种广泛表达的小GTP酶，参与与Rab11家族相互作用蛋白（FIPs）相互作用的再循环。虽然FIPs的功能还没有完全被理解，但它们可能部分地定义了回收系统中的不同子域。此外，初步数据表明，FIP 2以不同的构象存在于不同的亚细胞定位中。这些数据导致的假设，即离散的寡聚体的FIPs与Rab11相互作用，允许Rab11和FIP功能的时间和空间的特异性内的回收系统。提出了两个目标来测试这个想法。首先，FIPs的同源二聚化和异源二聚化能力将通过酵母双杂交、FRET显微镜和蛋白质共定位来评估，作为了解FIPs在回收系统内功能的一种方式。第二，FIP2构象将研究关于分子内和分子间的相互作用内的回收系统。通过阐明FIP复合物的组成及其亚细胞定位，Rab11和FIP作为质膜再循环的一般调节剂的重要性可能会被揭示。 意义：蛋白质的正确转运和定位对维持正常细胞功能至关重要。诸如囊性纤维化、尿崩症和阿尔茨海默病等疾病是由重要蛋白质的错误定位或异常运输引起的。该提案旨在通过研究重要的运输蛋白Rab11及其相互作用蛋白，增加对所有细胞类型共有的蛋白质运输基本机制的理解。

DESCRIPTION (provided by applicant): Protein trafficking and transport is a crucial aspect of cellular function because cells interact with the environment by internalization and release of proteins from the cell surface. Aberrant processing or transport of cell surface proteins contributes to diseases such as diabetes insipidus, cystic fibrosis, and Alzheimer's disease. While the particular cargo such as channels and receptors is unique to different cell types, the process of plasma membrane recycling is highly conserved among mammalian cells and regulated by a common set of trafficking proteins including the Rab small GTPases. Rab11 is a ubiquitously expressed small GTPase involved in recycling that interacts with the Rab11-Family Interacting Proteins (FIPs). Although the functions of FIPs are not fully understood, they may in part define distinct subdomains within the recycling system. Furthermore, preliminary data suggest that FIP2 exists in different conformations in different subcellular localizations. These data lead to the hypothesis that discrete oligomers of FIPs interacting with Rab11 allow for temporal and spatial specificity of Rab11 and FIP function within the recycling system. Two aims are proposed to test this idea. First, homodimerization and heterodimerization capabilities of FIPs will be assessed by yeast two hybrid, FRET microscopy, and protein colocalization as a way to understand FIP function within the recycling system. Second, FIP2 conformation will be studied with regards to intramolecular and intermolecular interactions within the recycling system. By elucidating the composition of FIP complexes and their subcellular localizations, the importance of Rab11 and FIPs as general regulators of plasma membrane recycling may be uncovered. SIGNIFICANCE: Proper protein transport and localization are crucial to maintenance of normal cellular function. Diseases such as cystic fibrosis, diabetes insipidus, and Alzheimer's disease result from mislocalization or aberrant trafficking of vital proteins. This proposal aims to increase the understanding of the basic mechanism of protein trafficking that is common to all cell types by studying an important trafficking protein, Rab11, and its interacting proteins.