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

描述(申请人提供)：蛋白质的运输和运输是细胞功能的一个重要方面，因为细胞通过从细胞表面内化和释放蛋白质与环境相互作用。细胞表面蛋白的异常加工或运输导致尿崩症、囊性纤维化和阿尔茨海默病等疾病。虽然通道和受体等特定的货物是不同类型的细胞所特有的，但质膜循环的过程在哺乳动物细胞中是高度保守的，并受到一系列共同的运输蛋白的调节，包括Rab小GTP酶。Rab11是一种广泛表达的小分子GTP酶，参与循环，与Rab11家族相互作用蛋白(FIP)相互作用。尽管对FIP的功能还没有完全了解，但它们可能在一定程度上定义了回收系统中不同的子域。此外，初步数据表明，FIP2在不同的亚细胞定位中以不同的构象存在。这些数据导致了这样的假设，即与Rab11相互作用的FIP的离散寡聚体允许Rab11的时间和空间特异性以及FIP在循环系统中的功能。提出了两个目标来检验这一想法。首先，将通过酵母双杂交、FRET显微镜和蛋白质共定位来评估FIP的同源二聚化和异源二聚化能力，以此来了解FIP在循环系统中的功能。第二，关于循环系统内分子内和分子间相互作用的FIP2构象将被研究。通过阐明FIP复合体的组成及其亚细胞定位，可以揭示Rab11和FIP作为质膜循环一般调节因子的重要性。 意义：正确的蛋白质转运和定位对于维持正常的细胞功能至关重要。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。这项建议旨在通过研究一种重要的运输蛋白Rab11及其相互作用蛋白来增加对所有类型细胞共同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.