Mechanisms and function of endosome-derived tubular carriers

内吞转运是正常细胞功能的中心，而失调是导致 疾病多种多样，如动脉粥样硬化、糖尿病和癌症。我的实验室致力于 了解膜和受体通过内吞循环的机制 从回收室(ERC)到质膜(PM)。内吞循环仍然是最少的循环之一 研究了内吞途径；特别是膜小管在循环过程中的参与 以及各种形式的管状内小体的作用和通过其机制 这些结构的产生和泡化仍然是一个主要的悬而未决的问题。确实有 多种“类型”的内体衍生管状载体(EDTC)，包括分选Nexin-bar(SNX-bar) 结构域和反聚体衍生的小管，以及内体小管的网络，如小管再循环 Mical-L1、Synd2(Synd2)和Eps15同源结构域(EHD)修饰的内小体(TrE) 蛋白质。关于这些明显不同的EDTC如何整合它们的功能，甚至是这样，我们知之甚少 逆转聚体和SXN-bar EDTC是否与TRE不同。最近的研究表明，EHD 蛋白质和Mical-L1与逆转聚体复合体的成分相互作用，表明这些小管 网络是相关的。我们的中心假设是重叠的和不同的管膜协调 从内体运输到PM和高尔基体。在本提案中，我们将揭示功能和 Tre、Retmer和SNX-bar衍生的EDTC之间的物理关系。此外，我们将重点关注一种 从机制上理解EDTC的产生方式，以及它们如何经历囊泡形成 以促进细胞内的运输。目的1：探讨EDTC的相互关系和功能作用。 管状内小体在胞膜转运中起着重要作用。EDTC包括一些 含有逆转聚体的结构，由SNX-bar结构域蛋白产生的逆转聚体非依赖的小管， Tre由Mical-L1、BAR结构域蛋白Synd2和EHD蛋白修饰。我们将研究 在回收的产生和裂变方面，这些途径的相互影响和交叉管理 小管和内吞循环的控制。目标2：定义和阐明时空 Tre生物发生和囊泡形成的调控和机制。我们的工作假设是 磷脂酸的产生导致膜中枢、Mical-L1和F-bar蛋白的募集 Synd2，以生成和重新建模Tre。我们进一步假设EHD3在这一过程中起着重要的作用 通过稳定Mical-L1-Synd2相互作用的过程。我们对细胞、细胞器、分子和 原子水平将使用从结构生物学到超分辨率显微镜和新颖的技术 生物物理囊泡分析，提供对知之甚少的人的功能作用的关键知识 TrE和Retmer-SNX-bar来源的小管及其生物发生和囊泡形成。

Endocytic trafficking is central to normal cell function, and dysregulation is the underlying cause for diseases as diverse as atherosclerosis, diabetes and cancer. My laboratory has contributed to understanding the mechanisms by which membranes and receptors are recycled via the endocytic recycling compartment (ERC) to the plasma membrane (PM). Endocytic recycling remains one of the least studied endocytic pathways; in particular, the involvement of membrane tubules in the recycling process is poorly understood, and the roles of the various forms of tubular endosomes, and the mechanisms by which these structures are generated and undergo vesiculation remain a major unanswered question. There are multiple `types' of endosome-derived tubular carriers (EDTC), including sorting nexin-BAR (SNX-BAR) domain and retromer-derived tubules, and networks of endosomal tubules, such as tubular recycling endosomes (TRE) decorated by MICAL-L1, Syndapin2 (Synd2) and Eps15 Homology Domain (EHD) proteins. Little is known about how these apparently different EDTC integrate their functions, or even whether retromer and SXN-BAR EDTC are distinct from TRE. Recent studies demonstrate that EHD proteins and MICAL-L1 interact with components of the retromer complex, suggesting that these tubular networks are related. Our central hypothesis is that overlapping and distinct tubular membranes coordinate transport from endosomes to the PM and the Golgi. In this proposal, we will uncover the functional and physical relationships between TRE, retromer and SNX-BAR-derived EDTC. Moreover, we will focus on a mechanistic understanding of the mode by which EDTC are generated, and how they undergo vesiculation to promote transport within the cell. Aim 1: To examine the relationship and functional roles of EDTC. Tubular endosomes play major roles in endocytic membrane trafficking. EDTC include a number of retromer-containing structures, retromer-independent tubules generated by SNX-BAR domain proteins, and TRE decorated by MICAL-L1, the BAR domain protein Synd2, and EHD proteins. We will examine the cross-talk and cross-regulation of these pathways with regard to the generation and fission of recycling tubules, and the control of endocytic recycling. Aim 2: To define and elucidate the spatio-temporal regulation and mechanism of TRE biogenesis and vesiculation. Our working hypothesis is that phosphatidic acid generation leads to recruitment of the membrane hub, MICAL-L1 and the F-BAR protein Synd2, to generate and remodel TRE. We further hypothesize that EHD3 plays an essential role in this process by stabilizing MICAL-L1-Synd2 interactions. Our studies on the cellular, organellar, molecular and atomic levels will use techniques ranging from structural biology to super-resolution microscopy and novel biophysical vesiculation assays, providing crucial knowledge of the functional role of the poorly understood TRE and retromer-SNX-BAR derived tubules, and their biogenesis and vesiculation.