Mechanisms of Endocytic Recycling

内吞回收机制

• 批准号: 9322098
• 负责人: VICTOR W HSU
• 金额: $ 9万
• 依托单位: BRIGHAM AND WOMEN'S HOSPITAL
• 依托单位国家: 美国
• 财政年份: 2015
• 资助国家: 美国
• 起止时间: 2015-07-01 至 2019-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9322098
• 关键词: ADP-ribosylation factor 6; Ankyrin Repeat; Binding; Capsid Proteins; Carrier Proteins; Cell division; Cell membrane; Cell surface; Cell-Cell Adhesion; Cells; Clathrin; Complex; Coupled; Critical Pathways; Cryoelectron Microscopy; Crystallography; Data; Disease; Endosomes; Ensure; Environment; Event; GTPase-Activating Proteins; Guanine Nucleotide Exchange Factors; Health; Integrins; Intracellular Transport; Mediating; Membrane; Monomeric GTP-Binding Proteins; Nutrient; PH Domain; Pathway interactions; Phosphatidylinositols; Phosphorylation; Phosphotransferases; Physiological; Play; Process; Proteins; Recruitment Activity; Recycling; Regulation; Resolution; Role; Seminal; Signal Transduction; Sorting - Cell Movement; Structure; Surface; Uncertainty; amphiphysin; cell growth; cell motility; human disease; innovation; insight; novel; particle; protein structure; receptor; uptake

 DESCRIPTION (provided by applicant): Endocytic recycling is critical for many physiologic events, including nutrient uptake, cell signaling, polarity, and cell adhesion and migration. In recent years, we have advanced a fundamental understanding of endocytic recycling by identifying the first coat complex to act in this process. This complex is composed of clathrin coupled to a novel adaptor, known as ACAP1 (Arfgap with Coil-coil, Ankyrin repeat and PH domain 1). We have also identified ARF6 (ADP-Ribosylation Factor 6) to regulate this coat complex, with Grp1 (General receptor for 3-phosphoinositides 1) acting as the guanine nucleotide exchange factor (GEF). More recently, our studies on these factors have uncovered significant new insights, which we propose to pursue in further detail. First, we have recently pursued a cutting-edge approach that combines single-particle cryo-electron microscopy with protein crystallography. This approach has shed unexpected new insight into how ACAP1 bends membrane. Thus, we propose to apply this combined approach to study how ARF6 bends membrane, and also examine how ARF6 cooperates with ACAP1 to impart membrane curvature. Second, we have also uncovered that Akt promotes endocytic recycling through a non-kinase function. Thus, we will examine three possibilities for this novel role: i) acting as a cargo adaptor to promote cargo binding by ACAP1, ii) regulating the GTPase activating protein (GAP) activity of ACAP1, or iii) modulating the ability of ACAP1 to bend membrane. Third, we have also uncovered new insights into the regulation of Grp1, which involves the phosphorylation of a key residue in its PH (Pleckstrin Homology) domain, resulting in Grp1 being targeted to the recycling endosome rather than the plasma membrane. We have also identified additional factors that promote this switch. Thus, we will elucidate how these different targeting mechanisms may be coordinated, and thereby contributing to a better understanding of how ARF GEFs are regulated in complex ways to ensure that a transport pathway is properly initiated. As endocytic recycling underlies many key physiologic events, we anticipate that our studies will not only advance a basic understanding of transport mechanisms, but also shed insights into how endocytic recycling is critical for both health and disease states.

 描述（由申请人提供）：内吞再循环对于许多生理事件至关重要，包括营养摄取、细胞信号传导、极性以及细胞粘附和迁移。近年来，我们已经通过识别在此过程中起作用的第一个外壳复合物，对内吞再循环有了基本的了解。这种复合物由网格蛋白偶联到称为ACAP 1（具有卷曲螺旋、锚蛋白重复和PH结构域1的Arfgap）的新型接头组成。我们还确定了ARF 6（ADP-核糖基化因子6）调节这种外壳复合物，Grp 1（3-磷酸肌醇1的一般受体）作为鸟嘌呤核苷酸交换因子（GEF）。最近，我们对这些因素的研究发现了重要的新见解，我们建议进一步详细研究。首先，我们最近采用了一种将单粒子冷冻电子显微镜与蛋白质晶体学相结合的尖端方法。这种方法为了解ACAP 1如何弯曲膜提供了意想不到的新见解。因此，我们建议应用这种组合的方法来研究ARF 6如何弯曲膜，并研究ARF 6如何与ACAP 1合作，赋予膜曲率。其次，我们还发现Akt通过非激酶功能促进内吞再循环。因此，我们将研究这种新作用的三种可能性：i）充当货物衔接子以促进ACAP 1的货物结合，ii）调节ACAP 1的GTP酶激活蛋白（GAP）活性，或iii）调节ACAP 1弯曲膜的能力。第三，我们还发现了对Grp 1调控的新见解，这涉及其PH（Pleckstrin同源性）结构域中关键残基的磷酸化，导致Grp 1靶向回收内体而不是质膜。我们还确定了促进这种转变的其他因素。因此，我们将阐明这些不同的 可以协调靶向机制，从而有助于更好地了解ARF GEF如何以复杂的方式进行监管，以确保正确启动运输途径。由于内吞再循环是许多关键生理事件的基础，我们预计我们的研究不仅将促进对运输机制的基本理解，而且还将深入了解内吞再循环对健康和疾病状态的关键作用。