Downregulation of Rab3D: Critical Role in Golgi Disorganization and the Pathogenesis of Alcoholic Liver Disease

Rab3D 下调：在高尔基体紊乱和酒精性肝病发病机制中的关键作用

• 批准号: 9885965
• 负责人: Carol A. Casey
• 金额: --
• 依托单位: OMAHA VA MEDICAL CENTER
• 依托单位国家: 美国
• 财政年份: 2020
• 资助国家: 美国
• 起止时间: 2020-04-01 至 2024-03-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9885965
• 关键词: Admission activity; Affect; Alcohol abuse; Alcoholic Hepatitis; Alcoholic Liver Cirrhosis; Alcoholic Liver Diseases; Alcoholic liver damage; Alcohols; Animal Model; Autophagocytosis; Autophagosome; Biochemical; Biology; Cell surface; Cells; Chronic; Cirrhosis; Coat Protein Complex I; Country; Cultured Cells; Data; Defect; Destinations; Development; Disease Progression; Down-Regulation; Endocytosis; Ethanol; Exocytosis; Face; Family; Fatty acid glycerol esters; Foundations; Functional disorder; General Population; Glutaminase; Goals; Golgi Apparatus; Guanosine Triphosphate Phosphohydrolases; Healthcare; Heavy Drinking; Hepatocyte; Hepatomegaly; Hepatotoxicity; Hypertrophy; Impairment; In Vitro; Incidence; Injury; Investigation; Knockout Mice; Laboratories; Lead; Link; Liver; Mediating; Medical; Medical center; Membrane; Military Personnel; Monomeric GTP-Binding Proteins; Morphology; Motor; Nonmuscle Myosin Type IIA; Nonmuscle Myosin Type IIB; Organ; Organelles; Pancreas; Pathogenesis; Pathway interactions; Patients; Pharmacology; Physiological; Play; Population; Prevalence; Principal Investigator; Process; Protein Isoforms; Protein Secretion; Proteins; Recovery; Recycling; Research Personnel; Role; Severities; Small Interfering RNA; Structure; Therapeutic; Toxic effect; Vacuole; Vesicle; Veterans; Work; alcohol effect; alcohol exposure; base; experience; human subject; improved; in vivo; innovation; insight; knock-down; knockout gene; lipid metabolism; lipid transport; liver injury; mouse model; non-muscle myosin; novel; novel strategies; novel therapeutics; protein metabolism; protein transport; receptor; reconstitution; restoration; success; therapeutic development; therapeutic target; trafficking

The goal of this VA Merit is to examine how ethanol exposure results in impaired function of the Golgi apparatus. The Golgi apparatus (also called the Golgi body or Golgi complex) packages proteins into membrane bound vesicles inside the cell before the vesicles are sent to their destination. As such, this organelle resides at the intersection of the secretory, lysosomal, and endocytic pathways; it is known to be of particular importance in processing proteins for secretion. Previous work from our laboratory has identified multiple defects in endocytosis, protein trafficking, and secretion, after alcohol administration, but we have not until now, examined a role for altered Golgi function in these processes. Because the incidence of alcoholic liver disease (ALD) is greater in the Veteran population and more than half of all medical admissions in VA Medical Centers across the country are linked to alcohol abuse, we are focusing efforts towards the identification of potential targets to intervene during the progressive injury which occurs after chronic alcohol administration, and perhaps the Golgi will prove to be such a target. Of central importance to our study is the role of a small GTPase, Rab3D, which is involved in exocytosis, secretion and vesicle trafficking. We have shown that Rab3D protein content was significantly decreased after alcohol administration, and recently we have obtained exciting new preliminary data that ethanol-impaired Rab3D function plays an important role in Golgi disorganization and fragmentation. The studies proposed in this application will extend our ongoing investigation of how ethanol alters hepatocyte biology, specifically in protein processing, to an examination of its role in transport through the Golgi. We provide a concept as to how alcohol-induced remodeling of Golgi morphology is a significant impairment of post-Golgi trafficking, and this leads to utilization of trans-Golgi membranes for the formation of autophagosomes. These recent and novel findings provide an excellent foundation for this proposal and support our central hypothesis that EtOH-induced down-regulation of Rab3D disrupts the assembly and function of Golgi apparatus leading to impaired protein trafficking and metabolism, contributing to liver injury. To examine this hypothesis, we have proposed three specific aims; in Aim 1 we will determine how the function of Rab3D regulates the integrity of the Golgi, in Aim 2 we will examine how autophagy is linked to EtOH-induced Golgi disorganization, and in Aim 3 we will explore recent preliminary data examining recovery of compact Golgi and reconstitution of trans-Golgi membranes. Altogether, successful completion of these aims will characterize the effect of EtOH on Golgi disorganization, and establish a role for altered Rab3D during this process. We will be able to correlate mechanisms of alcohol-mediated liver cell trafficking impairments with impaired Golgi function and provide key information that could lead to therapeutic strategies aimed at reducing or eliminating liver injury. Dr. Casey is the principal investigator, and she is joined by three outstanding co-investigators (Drs. Petrosyan, Thomes and Rasineni). The investigators have complementary strengths which are essential for the success of the project. All four are experienced with alcoholic liver injury, and Dr. Casey has specific expertise in protein trafficking and endocytosis, while Dr. Rasineni is currently examining Rab3D function in another organ, the pancreas. Dr. Petrosyan is an expert in Golgi morphology, and Dr. Thomes brings years of experience in autophagy for the project. Together with this expertise and our exciting and innovative approach, we anticipate that we will be successful in these studies and will be able to contribute to improved healthcare for Veteran patients by the identification of mechanisms involved in the alcoholic liver injury.

该 VA Merit 的目标是检查乙醇暴露如何导致高尔基体功能受损。 高尔基体（也称为高尔基体或高尔基复合体）将蛋白质包装成膜结合 在囊泡被发送到目的地之前，细胞内的囊泡。因此，该细胞器位于 分泌、溶酶体和内吞途径的交叉点；众所周知，它在以下方面特别重要： 加工蛋白质以供分泌。我们实验室之前的工作已经发现了多个缺陷 饮酒后的内吞作用、蛋白质运输和分泌，但我们直到现在才检查 高尔基体功能改变在这些过程中的作用。因为酒精性肝病（ALD）的发病率 退伍军人人口较多，占退伍军人事务部医疗中心所有医疗入院人数的一半以上 国家与酗酒有关，我们正在重点努力确定潜在目标 在长期饮酒后发生的进行性损伤期间进行干预，也许高尔基体 将被证明是这样一个目标。对我们的研究来说最重要的是一种小型 GTP 酶 Rab3D 的作用，它 参与胞吐作用、分泌和囊泡运输。我们已经证明 Rab3D 蛋白含量 饮酒后显着下降，最近我们获得了令人兴奋的新初步数据 乙醇损伤的 Rab3D 功能在高尔基体解体和破碎中起着重要作用。这 本申请中提出的研究将扩展我们正在进行的关于乙醇如何改变肝细胞的研究 生物学，特别是蛋白质加工，以检查其在高尔基体运输中的作用。我们提供 关于酒精诱导的高尔基体形态重塑如何对后高尔基体造成显着损害的概念 贩运，这导致利用跨高尔基体膜来形成自噬体。这些 最近的新颖发现为该提案提供了良好的基础，并支持我们的中心假设 EtOH 诱导的 Rab3D 下调会破坏高尔基体的组装和功能，从而导致 蛋白质运输和代谢受损，导致肝损伤。为了检验这个假设，我们有 提出了三个具体目标；在目标 1 中，我们将确定 Rab3D 的功能如何调节 高尔基体，在目标 2 中，我们将研究自噬如何与 EtOH 诱导的高尔基体解构相关，在目标 3 中 我们将探索最近的初步数据，检查紧凑型高尔基体的恢复和跨高尔基体的重建 膜。总而言之，成功完成这些目标将体现 EtOH 对高尔基体的影响 解组织，并在此过程中为改变的 Rab3D 建立角色。我们将能够关联 酒精介导的肝细胞运输障碍与高尔基体功能受损的机制，并提供关键 可能导致旨在减少或消除肝损伤的治疗策略的信息。凯西博士是 首席研究员，三位杰出的联合研究员（Petrosyan 博士、Thomes 博士和 拉西内尼）。研究人员具有互补的优势，这对于项目的成功至关重要。 所有四人都有酒精性肝损伤的经验，凯西博士在蛋白质运输和 Rasineni 博士目前正在检查 Rab3D 在另一个器官（胰腺）中的功能。博士。 Petrosyan 是高尔基体形态方面的专家，Thomes 博士为我们带来了多年的自噬经验 项目。凭借这些专业知识以及我们令人兴奋和创新的方法，我们预计我们将 在这些研究中取得成功，并将能够通过以下方式为改善退伍军人患者的医疗保健做出贡献 确定酒精性肝损伤的机制。