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: 10115517
• 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/10115517
• 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; 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; military veteran; mouse model; non-muscle myosin; novel; novel strategies; novel therapeutic intervention; 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.

本VA Merit的目的是研究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的形式。 在囊泡被送到目的地之前，细胞内的囊泡。因此，这个细胞器位于 分泌途径、溶酶体途径和内吞途径的交叉点;已知其在 加工蛋白质以供分泌。我们实验室以前的工作已经确定了多个缺陷， 内吞，蛋白运输和分泌，酒精管理后，但我们到现在为止，没有检查 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）的发病率是 更多的退伍军人人口和超过一半的所有医疗入院的VA医疗中心在整个 我们正在集中努力确定潜在的目标， 在慢性酒精给药后发生的进行性损伤期间进行干预，也许高尔基体 会成为这样的目标对我们的研究至关重要的是一个小的GTdR，Rab 3D， 参与胞吐、分泌和囊泡运输。我们已经证明Rab 3D蛋白质含量是 在服用酒精后显著降低，最近我们获得了令人兴奋的新的初步数据 乙醇损伤Rab 3D功能在高尔基体解体和分裂中起重要作用。的 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，以检查其在通过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是后高尔基体的显著损伤的概念 运输，这导致利用trans-Golgi膜形成自噬体。这些 最近的新发现为这一提议提供了极好的基础，并支持了我们的中心假设 乙醇诱导Rab 3D的下调破坏了高尔基体的组装和功能，导致 蛋白质运输和代谢受损，导致肝损伤。为了验证这个假设，我们有 提出了三个具体目标;在目标1中，我们将确定Rab 3D的功能如何调节 在目标2中，我们将研究自噬如何与EtOH诱导的高尔基体解体联系在一起，在目标3中， 我们将探讨最近的初步数据，检查恢复紧凑的高尔基体和重建的trans-Golgi 膜。总之，成功完成这些目标将表征乙醇对高尔基体的影响 破坏，并在此过程中建立改变Rab 3D的作用。我们就能把 酒精介导的肝细胞运输损伤与受损的高尔基体功能的机制，并提供关键 这些信息可能会导致旨在减少或消除肝损伤的治疗策略。凯西医生 主要研究者，她是由三个杰出的共同研究者（博士。 Rasineni）。调查人员具有互补的优势，这对项目的成功至关重要。 这四个人都有酒精性肝损伤的经验，凯西博士在蛋白质运输和 内吞作用，而Rasineni博士目前正在检查Rab 3D在另一个器官胰腺中的功能。博士 Petrosyan是高尔基体形态学方面的专家，Dr. 项目凭借这些专业知识和我们令人兴奋的创新方法，我们预计我们将成为 在这些研究中取得成功，并将能够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。