Role of Rab Proteins in Golgi Apparatus Structure and Function

Rab 蛋白在高尔基体结构和功能中的作用

基本信息

批准号：
8723844
负责人：
Brian Storrie
金额：
$ 31.84万
依托单位：
UNIV OF ARKANSAS FOR MED SCIS
依托单位国家：
美国
项目类别：
财政年份：
2011
资助国家：
美国
起止时间：
2011-09-30 至 2016-08-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/8723844
关键词：
Address Affect Aging Antigens Biogenesis Biological Assay Cell Fractionation Cells Chlamydia Infections Complex Defect Disease Electron Microscope Electron Microscopy Enzymes Event Glaucoma Golgi Apparatus Health Homeostasis Human In Situ Individual Infection Control Lead Maintenance Mediating Membrane Membrane Fusion Molecular Genetics Monomeric GTP-Binding Proteins Neurodegenerative Disorders Organelles Outcome Pathway interactions Pharmaceutical Preparations Phenotype Play Process Protein Isoforms Proteins RNA Interference Reagent Regulatory Pathway Relative (related person)Research Personnel Role Set protein Small Interfering RNA Structure Structure-Activity Relationship Sum Testing Textbooks Therapeutic Transport Vesicles Vesicle Viral Virus Diseases Vision Work base cell killing cell type human disease in vitro Assay novel novel strategies protein function research study screening small molecule tomography trafficking ward

项目摘要

DESCRIPTION (provided by applicant): Golgi associated Rab proteins - small GTPases of the Ras superfamily -- regulate critical, yet poorly understood, aspects of Golgi biogenesis, maintenance, and inheritance through affecting vesicle trafficking. The general (i.e., "textbook") paradigm holds that Golgi Rab proteins regulate vesicle targeting via the recruitment and activation of tethering factors that mediate initial steps of membrane fusion. Our Preliminary Studies support a different and novel mechanism. We screened for molecular genetic relationship(s) between Rab33b and Rab6 - two Golgi Rabs that our works implicates in an intra-Golgi Rab cascade -- and the putative tether proteins, COG and ZW10/RINT-1, central to retrograde Golgi trafficking. We found that both Rab6 and Rab33b acted upstream of, not through, the tether complexes. By electron microscope tomography (ET), we observed that Rab6 knockdown results in an apparent inhibition of Golgi vesicle budding/transport as evidenced by the pronounced accumulation of both coated and uncoated budding profiles/vesicles along Golgi cisternal membranes. This has led us to conclude that Rab6 is required for efficient Golgi vesicle release/scission. In sum, our results suggest that Golgi Rabs such as Rab6 and Rab33b serve important functions in very early events at the Golgi related to vesicle scission, and that they act upstream of tether factor recruitment in this fundamental process. Based on our Preliminary Studies, we thus contend that a critical and functionally important role of Golgi-associated Rabs (such as Rab6 and Rab33b) is to regulate, through context sensitive effector sets, the budding/release of distinct classes of transport vesicles from Golgi cisternae. Functionally, we predict that each vesicle class supports a distinct trafficking pathway(s) and as such are compositionally distinct. Cellularly, we hypothesize that cisternal Rab proteins such as Rab6 and likely Rab33b play an important/central role in Golgi homeostasis. This hypothesis is supported by our finding that compared to control cells, Rab6 depleted cells reproducibly reveal a significant increase in the number of Golgi cisternae per stack (4.2 + 0.32 versus 6.8 + 0.46, respectively). We propose that Rab proteins regulate the distribution of Golgi resident proteins between cisternae. We predict that in Rab knockdown experiments that the distribution of individual Golgi enzymes will be shifted cis- or trans-ward. Mechanistically, we hypothesize that distinct effectors, including already identified candidate protein and others novel, will modulate the budding of individual vesicle classes. We propose the following three Specific Aims to test these hypotheses. Specific Aim 1. We will test the hypothesis that Golgi associated Rab proteins such as Rab6 and Rab33b regulate the formation and/or release of multiple, distinct classes of vesicles from Golgi cisternae. Specific Aim 2. We will test the hypothesis that Golgi-associated Rabs regulate the distribution of Golgi resident proteins between cisternae. Specific Aim 3. We will test the hypothesis that mechanistically distinct protein sets, i.e., effectors, modulate the budding/release of individual vesicle classes. Characterization of structural/functional relationships within the mammalian Golgi apparatus and its regulatory pathways is important to human health. We and other investigators have found that such pathways are useful portals into the cell for the delivery of cell-killing reagents and antigens. Defects in these pathways can lead to human disease. Moreover, important machinery components in these pathways are involved in such processes as virus infection and aging. Modulation of Golgi associated Rabs may prove to be an important therapeutic approach.

描述（由申请人提供）：高尔基体相关的Rab蛋白 - RAS超家族的小GTPase-调节了批判性但知之甚少的，通过影响囊泡贩运来调节Golgi生物发生，维持和遗传的各个方面。一般（即“教科书”）范式认为，高尔基rab蛋白通过募集和激活的跨膜融合的初始步骤来调节囊泡的靶向。我们的初步研究支持一种不同的新型机制。我们筛选了Rab33b和Rab6之间的分子遗传关系 - 我们的作品与Golgi Rab cascade有关的两个高尔基兔，以及推定的链球蛋白，COG和ZW10/RINT-1，是逆行高尔基运输的中心。我们发现Rab6和Rab33b均作用于束缚复合物的上游。通过电子显微镜断层扫描（ET），我们观察到Rab6敲低导致高尔基囊泡发芽/转运的明显抑制，这证明了涂层和未涂层的萌芽和未涂层的萌芽曲线/囊泡沿高尔基体膜膜的明显积累。这使我们得出结论，有效的高尔基囊泡释放/分裂是必需的。总而言之，我们的结果表明，诸如Rab6和rab33b之类的高尔基龙在与囊泡分裂有关的高尔基体的非常早期的事件中起着重要的作用，并且它们在此基本过程中起着系带因子募集的上游作用。因此，基于我们的初步研究，我们争辩说，高尔基体相关的RAB（例如Rab6和Rab33b）的关键和功能重要的作用是通过上下文敏感效应子集调节，从高尔基硫氏菌的上下文敏感效应子集调节不同类别的运输囊泡。在功能上，我们预测每个囊泡类都支持一个不同的运输途径，因此在组成上是不同的。我们假设在细胞中，我们假设诸如Rab6和Rab33b之类的蓄水干蛋白在高尔基体稳态中起着重要/中心作用。我们的发现得到了这一假设，即与对照细胞相比，Rab6耗尽的细胞可重复显示每个堆栈的高尔基水库数量显着增加（分别为4.2 + 0.32对6.8 + 0.46）。我们提出，Rab蛋白调节高尔基体蛋白质蛋白之间的分布。我们预测在RAB敲低实验中，单个高尔基酶的分布将转移顺式或转移。从机械上讲，我们假设包括已经确定的候选蛋白和其他小说在内的不同效应子将调节单个囊泡类的萌芽。我们提出以下三个特定目的来检验这些假设。具体目的1。我们将检验以下假设：高尔基体与Rab6和Rab33b相关的Rab蛋白会调节来自高尔基硫磺岛的多种不同类囊泡的形成和/或释放。具体目的2。我们将检验以下假设：高尔基体相关的Rab调节硫斯基之间高尔基体蛋白质的分布。具体目标3。我们将测试以下假设：机械上不同的蛋白质集（即效应子）调节单个囊泡类的萌芽/释放。哺乳动物高尔基体及其调节途径内结构/功能关系的表征对人类健康很重要。我们和其他研究人员发现，此类途径是进入细胞的有用门户，用于输送细胞的试剂和抗原。这些途径中的缺陷会导致人类疾病。此外，这些途径中的重要机械组件参与了病毒感染和衰老等过程。对高尔基体相关的RAB的调节可能被证明是一种重要的治疗方法。

项目成果

期刊论文数量（9）

专著数量（0）

科研奖励数量（0）

会议论文数量（0）

专利数量（0）

Distinct sets of Rab6 effectors contribute to ZW10--and COG-dependent Golgi homeostasis.

DOI：
10.1111/tra.12167
发表时间：
2014-06
期刊：
Traffic (Copenhagen, Denmark)
影响因子：
0
作者：
Majeed W;Liu S;Storrie B
通讯作者：
Storrie B

How Rab proteins determine Golgi structure.

DOI：
10.1016/bs.ircmb.2014.12.002
发表时间：
2015
期刊：
International review of cell and molecular biology
影响因子：
0
作者：
Liu S;Storrie B
通讯作者：
Storrie B

Electron tomography reveals Rab6 is essential to the trafficking of trans-Golgi clathrin and COPI-coated vesicles and the maintenance of Golgi cisternal number.

DOI：
10.1111/j.1600-0854.2012.01343.x
发表时间：
2012-05
期刊：
Traffic (Copenhagen, Denmark)
影响因子：
0
作者：
Storrie B;Micaroni M;Morgan GP;Jones N;Kamykowski JA;Wilkins N;Pan TH;Marsh BJ
通讯作者：
Marsh BJ

Are Rab proteins the link between Golgi organization and membrane trafficking?

DOI：
10.1007/s00018-012-1021-6
发表时间：
2012-12
期刊：
CELLULAR AND MOLECULAR LIFE SCIENCES
影响因子：
8
作者：
Liu, Shijie;Storrie, Brian
通讯作者：
Storrie, Brian

Rab6a/a' are important Golgi regulators of pro-inflammatory TNF secretion in macrophages.

DOI：
10.1371/journal.pone.0057034
发表时间：
2013
期刊：
PloS one
影响因子：
3.7
作者：
Micaroni M;Stanley AC;Khromykh T;Venturato J;Wong CX;Lim JP;Marsh BJ;Storrie B;Gleeson PA;Stow JL
通讯作者：
Stow JL