A Novel Mechanism of GTPase Regulation: Arl2 Interaction with TBCD

GTPase 调节的新机制：Arl2 与 TBCD 的相互作用

• 批准号: 8919087
• 负责人: Joshua W Francis
• 金额: $ 4.31万
• 依托单位: EMORY UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2014
• 资助国家: 美国
• 起止时间: 2014-09-01 至 2017-08-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8919087
• 关键词: ADP-Ribosylation Factors; Binding; Biochemical; Biological; Biological Assay; Cardiac; Cell Proliferation; Cell division; Cell physiology; Cells; Centrosome; Complex; Cytosol; Data; Development; Disease; Dissociation; Dominant-Negative Mutation; Embryo; Energy Metabolism; Eukaryotic Cell; Family; Goals; Guanine Nucleotide Exchange Factors; Guanosine Triphosphate Phosphohydrolases; Health; Heart Diseases; Human; Hypoxia; In Vitro; Lead; Learning; Life; Link; Maintenance; Malignant Neoplasms; Mammalian Cell; Mammary Gland Parenchyma; Microtubule Polymerization; Microtubules; Mitotic spindle; Modeling; Molecular Chaperones; Nucleotides; Pancreas; Pathway interactions; Phenotype; Play; Polymers; Positioning Attribute; Preparation; Process; Property; Protein Binding; Proteins; Recombinants; Regulation; Retinal Degeneration; Role; Series; Signal Transduction; Solid Neoplasm; Source; System; Testing; Transcription Coactivator; Tubulin; Vascularization; Warburg Effect; base; cell growth; cofactor; design; human disease; insight; kidney cell; member; milligram; mutant; novel; overexpression; public health relevance; research study; tumor growth

DESCRIPTION (provided by applicant): The strict regulation of energy metabolism and cell proliferation is critical for the normal function of all eukaryotic life. Improper regulation of thse fundamental cellular processes is associated with multiple disease states, including heart disease and cancer. Examples include shifting the sources of ATP in cells as a result of the Warburg effect or hypoxia, resulting from solid tumor growth exceeding vascularization or cardiac insufficiency. ADP-ribosylation factor like-2 (Arl2), a member of the Arf family of regulatory GTPases, is an ancient and ubiquitous eukaryotic cell regulator that is involved in the maintenance of cellular ATP levels, the progression of cell division and the regulation of transcriptional activators involved in cell proliferation. Maintaining the necessary levels of activated Arl2 within a cell is crucial for the proper regulation of these essential processes. However, the biological mechanisms controlling Arl2 activity are unknown. The majority of cellular Arl2 is in a cytosolic complex with the tubulin-specific co-chaperone, cofactor D (TBCD). We will perform in vitro biochemical analyses to test the hypothesis that dissociation from TBCD is a critical, regulated step in Arl2 function in cells. Additionally, we will test the model that Arl2/TBCD is also an active complex required for proper regulation of centrosomal functions and microtubule dynamics. In specific aim 1, we will utilize recombinant preparations of Arl2 and TBCD to identify the mechanisms regulating assembly/disassembly of the Arl2/TBCD complex. In specific aim 2, we will use cell-based assays to determine the importance of Arl2 and TBCD interaction in regulating centrosomal and microtubule functions. The results of this study will provide fundamental groundwork in our long-term goal of explaining the mechanistic roles of Arl2 as a ubiquitous regulator that is indispensable to the regulation of several essential cellula processes. Further insight into the mechanisms underlying Arl2 regulation will ultimately lead to a better understanding of fundamental eukaryotic functions, such as energy metabolism and cell proliferation.

描述（由申请人提供）：能量代谢和细胞增殖的严格调节对于所有真核生物的正常功能至关重要。对这些基本细胞过程的不当调节与多种疾病状态有关，包括心脏病和癌症。实例包括由于瓦尔堡效应或缺氧（由实体瘤生长超过血管形成或心功能不全引起）而改变细胞中ATP的来源。ADP-核糖基化因子样-2（Arl 2）是调节性GTP酶Arf家族的成员，是一种古老且普遍存在的真核细胞调节因子，其参与维持细胞ATP水平、细胞分裂的进展以及参与细胞增殖的转录激活因子的调节。在细胞内维持必要水平的活化Arl 2对于这些基本过程的适当调节至关重要。然而，控制Arl 2活性的生物学机制是未知的。大多数细胞Arl 2与微管蛋白特异性辅伴侣，辅因子D（TBCD）在胞质复合物中。我们将进行体外生物化学分析，以检验从TBCD解离是细胞中Arl 2功能的关键调节步骤的假设。此外，我们将测试Arl 2/TBCD也是适当调节中心体功能和微管动力学所需的活性复合物的模型。在具体目标1中，我们将利用Arl 2和TBCD的重组制剂来鉴定调节Arl 2/TBCD复合物的组装/分解的机制。在具体目标2中，我们将使用基于细胞的测定来确定Arl 2和TBCD相互作用在调节中心体和微管功能中的重要性。这项研究的结果将为我们的长期目标提供基本的基础，即解释Arl 2作为一种普遍存在的调节剂的机制作用，这种调节剂对几种基本的细胞过程的调节是不可或缺的。进一步深入了解Arl 2调控的机制将最终导致更好地理解基本的真核生物功能，如能量代谢和细胞增殖。