Roles of Phospholipase D in AT1 Receptor Endocytosis

磷脂酶 D 在 AT1 受体胞吞作用中的作用

• 批准号: 7804120
• 负责人: GUANGWEI DU
• 金额: $ 7.96万
• 依托单位: UNIVERSITY OF TEXAS HLTH SCI CTR HOUSTON
• 依托单位国家: 美国
• 财政年份: 2006
• 资助国家: 美国
• 起止时间: 2006-03-01 至 2011-02-28
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7804120
• 关键词: Actins; Affect; Agonist; Angiotensin II; Appearance; Cardiovascular Diseases; Cell Proliferation; Cell membrane; Cell surface; Choline; Clathrin; Cytoskeletal Modeling; Down-Regulation; Endocytic Vesicle; Endocytosis; Endosomes; Equilibrium; Event; Family; Functional disorder; G Protein-Coupled Receptor Signaling; G-Protein-Coupled Receptors; Goals; Heart Hypertrophy; Heterogeneity; Hydrolysis; Kinetics; Lecithin; Location; Mediating; Membrane; Membrane Lipids; Membrane Microdomains; Membrane Protein Traffic; Membrane Proteins; Metabolism; Modeling; Output; Pathway interactions; Pattern; Phosphatidic Acid; Phosphatidylinositols; Phospholipase D; Phospholipid Metabolism; Phospholipids; Play; Process; Production; Protein Isoforms; Proteins; Publishing; Regulation; Reporting; Research Personnel; Role; Signal Pathway; Signal Transduction; Signaling Molecule; Signaling Protein; Site; Testing; Work; base; blood pressure regulation; cell growth; coated pit; desensitization; essential phospholipids; genetic regulatory protein; insight; member; novel therapeutic intervention; programs; protein function; receptor; receptor binding; research study; response; segregation; small hairpin RNA; spatiotemporal; trafficking

DESCRIPTION (provided by applicant): The angiotensin II type I receptor (AT1R) mediates diverse intracellular responses to angiotensin II (Ang II) in the regulation of blood pressure, hydromineral balance, cardiac hypertrophy and cell proliferation. Endocytosis of the Ang ll-bound receptor not only desensitizes AT1R signaling from the cell surface but also activates specific intracellular signaling pathways. The long-term goal of this project is to elucidate the signaling pathways that regulate AT1R subcellular trafficking. Clarifying the events that ensue will potentially suggest approaches for novel therapeutics for cardiovascular diseases related to AT1R dysfunction, and will provide new insights into the mechanism underlying receptor endocytosis in general. Although roles for many proteins in AT1R trafficking are becoming clear, less is known about the specific functions of membrane lipids. It has been recognized that interaction between proteins and membrane (particularly phospholipids) drives initiation, fission, and fusion of endocytic vesicles. However, how phospholipid metabolism is regulated during this process and, in turn, triggers signals for the different receptor endocytic steps, remains unclear. The Phospholipase D (PLD) family contains two members, PLD1 and PLD2, both of which catalyze the hydrolysis of phosphatidylcholine (PC) to generate phosphatidic acid (PA) and choline. PLD2-generated PA has been proposed to regulate a number of endocytic and actin regulatory proteins and several types of membrane trafficking processes. The hypothesis of this proposal, based on our published work and preliminary evidence, is that PLD2-generated PA plays a central signaling role in regulation of AT1R endocytosis and signaling. In specific aim 1, we will test the hypothesis that PLD2 facilitates AT1R endocytosis by regulating spatiotemporal dynamics of phosphatidylinositol 4,5-bisphosphate {PI(4,5)P2}, which is an essential phospholipid in endocytosis. Specific aim 2 will test whether PLD2 regulates formation of clathrin-coated pits (CCPs) by determining which step in CCP formation is blocked when PLD2 is downregulated by small hairpin RNA (shRNA). We will also examine whether membrane microdomains regulated by PLD2 signaling are critical for CCP formation. Specific aim 3 will test whether differential distribution of AT1R as regulated by PLD2 determines the output of AT1R signaling. AT1R activation in different subcellular locations results in distinct signaling outputs. We will determine whether PLD2 downregulation changes the kinetic and spatial patterns of Ang II signaling, and as a result, affects cell growth.

描述（由申请人提供）：血管紧张素I II型受体（AT1R）在调节血压，氢化层平衡，心脏肥大和细胞增殖方面介导了对血管紧张素II（ANG II）的细胞内反应。 ANG结合受体的内吞作用不仅使细胞表面的AT1R信号脱敏，而且激活了特定的细胞内信号通路。该项目的长期目标是阐明调节AT1R亚细胞运输的信号传导途径。澄清随之而来的事件将有可能提出与AT1R功能障碍有关的心血管疾病的新型治疗方法的方法，并将提供有关总体受体内吞作用机制的新见解。尽管许多蛋白质在AT1R运输中的作用变得越来越清晰，但对膜脂质的特定功能知之甚少。人们已经认识到，蛋白质与膜（尤其是磷脂）之间的相互作用可以驱动内吞囊泡的起始，裂变和融合。然而，在此过程中如何调节磷脂代谢，而反过来又触发了不同受体内吞步骤的信号，尚不清楚。磷脂酶D（PLD）家族包含两个成员PLD1和PLD2，这两者都催化磷脂酰胆碱（PC）的水解产生磷脂酸（PA）和胆碱。已提出PLD2生成的PA来调节许多内吞和肌动蛋白调节蛋白以及几种类型的膜运输过程。根据我们已发表的工作和初步证据，该提案的假设是PLD2生成的PA在调节AT1R内吞和信号传导的调节中起着中心信号的作用。在特定目标1中，我们将通过调节磷脂酰肌醇4,5-双磷酸{pi（4,5）p2}的时空动力学来促进AT1R内吞作用的假设，这是内胞病中必不可少的磷脂。特定的目标2将测试PLD2是否通过确定在PLD2被小型发夹RNA（SHRNA）下调时，是否通过确定CCP形成中的哪个步骤被阻断来调节涂有网格蛋白涂层的坑（CCP）。我们还将检查由PLD2信号调节的膜微区域是否对于CCP形成至关重要。特定的目标3将测试由PLD2调节的AT1R的差分分布确定AT1R信号的输出。不同亚细胞位置的AT1R激活导致不同的信号输出。我们将确定PLD2下调是否会改变ANG II信号的动力学和空间模式，从而影响细胞生长。