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Novel Roles for Phosphoinositide Signaling in alpha-Granule Biogenesis

磷酸肌醇信号传导在 α 颗粒生物发生中的新作用

基本信息

批准号：
9884351
负责人：
CHARLES S. ABRAMS
金额：
$ 11.01万
依托单位：
UNIVERSITY OF PENNSYLVANIA
依托单位国家：
美国
项目类别：
财政年份：
2020
资助国家：
美国
起止时间：
2020-02-01 至 2020-08-17
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/9884351
关键词：
Address Affect Alpha Granule Binding Binding Proteins Biochemical Biogenesis Biology Blood Blood Cells Blood Platelets Blood coagulation Bone Marrow Cell membrane Cells Collaborations Cytoplasm Cytoplasmic Granules Data Defect Development Environment Fatty Acids GOLPH3 gene Golgi Apparatus Growth Factor Hand Hematopoiesis Hematopoietic stem cells Human Hydrophobicity Impairment Indiana Individual Inflammation Knock-out Left Link Maintenance Mediating Megakaryocytes Megakaryocytopoieses Membrane Morphology Multivesicular Body Mus Mutate Mutation Myocardial Infarction Neurons Patients Phenotype Phosphatidylinositol Transfer Protein Phosphatidylinositols Phospholipids Platelet Activation Play Process Production Property Protein Family Protein Isoforms Proteins Publications Publishing Regulation Role Second Messenger Systems Signal Pathway Signal Transduction Stroke Syndrome Testing Thrombosis Transforming Growth Factor beta Transmembrane Transport Universities aqueous cytokine human disease in vivo member monomer novel protein transport recruit trafficking trans-Golgi Network

项目摘要

Project Summary Phosphorylated phosphatidylinositols (phosphoinositides) are a type of membrane bound phospholipid that impact multiple diverse processes required for megakaryopoiesis and the activation of platelets. We have recently published in Developmental Cell that phosphoinositides in neuronal cells initiate intracellular trafficking by recruiting effector proteins such as GOLPH3 that are involved in vesicular fusion and budding of plasma membranes during Golgi biogenesis. Since megakaryocyte α-granules are derived from the trans-Golgi network and Multi-Vesicular Bodies, I hypothesize that phosphoinositide signaling is necessary for the intracellular trafficking required for the biogenesis of α-granules. PhosphatidylInositol Transfer Proteins (PITPs) are members of a small protein family that bind and transfer phosphoinositide monomers from one cellular compartment to another and thereby enable phosphoinositide synthesis. We have made the unexpected observation that the two predominant PITP isoforms found within megakaryocytes, PITPα and PITPβ play previously unrecognized but essential roles in the trafficking of cargo from the Multi-Vesicular Body to α-granules. Loss of PITP-mediated phosphoinositide synthesis produces morphologic defects similar to what is seen in humans with Gray Platelet Syndrome. The overall hypothesis of this Proposal is that phosphoinositide signaling mediated by PITPs is necessary for the membrane dynamics and protein trafficking required for the biogenesis and maintenance of megakaryocyte α-granules. In Aim 1 of the Project, we will rigorously analyze the discrete biochemical properties of individual PITP isoforms in megakaryocytes. Our preliminary data shows that the two PITP isoforms control phosphoinositide signaling through biochemically distinct mechanisms. In Aim 2, we will determine how phosphoinositide signaling contributes to alpha granule biogenesis and function. We will test the hypothesis that phosphoinositide synthesis within discrete microdomains of megakaryocytes and platelets regulates effector proteins such as NBEAL2 (the mutated protein responsible for Gray Platelet Syndrome). This signaling cascade modulates NBEAL2’s ability to mediate membrane dynamics and protein trafficking. We will also analyze in detail the functional roles of α-granules with ex vivo rheologic and ultramicroscopy studies, in vivo thrombosis studies, and in vivo inflammation studies.

项目摘要磷酸化磷脂酰肌醇（磷酸肌醇）是一种膜结合型磷脂酰肌醇。磷脂影响巨核细胞生成所需的多种不同过程，激活血小板。我们最近在《发育细胞》上发表了磷酸肌醇在神经元细胞中通过募集效应蛋白如GOLPH 3启动细胞内运输它们参与高尔基体生物发生过程中质膜的泡状融合和出芽。由于巨核细胞α-颗粒来源于高尔基体网络和多囊泡细胞，因此，我假设磷酸肌醇信号传导对于细胞内运输是必要的是α颗粒生物发生所必需的。磷脂酰肌醇转移蛋白（PITP）是一个小的蛋白质家族的成员，结合和转移磷酸肌醇单体从一个细胞隔室彼此相连，从而使磷酸肌醇合成成为可能。我们取得了出乎意料的观察结果是，在细胞内发现的两种主要的PITP同种型，在巨核细胞中，PITPα和PITPβ发挥着以前未被认识到的但重要的作用，将货物从多囊泡体运输到α颗粒。PITP介导的丢失磷酸肌醇合成产生的形态缺陷类似于在人类中观察到的，灰色血小板综合征。该提案的总体假设是磷酸肌醇信号传导由PITP介导的是必要的膜动力学和蛋白质运输所需的巨核细胞α颗粒的生物发生和维持。在项目目标1中，我们将严格分析单个PITP亚型的离散生化特性，巨核细胞我们的初步数据表明，两个PITP亚型控制磷酸肌醇通过生物化学上不同的机制发出信号。在目标2中，我们将确定如何磷酸肌醇信号传导有助于α颗粒生物发生和功能。我们将测试巨核细胞离散微区内磷酸肌醇合成血小板调节效应蛋白，如NBEAL 2（负责Gray的突变蛋白），血小板综合征）。这种信号级联调节NBEAL 2介导膜粘附的能力。动力学和蛋白质运输。我们还将详细分析α颗粒的功能作用离体流变学和超显微镜研究、体内血栓形成研究和体内炎症研究。