Runx1 Haplodeficiency, Endocytosis and Vesicle transport

Runx1 单倍体缺陷、内吞作用和囊泡运输

• 批准号: 10084304
• 负责人: Angara Koneti Rao
• 金额: $ 44.92万
• 依托单位: TEMPLE UNIV OF THE COMMONWEALTH
• 依托单位国家: 美国
• 财政年份: 2018
• 资助国家: 美国
• 起止时间: 2018-02-05 至 2024-01-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10084304
• 关键词: 12-HETE; AGFG1 gene; Abnormal Platelet; Acute leukemia; Albumins; Alpha Granule; Atherosclerosis; Biology; Blood Cells; Blood Coagulation Disorders; Blood Platelets; Blood coagulation; Cardiovascular Diseases; Cells; Cellular biology; Collaborations; Cytoplasmic Granules; Defect; Down-Regulation; Dynamin III; Endocytosis; Expression Profiling; Factor V; Fibrinogen; Foundations; Functional disorder; GATA1 gene; GFI1B gene; Genes; Genetic Transcription; Guanosine Triphosphate Phosphohydrolases; Hematopoietic; Hemorrhage; Hemostatic function; Human; Immunoglobulin G; Impairment; Inflammation; Inherited; Injury; Lead; Link; Mediating; Megakaryocytes; Megakaryocytopoieses; Messenger RNA; MicroRNAs; Modeling; Molecular; Molecular Biology; Mutation; Myocardial Infarction; Myosin Light Chains; P-Selectin; Pathway interactions; Patients; Phenotype; Phosphorylation; Platelet aggregation; Play; Predisposition; Process; Production; Protein Kinase C; Proteins; RUNX1 gene; Reporting; Role; Small Interfering RNA; Stroke; Technology; Thrombocytopenia; Thrombosis; Time; Vesicle; experience; human model; induced pluripotent stem cell; insight; interest; knock-down; novel therapeutic intervention; particle; pathogen; platelet function; platelet protein P47; protein transport; receptor; stem cells; trafficking; transcription factor; tumor; uptake; vesicle transport

Project Summary RUNX1 is a major hematopoietic transcription factor and RUNX1 haplodeficiency (RHD) is characterized by familial thrombocytopenia and impaired platelet function. We have a longstanding interest in the molecular basis of inherited platelet dysfunction, particularly related to RHD. Numerous platelet abnormalities have been described in RHD, several by us, and include deficiencies of dense granule (DG) and α-granules (AG). Our platelet expression profiling studies in a patient with RHD showed that several genes are down regulated (MYL9, PRKCQ, ALOX12, PF4, PLDN, PCTP); and we have shown that these are direct RUNX1 transcriptional targets and impact platelet/megakaryocyte (MK) biology. The overall objective of this proposal is to obtain new insights into the mechanisms of endocytosis, vesicle trafficking, and α-granule formation in platelets/MK through the study of phenotypic abnormalities and genes dysregulated in RHD. This project builds on unique abnormalities identified by us in RHD. We have reported in our patient that platelet albumin and IgG (incorporated by bulk endocytosis into AG) are decreased. Our expression profiling studies show that platelet RAB1B, RAB31 and DNM3 – three GTPases closely linked to vesicle trafficking are decreased. Little is currently known regarding the mechanisms regulating endocytosis in platelets/MK or the role of these GTPases. Our hypothesis is that mechanisms of endocytosis and vesicle trafficking are impaired in RHD. Aim 1 is to obtain insights into mechanisms regulating endocytosis in platelets/MK through the study of mechanisms leading to the decreased platelet albumin and IgG in RHD. We will study uptake and transport of albumin and IgG in normal and RHD platelets, study the effect of downregulation of RUNX1, RAB1B, RAB31 and DNM3 on these processes in MK, perform studies using markers of secretory and endocytic pathways of vesicle trafficking. Aim 2 is to understand the mechanisms leading to AG deficiency in RHD. We will study: platelet AG in normal and RHD platelets focusing on selected AG proteins; the effect of downregulation of RUNXI, RAB1B, RAB31and DNM3 on AG and trafficking of AG proteins in MK. These studies will be performed on 5-6 patients with RHD and using induced pluripotent stem cells (IPSCs) already developed from a RHD patient. They will provide important new insights into endocytosis and vesicle trafficking in platelets/MK, about which little is presently known. Relevance Platelets play a major role in hemostasis, thrombosis, inflammation, atherosclerosis and handling of pathogens. Our studies will provide new information on the basic aspects of platelet/MK function through studies in human RUNX1 haplodeficiency, a unique reservoir of information. This information will lay the foundation for new therapeutic approaches for both thrombotic and bleeding disorders.

项目摘要 RUNX1是一种主要的造血转录因子和RUNX1单倍体缺乏症(RHD) 以家族性血小板减少和血小板功能受损为特征。我们有一个 对遗传性血小板功能障碍的分子基础的长期兴趣，特别是 与风湿性心脏病有关。风湿性心脏病患者中有大量的血小板异常，有几个 包括致密颗粒(DG)和α颗粒(AG)的不足。我们的血小板 风湿性心脏病患者的表达谱研究显示有几个基因下调 受监管的(MYL9、PRKCQ、ALOX12、PF4、PLDN、PCTP)；我们已经证明这些 是RUNX1的直接转录靶点，影响血小板/巨核细胞(MK) 生物学。这项建议的总体目标是获得对 血小板/巨噬细胞吞噬、囊泡转运和α颗粒形成的机制 通过对风湿性心脏病表型异常和基因异常的研究。这 该项目建立在我们在RHD中发现的独特异常的基础上。我们已经在我们的 患者称血小板白蛋白和免疫球蛋白(通过大量内吞作用进入AG) 减少了。我们的表达谱研究表明，血小板RAB1B、RAB31和 DNM3-与囊泡运输密切相关的三种GTP酶减少。小才是 目前已知的关于调节血小板/MK的内吞作用的机制或 这些GTP酶的作用。我们的假设是，内吞作用和囊泡的机制 贩卖人口在RHD中受到损害。目标1是深入了解调控机制 通过研究内吞作用导致血小板/巨噬细胞集落刺激因子降低的机制 风湿性心脏病患者的血小板白蛋白和免疫球蛋白。我们将研究白蛋白和免疫球蛋白的摄取和转运。 在正常和RHD血小板中，研究RUNX1、RAB1B、RUNX1、RAB1B、 RAB31和DNM3在MK的这些过程上，使用标记进行研究 囊泡运输的分泌和内吞途径。目标2是了解 风湿性心脏病AG缺乏的机制。我们将研究：正常和正常的血小板AG RHD血小板聚焦于选定的AG蛋白；RUNXI下调的影响， RAB1B、RAB31和DNM3对AG的影响及AG蛋白在小鼠体内的转运这些研究 将在5-6名风湿性心脏病患者身上进行，并使用诱导的多能干细胞 (IPSCs)已经从一名风湿性心脏病患者发展而来。它们将提供重要的新见解 血小板/单核细胞的内吞作用和囊泡运输，目前对此知之甚少 为人所知。 相关性 血小板在止血、血栓形成、炎症、动脉粥样硬化和 病原体的处理。我们的研究将提供有关基本方面的新信息 一种独特的人类RUNX1单倍体缺陷症研究中的血小板/MK功能 信息库。这一信息将为新的治疗方法奠定基础 血栓性和出血性疾病的治疗方法。