ROLE OF STATHMIN IN MEGAKARYOPOIESIS AND PLATELET PRODUCTION

描述（由申请人提供）： 候选人的长期职业目标是了解巨核细胞生成和血小板生成的未定义机制，并应用这些见解来理解和治疗血液系统疾病。巨核细胞生成是一个复杂的过程，其中骨髓巨核细胞产生循环血小板。巨核细胞中微管细胞骨架具有独特的结构，对血小板的正常生成至关重要。本项目的总体目标是研究stathmin（一种主要的微管调节蛋白）在原代巨核细胞多倍体化（即获得超过4 N的DNA含量）和血小板生成过程中的作用。Stathmin在巨核细胞生成过程中生理性下调，并且在血小板中不存在。stathmin的表达水平与巨核细胞的多倍化呈负相关。抑制stathmin增加巨核细胞系的倍性，而其过表达降低倍性。基于此，我们的假设是，stathmin的表达下调是必要的多倍化和血小板的生产过程中正常的巨核细胞。因此，我们建议在转基因小鼠体内和体外研究这种干预对多倍化和血小板产生的影响，以防止在原代巨核细胞和血小板中stathmin的生理下调。首先，我们将研究慢病毒介导的stathmin过表达对多倍化的影响和这一过程的潜在机制（具体目标1）。第二，我们将产生在巨核细胞中有条件地过表达stathmin的转基因小鼠（具体目标2）。第三，我们将研究转基因巨核细胞中的巨核细胞生成和血小板形成（具体目标3）。这些研究应该能够更好地了解正常的巨核细胞生成的过程，并为今后研究stathmin和/或微管在异常巨核细胞生成和血小板生成中的潜在作用提供基础。他们还应该有可能验证微管作为血小板疾病治疗靶点的潜在用途。

DESCRIPTION (provided by applicant): The long-term career goal of the candidate is to understand the as yes undefined mechanisms of megakaryopoiesis and platelet production and to apply the insights to understand and possibly treat hematological disorders. Megakaryopoiesis is a complex process in which bone marrow megakaryocytes give rise to circulating platelets. Microtubule cytoskeleton has a unique organization in megakaryocytes which is essential for normal platelet production. The overall objective of this project is to investigate the role of stathmin, a major microtubule-regulatory protein, in the process of polyploidization (i.e. acquisition of more than 4N DNA content) and platelet production in primary megakaryocytes. Stathmin is physiologically downregulated during megakaryopoiesis and is absent in platelets. The levels of stathmin expression correlate inversely with megakaryocytes polyploidization. Inhibition of stathmin increases ploidy while its overexpression decreases ploidy of megakaryocytic cell lines. Based on this, our hypothesis is that downregulation of stathmin expression is necessary for polyploidization and platelet production during normal megakaryopoiesis. Thus, we propose to prevent physiological downregulation of stathmin in primary megakaryocytes and platelets ex vivo and in vivo in transgenic mice study the effects of this intervention on polyploidization and platelet production. First, we will investigate the effects of lentivirus-mediated overexpression of stathmin on polyploidization and the potential mechanisms underlying this process (Specific Aim 1). Second, we will generate transgenic mice that overexpress stathmin conditionally in megakaryocytes (Specific Aim 2). Third, we will investigate megakaryopoiesis and platelet formation in transgenic megakaryopcytes (Specific Aim 3). These studies should make it possible to better understand the process of normal megakaryopoiesis and provide the basis for the future investigation of the potential role of stathmin and/or microtubules in abnormal megakaryopoiesis and platelet production. They should also make possible to validate the potential use of microtubules as therapeutic targets in platelet disorders.